Publications

Recent functional and anatomical discoveries of brain-immune relationships have overturned previous beliefs regarding the brain's immune privilege. Here, we propose that the brain and immune cells at its borders operate as an "ecosystem" to support the brain's robustness and resilience. Modulation of this ecosystem can be harnessed in the clinic.

For decades, neurodegenerative diseases were thought to be caused by the accumulation of toxic compounds, exacerbated by local inflammation, which together lead to neuronal loss and cognitive impairment . An additional factor that was long overlooked , is the role of the systemic immune system, which provides a defense mechanism against internal and external intruders in all bodily tissues. The evolving understanding of the life-long cross-talk between the CNS and the immune system led to an awareness of the function of systemic adaptive immunity in containing emerging destructive factors within the brain. . This includes harnessing of circulating myeloid cells to help the brain. However, as damage accumulates within the brain, the systemic immune system loses its protective capacity. Under such conditions, the dysregulated immune system becomes an escalating factor itself, thereby driving a vicious cycle that must be arrested.
•The brain's privilege does not imply immune isolation, but rather, a unique mode of brain–immune communication.•Both adaptive and innate immune support are needed for brain maintenance and repair.•Neurodegenerative diseases have an essential peripheral component, which is often manifested by immune dysfunction.•While not a primary cause, the immune dysfunction develops at an early stage in neurodegenerative disease, presumably before symptoms appear, resulting in a vicious cycle that further escalates disease progression.•Breaking the vicious cycle by activating the systemic immune system can harness bone marrow-derived macrophages and regulatory T cells to the brain to help defeat disease.

For decades, it was commonly accepted that the brain is secluded from peripheral immune activity and is self-sufficient for its maintenance and repair. This simplistic perception was based on the presence of resident immune cells, the microglia, and barrier systems within the brain, and the assumption that the central nervous system (CNS) lacks lymphatic drainage. This view was revised with the discoveries that higher functions of the CNS, homeostasis and repair are supported by peripheral innate and adaptive immune cells. The findings of bone marrow-derived immune cells in specialized niches, and the renewed observation that a lymphatic drainage system exists within the brain, further contributed to this revised model. In this Review, we describe the immune niches within the brain, the contribution of professional immune cells to brain functions, the bidirectional relationships between the CNS and the immune system and the relevance of immune components to brain aging and neurodegenerative diseases.

The role of non-neuronal cells in Alzheimer's disease progression has not been fully elucidated. Using single-nucleus RNA sequencing, we identified a population of disease-associated astrocytes in an Alzheimer's disease mouse model. These disease-associated astrocytes appeared at early disease stages and increased in abundance with disease progression. We discovered that similar astrocytes appeared in aged wild-type mice and in aging human brains, suggesting their linkage to genetic and age-related factors.

The immune system supports brain plasticity and homeostasis, yet it is prone to changes following psychological stress. Thus, it remains unclear whether and how stress-induced immune alterations contribute to the development of mental pathologies. Here, we show that following severe stress in mice, leukocyte trafficking through the choroid plexus (CP), a compartment that mediates physiological immune-brain communication, is impaired. Blocking glucocorticoid receptor signaling, either systemically or locally through its genetic knockdown at the CP, facilitated the recruitment of Gata3- and Foxp3-expressing T cells to the brain and attenuated post-traumatic behavioral deficits. These findings functionally link post-traumatic stress behavior with elevated stress-related corticosteroid signaling at the brain-immune interface and suggest a novel therapeutic target to attenuate the consequences of severe psychological stress.

Microglia differentiate from progenitors that infiltrate the nascent CNS during early embryonic development. They then remain in this unique immune-privileged environment throughout life. Multiple immune mechanisms, which we collectively refer to as microglial checkpoints, ensure efficient and tightly regulated microglial responses to perturbations in the CNS milieu. Such mechanisms are essential for proper CNS development and optimal physiological function. However, in chronic disease or aging, when a robust immune response is required, such checkpoint mechanisms may limit the ability of microglia to protect the CNS. Here we survey microglial checkpoint mechanisms and their roles in controlling microglial function throughout life and in disease, and discuss how they may be targeted therapeutically.

Hypoxia occurs physiologically in the developing body, and changing oxygen tensions are known to direct tissue differentiation; however, in the context of pathology, the same hypoxia-activated mechanisms may negatively affect tissue function. In this issue of The EMBO Journal, Esteban-Martinez et al (2017) report that programmed mitophagy, dependent on hypoxia-induced NIP-3-like protein X (BNIP3L, best known as NIX), is an essential step in differentiation of both retinal neurons and inflammatory macrophages.

For decades, the brain was considered to be a tissue behind barriers, and thus autonomous with respect to its maintenance needs. Several years ago, our group suggested that the ability of the brain to operate optimally, with relatively little deterioration throughout life, is dependent on continuous support from circulating immune cells. We envisioned that such immune support is provided from specific permissive sites that lie between the brain and the immune system, enabling immune-brain dialogue without the risk of an inflammatory autoimmune response. The brain's choroid plexus has been identified as an active neuroimmunological interface that is continuously exposed to signals from the brain and from the periphery. The interplay between these bidirectional signals shapes brain function in health and disease, including in multiple sclerosis. Accumulating evidence suggests that autoimmune T cells that recognize brain antigens are involved in this dialogue. Here, we discuss the fate of this dialogue under neurodegenerative conditions, and how boosting autoimmunity, rather than suppressing it, by either active vaccination with central nervous system-derived antigens, or by breaking peripheral immune suppression, could be harnessed for mitigating neurodegenerative diseases.

Hippocampal neurogenesis is affected throughout life by factors external to the brain, including what we eat, our gut microbiota, and the immune system. However, the mechanisms that link microbiota to neurogenesis are still puzzling. Now in Cell Reports, Mohle et al. (2016) attribute a role to Ly6C(hi) monocytes in this gut-immune-brain axis.

Hemorrhagic stroke, primarily caused by rupture of blood vessels in the brain, is a leading cause of death and disability in adults. In this issue of Immunity, Liu et al. (2016) demonstrate that repair of cerebrovascular ruptures can be directly mediated by myeloid cells.

Recent findings have revealed distinct roles for type I and II interferons (IFN-I and IFN-gamma) in the recruitment of immune cells to the central nervous system (CNS) and highlighted the importance of this process for brain maintenance and protection/repair. Furthermore, manipulation of IFN-I and IFN-gamma pathways in pathological contexts has yielded conflicting results. We discuss these findings, focusing on two distinct conditions; relapsing remitting multiple sclerosis (RRMS) and brain aging. Using these examples, we propose that regulation of immune cell entry to the CNS is a mechanism through which interaction between IFN-I and -II can affect brain function from its anatomical borders. Deviation from homeostatic IFN-I/-II balance may contribute to distinct brain pathologies, resulting from either insufficient immune surveillance of the CNS and loss of immune-dependent protection, or overwhelming leukocyte entry and immune-mediated destruction.

In the article “Passive or Active Immunization with Myelin Basic Protein Promotes Recovery from Spinal Cord Injury Contusion” by Ehud Hauben, Oleg Butovsky, Uri Nevo, Eti Yoles, Gila Moalem, Eugenia Agranov, Felix Mor, Raya Leibowitz-Amit, Evgenie Pevsner, Solange Akselrod, Michal Neeman, Irun R. Cohen, and Michal Schwartz, which appeared on pages 6421–6430 of the September 1, 2000 issue, the following error was discovered:In the legend of Figure 4A, a reference is missing. Hauben E, Nevo U, Yoles E, Moalem G, Agranov E, Mor F, Akselrod S, Neeman M, Cohen IR, Schwartz M (2000) Autoimmune T cells as potential neuroprotective therapy for spinal cord injury. Lancet 355:28–287, which is cited in the text of the article, should be cited in the legend as well. The sentence in the legend that reads “For comparison, a similar experiment using five PBS-treated and six rats treated immediately with anti-MBP T cells is shown here” should be replaced by “For comparison, a similar experiment, taken from Hauben et al. (2000), of six rats treated immediately with anti-MBP is shown.”

Chronic neuroinflammation is evident in brain aging and neurodegenerative disorders and is often associated with excessive nitric oxide (NO) production within the central nervous system (CNS). Under such conditions, increased NO levels are observed at the choroid plexus (CP), an epithelial layer that forms the blood-cerebrospinal fluid barrier (BCSFB) and serves as a selective gateway for leukocyte entry to the CNS in homeostasis and following injury. Here, we hypothesized that elevated cerebral NO levels interfere with CP gateway activity. We found that induction of leukocyte trafficking determinants by the CP and sequential leukocyte entry to the CSF are dependent on the CP epithelial NFκB/p65 signaling pathway, which was inhibited upon exposure to NO. Examining the CP in 5XFAD transgenic mouse model of Alzheimer's disease (AD-Tg) revealed impaired ability to mount an NFκB/p65-dependent response. Systemic administration of an NO scavenger in AD-Tg mice alleviated NFκB/p65 suppression at the CP and augmented its gateway activity. Together, our findings identify cerebral NO as a negative regulator of CP gateway activity for immune cell trafficking to the CNS.

Inadequate axonal regeneration is a common phenomenon occurring following acute injury to the central nervous system (CNS), and is often associated with permanent neurological deficits. The injured axons attempting to regenerate face the inhospitable environment of the CNS scar, which can hinder axonal growth and sprouting. In addition, in response to the insult, intense activation and infiltration of immune cells take place. Both the scar tissue and immune response, which have received a bad reputation in the context of CNS repair are essential for the overall recovery from CNS injuries, but are not optimally controlled. The glial scar contributes to protection of the spared neural tissues by establishing a boundary between damaged and salvageable tissue, and by educating the immune cells to promote the healing of the CNS tissue. In turn, the immune cells, and in particular the infiltrating macrophages, exert several functions at the lesion site, including resolution of the microglial response, control of scar tissue degradation, and production of growth factors; thereby, promoting neuronal survival, axonal regeneration, and tissue remodeling. As axonal regeneration and tissue remodeling are viewed as critical steps for the overall functional recovery following CNS injury, a detailed understanding of the mechanisms underlying the timely formation and degradation of the CNS scar, and its crosstalk with the inflammatory response, are of great importance, both biologically and clinically.

Keywords: Immunology; Neurosciences

Aging-associated cognitive decline is affected by factors produced inside and outside the brain. By using multiorgan genome-wide analysis of aged mice, we found that the choroid plexus, an interface between the brain and the circulation, shows a type I interferon (IFN-I)-dependent gene expression profile that was also found in aged human brains. In aged mice, this response was induced by brain-derived signals, present in the cerebrospinal fluid. Blocking IFN-I signaling within the aged brain partially restored cognitive function and hippocampal neurogenesis and reestablished IFN-II-dependent choroid plexus activity, which is lost in aging. Our data identify a chronic aging-induced IFN-I signature, often associated with antiviral response, at the brain's choroid plexus and demonstrate its negative influence on brain function, thereby suggesting a target for ameliorating cognitive decline in aging.

Keywords: Immunology; Neurosciences

Immune activity in the CNS parenchyma under various acute and chronic neurodegenerative conditions has been often interpreted as a sign of pathological inflammation. The apparent resemblance of the local neuroinflammatory processes to autoimmune diseases, such as multiple sclerosis (MS), generated the view that, despite differences in etiology and pathology, neurodegenerative disorders with a local inflammatory component can benefit from systemic anti-inflammatory therapy. In addition, as CNS self-reactive T cells are associated with the etiology of MS, autoimmunity was assumed to solely reflect pathology, and therefore, was universally linked to autoimmune disease. Yet, it is becoming increasingly clear that CNS-specific T cells, along with circulating and local innate immune cells, can enhance CNS healing processes following non-infectious injuries, or any deviation from homeostasis, including chronic pathological conditions. Here, we discuss the theory of "protective autoimmunity," which describes the activity of an immune cell network encompassing effector and regulatory T cells with specificity for CNS antigens, in CNS maintenance and repair. Such an immune network, evoked in response to external and internal threats, functions in a tightly regulated way, ensuring restoration of the brain's equilibrium and return to homeostasis.

For decades, several axioms have prevailed with respect to the relationships between the CNS and circulating immune cells. Specifically, immune cell entry was largely considered to be pathological or to mark the beginning of pathology within the brain. Moreover, local inflammation associated with neurodegenerative diseases such Alzheimer's disease or amyotrophic lateral sclerosis, were considered similar in their etiology to inflammatory diseases, such as remitting relapsing-multiple sclerosis. The ensuing confusion reflected a lack of awareness that the etiology of the disease as well as the origin of the immune cells determines the nature of the inflammatory response, and that inflammation resolution is an active cellular process. The last two decades have seen a revolution in these prevailing dogmas, with a significant contribution made by the authors. Microglia and infiltrating monocyte-derived macrophages are now known to be functionally distinct and of separate origin. Innate and adaptive immune cells are now known to have protective/healing properties in the CNS, as long as their activity is regulated, and their recruitment is well controlled; their role is appreciated in maintenance of brain plasticity in health, aging, and chronic neurodevelopmental and neurodegenerative diseases. Moreover, it is now understood that the barriers of the brain are not uniform in their interactions with the circulating immune cells. The implications of these new findings to the basic understanding of CNS repair processes, brain aging, and a wide spectrum of CNS disorders, including acute injuries, Rett syndrome, Alzheimer's disease, and multiple sclerosis, will be discussed.

Functional macrophage heterogeneity is recognized outside the central nervous system (CNS), where alternatively activated macrophages can perform immune-resolving functions. Such functional heterogeneity was largely ignored in the CNS, with respect to the resident microglia and the myeloid-derived cells recruited from the blood following injury or disease, previously defined as blood-derived microglia; both were indistinguishably perceived detrimental. Our studies have led us to view the myeloid-derived infiltrating cells as functionally distinct from the resident microglia, and accordingly, to name them monocyte-derived macrophages (mo-M Phi). Although microglia perform various maintenance and protective roles, under certain conditions when they can no longer provide protection, mo-M Phi are recruited to the damaged CNS; there, they act not as microglial replacements but rather assistant cells, providing activities that cannot be timely performed by the resident cells. Here, we focus on the functional heterogeneity of microglia/mo-M Phi, emphasizing that, as opposed to the mo-M Phi, microglia often fail to timely acquire the phenotype essential for CNS repair.

Complex barriers separate immune-privileged tissues from the circulation. Here, we propose that cell entry to immune-privileged sites through barriers composed of tight junction-interconnected endothelium is associated with destructive inflammation, whereas border structures comprised of fenestrated vasculature enveloped by tightly regulated epithelium serve as active and selective immune-skewing gates in the steady state. Based on emerging knowledge of the central nervous system and information from other immune-privileged sites, we propose that these sites are endowed either with absolute endothelial-based barriers and epithelial gates that enable selective and educative transfer of trafficking leukocytes or with selective epithelial gates only.

The poor recovery of the central nervous system (CNS) after injury, coupled with its complex and immunologically-privileged nature, led to the belief that CNS repair is different from the repair of other tissues. Here, we consider CNS repair from a novel perspective, suggesting that CNS responses to injury resemble wound healing. Extrapolating the classical wound healing model suggests that poor CNS recovery is an outcome of insufficient resolution of interim reparative events that precede tissue regeneration and renewal, a state reminiscent of chronic/unresolved wounds. This comparison requires reevaluation of the inflammatory response, glial scarring, and barrier permeability, traditionally considered obstacles to CNS repair. Understanding the similarity to wound healing suggests new research directions and therapeutic avenues for CNS injuries.

The adaptive arm of the immune system has been suggested as an important factor in brain function. However, given the fact that interactions of neurons or glial cells with T lymphocytes rarely occur within the healthy CNS parenchyma, the underlying mechanism is still a mystery. Here we found that at the interface between the brain and blood circulation, the epithelial layers of the choroid plexus (CP) are constitutively populated with CD4(+) effector memory cells with a T-cell receptor repertoire specific to CNS antigens. With age, whereas CNS specificity in this compartment was largely maintained, the cytokine balance shifted in favor of the T helper type 2 (Th2) response; the Th2-derived cytokine IL-4 was elevated in the CP of old mice, relative to IFN-gamma, which decreased. We found this local cytokine shift to critically affect the CP epithelium, triggering it to produce the chemokine CCL11 shown to be associated with cognitive dysfunction. Partial restoration of cognitive ability in aged mice, by lymphopenia-induced homeostasis-driven proliferation of memory T cells, was correlated with restoration of the IL-4:IFN-gamma ratio at the CP and modulated the expression of plasticity-related genes at the hippocampus. Our data indicate that the cytokine milieu at the CP epithelium is affected by peripheral immunosenescence, with detrimental consequences to the aged brain. Amenable to immunomodulation, this interface is a unique target for arresting age-related cognitive decline.

Reduction in T cell receptor (TCR) diversity in old age is considered as a major cause for immune complications in the elderly population. Here, we explored the consequences of aging on the TCR repertoire in mice using high-throughput sequencing (TCR-seq). We mapped the TCR beta repertoire of CD4+ T cells isolated from bone marrow (BM) and spleen of young and old mice. We found that TCR beta diversity is reduced in spleens of aged mice but not in their BM. Splenic CD4+ T cells were also skewed toward an effector memory phenotype in old mice, while BM cells preserved their memory phenotype with age. Analysis of V beta and J beta gene usage across samples, as well as comparison of CDR3 length distributions, showed no significant age dependent changes. However, comparison of the frequencies of amino-acid (AA) TCR beta sequences between samples revealed repertoire changes that occurred at a more refined scale. The BM-derived TCR beta repertoire was found to be similar among individual mice regardless of their age. In contrast, the splenic repertoire of old mice was not similar to those of young mice, but showed an increased similarity with the BM repertoire. Each old-mouse had a private set of expanded TCR beta sequences. Interestingly, a fraction of these sequences was found also in the BM of the same individual, sharing the same nucleotide sequence. Together, these findings show that the composition and phenotype of the CD4+ T cell BM repertoire are relatively stable with age, while diversity of the splenic repertoire is severely reduced. This reduction is caused by idiosyncratic expansions of tens to hundreds of T cell clonotypes, which dominate the repertoire of each individual. We suggest that these private and abundant clonotypes are generated by sporadic clonal expansions, some of which correspond to pre-existing BM clonotypes. These organ- and age-specific changes of the TCR beta repertoire have implications for understanding and manipulating age-associated immune decline.

Understanding of ocular diseases and the search for their cure have been based on the common assumption that the eye is an immune privileged site, and the consequent conclusion that entry of immune cells to this organ is forbidden. Accordingly, it was assumed that when immune cell entry does occur, this reflects an undesired outcome of breached barriers. However, studies spanning more than a decade have demonstrated that acute insults to the retina, or chronic conditions resulting in retinal ganglion cell loss, such as in glaucoma, result in an inferior outcome in immunocompromised mice; likewise, steroidal treatment was found to be detrimental under these conditions. Moreover, even conditions that are associated with inflammation, such as age-related macular degeneration, are not currently believed to require immune suppression for treatment, but rather, are thought to benefit from immune modulation. Here, we propose that the immune privilege of the eye is its ability to enable, upon need, the entry of selected immune cells for its repair and healing, rather than to altogether prevent immune cell entry. The implications for acute and chronic degenerative diseases, as well as for infection and inflammatory diseases, are discussed.

The brain has been commonly regarded as a "tissue behind walls." Appearance of immune cells in the brain has been taken as a sign of pathology. Moreover, since infiltrating monocyte-derived macrophages and activated resident microglia were indistinguishable by conventional means, both populations were considered together as inflammatory cells that should be mitigated. Yet, because the microglia permanently reside in the brain, attributing to them negative properties evoked an ongoing debate; why cells that are supposed to be the brain guardians acquire only destructive potential? Studies over the last two decades in the immune arena in general, and in the context of central nervous system pathology in particular, have resulted in a paradigm shift toward a more balanced appreciation of the contributions of immune cells in the context of brain maintenance and repair, and toward the recognition of distinct roles of resident microglia and infiltrating monocyte-derived macrophages.

Background: Circulating immune cells including autoreactive T cells and monocytes have been documented as key players in maintaining, protecting and repairing the central nervous system (CNS) in health and disease. Here, we hypothesized that neurodegenerative diseases might be associated, similarly to tumors, with increased levels of circulating peripheral myeloid derived suppressor cells (MDSCs), representing a subset of suppressor cells that often expand under pathological conditions and inhibit possible recruitment of helper T cells needed for fighting off the disease. Methods and Findings: We tested this working hypothesis in amyotrophic lateral sclerosis (ALS) and its mouse model, which are characterized by a rapid progression once clinical symptoms are evident. Adaptive transfer of alternatively activated myeloid (M2) cells, which homed to the spleen and exhibited immune suppressive activity in G93A mutant superoxide dismutase-1 (mSOD1) mice at a stage before emergence of disease symptoms, resulted in earlier appearance of disease symptoms and shorter life expectancy. The same protocol mitigated the inflammation-induced disease model of multiple sclerosis, experimental autoimmune encephalomyelitis (EAE), which requires circulating T cells for disease induction. Analysis of whole peripheral blood samples obtained from 28 patients suffering from sporadic ALS (sALS), revealed a two-fold increase in the percentage of circulating MDSCs (LIN(-/Low)HLA-DR(-)CD33(+)) compared to controls. Conclusions: Taken together, these results emphasize the distinct requirements for fighting the inflammatory neurodegenerative disease, multiple sclerosis, and the neurodegenerative disease, ALS, though both share a local inflammatory component. Moreover, the increased levels of circulating MDSCs in ALS patients indicates the operation of systemic mechanisms that might lead to an impairment of T cell reactivity needed to overcome the disease conditions within the CNS. This high le

Amyotrophic lateral sclerosis (ALS) is a rapidly progressing fatal neurodegenerative disorder characterized by the selective death of motor neurons (MN) in the spinal cord, and is associated with local neuroinflammation. Circulating CD4(+) T cells are required for controlling the local detrimental inflammation in neurodegenerative diseases, and for supporting neuronal survival, including that of MN. T-cell deficiency increases neuronal loss, while boosting T cell levels reduces it. Here, we show that in the mutant superoxide dismutase 1 G93A (mSOD1) mouse model of ALS, the levels of natural killer T (NKT) cells increased dramatically, and T-cell distribution was altered both in lymphoid organs and in the spinal cord relative to wildtype mice. The most significant elevation of NKT cells was observed in the liver, concomitant with organ atrophy. Hepatic expression levels of insulin-like growth factor (IGF)-1 decreased, while the expression of IGF binding protein (IGFBP)-1 was augmented by more than 20-fold in mSOD1 mice relative to wild-type animals. Moreover, hepatic lymphocytes of pre-symptomatic mSOD1 mice were found to secrete significantly higher levels of cytokines when stimulated with an NKT ligand, ex-vivo. Immunomodulation of NKT cells using an analogue of alpha-galactosyl ceramide (alpha-GalCer), in a specific regimen, diminished the number of these cells in the periphery, and induced recruitment of T cells into the affected spinal cord, leading to a modest but significant prolongation of life span of mSOD1 mice. These results identify NKT cells as potential players in ALS, and the liver as an additional site of major pathology in this disease, thereby emphasizing that ALS is not only a non-cell autonomous, but a non-tissue autonomous disease, as well. Moreover, the results suggest potential new therapeutic targets such as the liver for immunomodulatory intervention for modifying the disease, in addition to MN-based neuroprotection and systemic treatments

Noninvasive monitoring of beta-amyloid (A beta) plaques, the neuropathological hallmarks of Alzheimer's disease (AD), is critical for AD diagnosis and prognosis. Current visualization of A beta plaques in brains of live patients and animal models is limited in specificity and resolution. The retina as an extension of the brain presents an appealing target for a live, noninvasive optical imaging of AD if disease pathology is manifested there. We identified retinal A beta plaques in postmortem eyes from AD patients (n = 8) and in suspected early stage cases (n=5), consistent with brain pathology and clinical reports; plaques were undetectable in age-matched non-AD individuals (n = 5). In APP(SWE)/PS1(Delta E9) transgenic mice (AD-Tg; n = 18) but not in non-Tg wt mice (n = 10), retinal A beta plaques were detected following systemic administration of curcumin, a safe plaque-labeling fluorochrome. Moreover, retinal plaques were detectable earlier than in the brain and accumulated with disease progression. An immune-based therapy effective in reducing brain plaques, significantly reduced retinal A beta plaque burden in immunized versus non-immunized AD mice (n = 4 mice per group). In live AD-Tg mice (n = 24), systemic administration of curcumin allowed noninvasive optical imaging of retinal A beta plaques in vivo with high resolution and specificity; plaques were undetectable in non-Tg wt mice (n = 11). Our discovery of A beta specific plaques in retinas from AD patients, and the ability to noninvasively detect individual retinal plaques in live AD mice establish the basis for developing high-resolution optical imaging for early AD diagnosis, prognosis assessment and response to therapies. (C) 2010 Elsevier Inc. All rights reserved.

Amyotrophic lateral sclerosis (ALS) is a devastating disease, characterized by extremely rapid loss of motor neurons. Our studies over the last decade have established CD4+ T cells as important players in central nervous system maintenance and repair. Those results, together with recent findings that CD4+ T cells play a protective role in mouse models of ALS, led us to the current hypothesis that in ALS, a rapid T-cell malfunction may develop in parallel to the motor neuron dysfunction. Here, we tested this hypothesis by assessing thymic function, which serves as a measure of peripheral T-cell availability, in an animal model of ALS (mSOD1 [superoxide dismutase] mice; G93A) and in human patients. We found a significant reduction in thymic progenitor-cell content, and abnormal thymic histology in 3-4-month-old mSOD1 mice. In ALS patients, we found a decline in thymic output, manifested in the reduction in blood levels of T-cell receptor rearrangement excision circles, a non-invasive measure of thymic function, and demonstrated a restricted T-cell repertoire. The morbidity of the peripheral immune cells was also manifested in the increase of pro-apoptotic BAX/BCXL2 expression ratio in peripheral blood mononuclear cells (PBMCs) of these patients. In addition, gene expression screening in the same PBMCs, revealed in the ALS patients a reduction in key genes known to be associated with T-cell activity, including: CD80, CD86, IFNG and IL18. In light of the reported beneficial role of T cells in animal models of ALS, the present observation of thymic dysfunction, both in human patients and in an animal model, might be a co-pathological factor in ALS, regardless of the disease aetiology. These findings may lead to the development of novel therapeutic approaches directed at overcoming the thymic defect and T-cell deficiency.

Circulating immune cells support hippocampal neurogenesis, spatial memory, expression of brain-derived neurotrophic factor, and resilience to stress. Nevertheless, considering the immune privileged status of the central nervous system (CNS), such cells were assumed to be excluded from the healthy brain. It is evident, however, that the CNS is continuously surveyed by leukocytes, though their function is still a mystery. Coupling this routine leukocyte trafficking with the function attributed to circulating T cells in brain plasticity led us to propose here that CNS immunosurveillance is an integral part of the functioning brain. Anatomical restriction of selected self-recognizing leukocytes to the brain's borders and fluids (cerebrospinal fluid) not only supports the brain's activity, but also controls the potential aggressiveness of such cells. Accordingly, the brain's 'privilege' is its acquisition of a private peripheral immunological niche under its own control, which supports brain function. Immune malfunction may comprise a missing link between a healthy and diseased mind. Molecular Psychiatry (2010) 15, 342-354; doi: 10.1038/mp.2010.31

The factors that determine brain aging remain a mystery. Do brain aging and memory loss reflect processes occurring only within the brain? Here, we present a novel view, linking aging of adaptive immunity to brain senescence and specifically to spatial memory deterioration. Inborn immune deficiency, in addition to sudden imposition of immune malfunction in young animals, results in cognitive impairment. As a corollary, immune restoration at adulthood or in the elderly results in a reversal of memory loss. These results, together with the known deterioration of adaptive immunity in the elderly, suggest that memory loss does not solely reflect chronological age; rather, it is an outcome of the gap between an increasing demand for maintenance (age-related risk-factor accumulation) and the reduced ability of the immune system to meet these needs.

The psychobiological mechanisms that contribute to the development of stress resilience are not fully elucidated. One potential approach for enhancing resilience is the exposure to mild challenges. According to this approach, a mildly stressful episode may immunize the individual, thereby strengthening resistance to subsequent stressors. This phenomenon is often viewed as a form of behavioral immunization. Although, the term 'behavioral immunization' was borrowed from the field of immunology, the involvement of the adaptive immune system in stress resilience was never investigated. However, based on accumulated new data, we suggest that the immunological memory does have a significant role in developing coping responses to stress. Although, immune deficiency results in an impaired ability to cope with stress, boosting immunological memory can increase stress resilience. Therefore, we propose that defense against mental challenge, similarly to defense against intruders, involves an immunological mechanism, which establishes stress resilience to a later challenge. Here, we review the involvement of the adaptive immune system in coping mechanisms in response to psychological stress, and discuss the connection between cognitive memory and immunological memory in establishing ability to efficiently cope with stressful episodes.

Our research group has been working for more than a decade on the cross-talk between the immune and the nervous systems. Due to the unique nature of the central nervous system (CNS) as an immune privileged site, it was commonly believed that the nervous system functions optimally without any immune intervention, and that any immune cell infiltration to the CNS is a sign of pathology. However, since the immune system constitutes the body's major defense and repair mechanism, it seemed unreasonable that the CNS would have completely lost the need for assistance from this system. This insight prompted us to revisit the entire question of whether immune cells are required for recovery from CNS injuries. We subsequently made numerous fundamental observations that led us to formulate a unified theory linking all neurodegenerative conditions; thus, we suggested that "T-cell immunity to self maintains the self," at least in the CNS. According to this view, immunity to self ("protective autoimmunity") provides a pivotal role in maintenance, protection, and repair of the healthy and diseased CNS. We further showed that the T cells mediate their effect, at least under pathological conditions, by controlling the recruitment of blood-borne monocytes, which play a crucial local role that cannot be replaced by the resident microglia. Boosting of such a T cell response specific for brain proteins, while carefully choosing the antigen, the carrier, timing, dosing, and regimen should be considered as a way of augmenting a physiological repair mechanism needed to ameliorate disease conditions while restoring equilibrium needed for protection, repair and renewal; such therapy is not intended to modify a single mediator of a single disease, but rather, would serve as an approach for adjusting the levels of the immune response needed to restore a lost balance. (C) 2009 IBRO. Published by Elsevier Ltd. All rights reserved.

Retinal neurogenesis ceases by the early postnatal period, although retinal progenitor cells (RPCs) persist throughout life. In this study, we show that in the mammalian eye, the function of Toll-like receptor 4 (TLR4) extends beyond regulation of the innate immune response; it restricts RPC proliferation. In TLR4-deficient mice, enhanced proliferation of cells reminiscent of RPCs is evident during the early postnatal period. In vitro experiments demonstrate that TLR4 acts as an intrinsic regulator of RPC fate decision. Increased TLR4 expression in the eye correlates with the postnatal cessation of cell proliferation. However, deficient TLR4 expression is not sufficient to extend the proliferative period but rather contributes to resumption of proliferation in combination with growth factors. Proliferation in vivo is inhibited by both MyD88-dependent and -independent pathways, similar to the mechanisms activated by TLR4 in immune cells. Thus, our study attributes a novel role to TLR4 as a negative regulator of RPC proliferation.

Aging is often associated with a decline in hippocampus-dependent spatial memory. Here, we show that functional cell-mediated immunity is required for the maintenance of hippocampus-dependent spatial memory. Sudden imposition of immune compromise in young mice caused spatial memory impairment, whereas immune reconstitution reversed memory deficit in immune-deficient mice. Analysis of hippocampal gene expression suggested that immune-dependent spatial memory performance was associated with the expression of insulin-like growth factor (Igf1) and of genes encoding proteins related to presynaptic activity (Syt10, Cplx2). We further showed that memory loss in aged mice could be attributed to age-related attenuation of the immune response and could be reversed by immune system activation. Homeostatic-driven proliferation of lymphocytes, which expands the existing T cell repertoire, restored spatial memory deficits in aged mice. Thus, our results identify a novel function of the immune system in the maintenance of spatial memory and suggest an original approach for arresting or reversing age-associated memory loss.

Background Chondroitin sulfate proteoglycan (CSPG) is a major component of the glial scar. It is considered to be a major obstacle for central nervous system (CNS) recovery after injury, especially in light of its well-known activity in limiting axonal growth. Therefore, its degradation has become a key therapeutic goal in the field of CNS regeneration. Yet, the abundant de novo synthesis of CSPG in response to CNS injury is puzzling. This apparent dichotomy led us to hypothesize that CSPG plays a beneficial role in the repair process, which might have been previously overlooked because of nonoptimal regulation of its levels. This hypothesis is tested in the present study. Methods and Findings We inflicted spinal cord injury in adult mice and examined the effects of CSPG on the recovery process. We used xyloside to inhibit CSPG formation at different time points after the injury and analyzed the phenotype acquired by the microglia/macrophages in the lesion site. To distinguish between the resident microglia and infiltrating monocytes, we used chimeric mice whose bone marrow-derived myeloid cells expressed GFP. We found that CSPG plays a key role during the acute recovery stage after spinal cord injury in mice. Inhibition of CSPG synthesis immediately after injury impaired functional motor recovery and increased tissue loss. Using the chimeric mice we found that the immediate inhibition of CSPG production caused a dramatic effect on the spatial organization of the infiltrating myeloid cells around the lesion site, decreased insulin-like growth factor 1 (IGF-1) production by microglia/macrophages, and increased tumor necrosis factor alpha (TNF-alpha) levels. In contrast, delayed inhibition, allowing CSPG synthesis during the first 2 d following injury, with subsequent inhibition, improved recovery. Using in vitro studies, we showed that CSPG directly activated microglia/macrophages via the CD44 receptor and modulated neurotrophic factor secretion by these cells. Co

Neurogenesis, the formation of new neurons from stem/progenitor cells, occurs in the hippocampal dentate gyrus throughout life. Although the exact function of adult hippocampal neurogenesis is currently unknown, recent studies suggest that the newly formed neuronal population plays an important role in hippocampal-dependent cognitive abilities, including declarative memory. The process of adult neurogenesis is greatly influenced by the interaction between cells of the adaptive immune system and CNS-resident immune cells. Our laboratory has recently demonstrated that immune cells contribute to maintaining life-long hippocampal neurogenesis. The regulation of such immune-cell activity is crucial: too little immune activity (as in immune deficiency syndromes) or too much immune activity (as in severe inflammatory diseases) can lead to impaired hippocampal neurogenesis, which could then result in impaired hippocampal-dependent cognitive abilities. From these converging discoveries arise a mechanism that can explain one route by which our body affects our mind. (C) 2007 Elsevier Inc. All rights reserved.

Purpose: Drusen formation in age-related macular degeneration (AMD) shares some similarities with Alzheimer's disease (AD), which is associated with amyloid deposits. Aggregated beta-amyloid induces microglia to become cytotoxic and block neurogenesis. Recent evidence showed that T cell-based vaccination with Copaxone in AD mice model resulted in modulation of microglia into neuroprotective phenotype and as a result in reduction of cognitive decline, elimination of plaque formation, and induction of neuronal survival and neurogenesis. The aim was to investigate whether the effect of Copaxone on drusen in dry AMD is similar to that on deposits of other age-related chronic neurodegenerative diseases such as Alzheimer disease (AD). Materials and Methods: Patients over 50 years of age with intermediate dry AMD in both eyes were randomized to receive Copaxone or sham injections and were weekly treated by subcutaneous injections of Copaxone (dose of 20 mg) or sham injections for 12 weeks. At baseline, 6-week, and 12-week visits, visual acuity, contrast sensitivity, fundus examination and photography, fluorescein angiography, and ocular coherent tomography were performed. Main outcome measure was a change in total drusen area (TDA) measured by Image-Pro software and presented in arbitrary units (AU). Results: Eight studied eyes of four treated patients showed a decrease in TDA from 48130 to 16205 AU at 12 weeks as compared to baseline. In contrast, four control eyes (two patients) demonstrated almost no change in TDA (from 32294 to 32781 AU). Conclusion: These preliminary results show that Copaxone reduces drusen area.

Neural stem/progenitor cells are known to exist in the intact spinal cord, but the presence of newly formed neurons during adulthood has not been documented there to date. Here, we report the appearance of newly formed neurons under normal physiological conditions. These neurons are immature, express a GABAergic phenotype' and are primarily located in the dorsal part of the spinal cord. This localization appeared to be mediated by stromal-derived factor- 1/CXC-chemokine receptor-4 signaling in the dorsal region. The extent of spinal cord neurogenesis was found to be greatly influenced by immune system integrity and in particular by myelin-specific T cells. These observations provide evidence for in vivo spinal cord neurogenesis under nonpathological conditions and introduce novel mechanisms regulating adult spinal cord plasticity.

We have recently shown that the ability of microglia to effectively fight off aggregated beta-amyloid plaque formation and cognitive loss in transgenic mouse models of Alzheimer's disease (Tg-AD), is augmented in response to T-cell-based immunization, using glatiramer acetate (GA). The immunization increases incidence of local CD11c(+) dendritic-like cells. It is unclear, however, whether these dendritic cells are derived from resident microglia or from the bone marrow. To determine the origin of this dendritic-cell population, we used chimeric mice whose bone marrow-derived cells express a transgene that allows the cells to be specifically ablated by diphtheria toxin. We show here that T-cell-based immunization of these mice, using GA, induced the recruitment of bone marrow-derived dendritic cells. Depletion of the dendritic cells by systemic injection of diphtheria toxin resulted in significantly increased formation of amyloid plaques. Thus, recruitment of bone marrow-derived dendritic cells evidently plays a role in reducing plaque formation in a mouse model of Alzheimer's disease.

Glaucomatous neuropathy, like other neurodegenerative diseases, is a multi-dimensional disease in which various molecular and cellular factors contribute to the pathological process. These factors, while not initially causative, are key elements in disease progression and may continue to contribute even after the primary pathology is alleviated. An entire field of research of neuroprotection and restoration has opened up as part of the search for additional ways of slowing disease progression. We have proposed, on the basis of experimental evidence, that a vaccine could be a means of recruiting the immune system to help eliminate many of the factors associated with glaucomatous neurodegeneration and thus prevent disease progression, though not its onset. This immune defence involves lymphocytes, resident and infiltrating innate immune cells, the microglia, and macrophages. The antigens of choice are synthetic antigens, such as glatiramer acetate, that weakly cross-react with self-antigens in the retina and optic nerves. The vaccine induces a beneficial immune response that recruits immune effector cells to counteract or neutralize many of the compounds and factors that contribute to ongoing destruction, and in addition supports cell renewal and repair.

The ability of the central nervous system to cope with stressful conditions was shown to be dependent on proper T-cell-mediated immune response. Because the therapeutic window for neuroprotection after acute insults such as stroke is relatively narrow, we searched for a procedure that would allow the relevant T cells to be recruited rapidly. Permanent middle cerebral artery occlusion was induced in adult rats. To facilitate a rapid poststroke T cell activity, rats were treated with poly-YE using different regimens. Control and poly-YE-treated rats were assessed for functional recovery using neurological severity score and Morris water maze. Neuroprotection, neurogenesis, growth factor expression, and microglial phenotype were assessed using histological and immunofluorescence methods. Administration of poly-YE as late as 24 hours after middle cerebral artery occlusion yielded a beneficial effect manifested by better neurological performance, reduced neuronal loss, attenuation of behavioral deficits, and increased hippocampal and cortical neurogenesis. This compound affected the subacute phase by modulating microglial response and by increasing local production of insulin-like growth factor-I, known to be a key player in neuronal survival and neurogenesis. The relative wide therapeutic window, coupled with its efficacy in attenuating further degeneration and enhancing restoration, makes poly-YE a promising immune-based candidate for stroke therapy. (Stroke. 2007;38[part 2]:774-782.)

Neurogenesis is known to continuously take place in certain ‘neurogenic’ areas of the adult central nervous system (CNS) and can be induced in ‘non-neurogenic’ areas under traumatic or degenerative conditions. Here we introduce T cells and CNS-resident microglia as important players in the regulation of adult neurogenesis. Under normal conditions, immune deficient mice (SCID and nude) and transgenic mice that most of their T-cell pool is specific for an irrelevant antigen (ovalbumin) exhibited impaired hippocampal neurogenesis. In contrast, mice in which the majority of T cells specifically recognize the CNSabundant antigen myelin basic protein showed normal neurogenesis. CNSspecific T cells were also found to be important for spatial learning abilities and for brain-derived neurotrophic factor expression in the dentate gyrus. Environmental enrichment did not evoke enhanced neurogenesis in immune-deficient animals, whereas in wild-type animals it led to enhanced hippocampal neurogenesis coupled with recruitment of T cells and activation of microglia. The clinical implications of these findings were tested using a rat model of cerebral ischemic insult. We demonstrate that Tcell based immune activation following stroke induces a robust elevation of neurogenesis in the hippocampus as well as in the cerebral cortex. Our results suggest that T cells, acting via resident antigen presenting cells, are important regulators of adult neurogenesis under both physiological and pathological conditions.

The well regulated activities of microglia and T cells specific to central nervous system (CNS) antigens can contribute to the protection of CNS neural cells and their renewal from adult neural stem/progenitor cells (aNPCs). Here we report that T cell-based vaccination of mice with a myelin-derived peptide, when combined with transplantation of aNPCs into the cerebrospinal fluid (CSF), synergistically promoted functional recovery after spinal cord injury. The synergistic effect was correlated with modulation of the nature and intensity of the local T cell and microglial response, expression of brain-derived neurotrophic factor and noggin protein, and appearance of newly formed neurons from endogenous precursor-cell pools. These results substantiate the contention that the local immune response plays a crucial role in recruitment of aNPCs to the lesion site, and suggest that similar immunological manipulations might also serve as a therapeutic means for controlled migration of stem/progenitor cells to other acutely injured CNS sites.

The ability to cope with ongoing neurodegeneration after injury to the central nervous system of mammals differs among strains and depends in part on the animal's ability to manifest a T-cell-mediated protective response. After CNS injury, strain-related differences were observed. Moreover, the post-injury effect of naturally occurring regulatory CD4+CD25+ T cells was found to differ in different strains. In this study, using partially injured optic nerves of Balb/c/OLA and C57BL/6J mice as models, we observed strain-related differences in the T-cell-mediated protection obtained by antigens administered via the nasal route. Active immunization with myelin-related antigens emulsified in complete Freund's adjuvant had a beneficial effect on both strains, whereas mucosal administration of the same antigens was destructive in mice of the Balb/c/OLA strain but protective in C57BL/6J mice. (c) 2006 Elsevier B.V. All rights reserved.

The role of activated microglia (MG) in demyelinating neurodegenerative diseases such as multiple sclerosis is controversial. Here we show that high, but not low, levels of IFN-gamma (a cytokine associated with inflammatory autoimmune diseases) conferred on rodent MG a phenotype that impeded oligodendrogenesis from adult neural stem/progenitor cells. IL-4 reversed the impediment, attenuated TNF-alpha. production, and overcame blockage of IGF-I production caused by IFN-gamma. In rodents with acute or chronic EAE, injection of IL-4-activated MG into the cerebrospinal fluid resulted in increased oligodendrogenesis in the spinal cord and improved clinical symptoms. The newly formed oligodendrocytes were spatially associated with MG expressing MHC class II proteins and IGF-I. These results point to what we believe to be a novel role for MG in oligodendrogenesis from the endogenous stem cell pool.

Accumulating evidence suggests that neurodegenerative diseases of the central nervous system (CNS) are associated with a local inflammatory response. CNS autoimmune diseases are also associated with inflammation. Does this mean that all neurodegenerative diseases are autoimmune in nature? Does it imply that autoimmune and neurodegenerative diseases are both eligible for the same therapy? What distinguishes between the two types of disease? Do they differ both in etiology and in pathology, or do they have different etiologies but similar pathology and progression? In this minireview we offer a new view of the inflammatory differences between neurodegenerative and autoimmune diseases in the CNS and discuss the implications for therapy.

Neurogenesis is known to take place in the adult brain. This work identifies T lymphocytes and microglia as being important to the maintenance of hippocampal neurogenesis and spatial learning abilities in adulthood. Hippocampal neurogenesis induced by an enriched environment was associated with the recruitment of T cells and the activation of microglia. In immune-deficient mice, hippocampal neurogenesis was markedly impaired and could not be enhanced by environmental enrichment, but was restored and boosted by T cells recognizing a specific CNS antigen. CNS-specific T cells were also found to be required for spatial learning and memory and for the expression of brain-derived neurotrophic factor in the dentate gyrus, implying that a common immune-associated mechanism underlies different aspects of hippocampal plasticity and cell renewal in the adult brain.

Chondroitin sulfate proteoglycan (CSPG), a matrix protein that occurs naturally in the central nervous system (CNS), is considered to be a major inhibitor of axonal regeneration and is known to participate in activation of the inflammatory response. The degradation of CSPG by a specific enzyme, chondroitinase ABC, promotes repair. We postulated that a disaccharidic degradation product of this glycoprotein (CSPG-DS), generated following such degradation, participates in the modulation of the inflammatory responses and can, therefore, promote recovery in immune-induced neuropathologies of the CNS, such as experimental autoimmune encephalomyelitis (EAE) and experimental autoimmune uveitis (EAU). In these pathologies, the dramatic increase in T cells infiltrating the CNS is far in excess of the numbers needed for regular maintenance. Here, we show that CSPG-DS markedly alleviated the clinical symptoms of EAE and protected against the neuronal loss in EAU. The last effect was associated with a reduction in the numbers of infiltrating T cells and marked microglia activation. This is further supported by our in vitro results indicating that CSPG-DS attenuated T cell motility and decreased secretion of the cytokines interferon-gamma and tumor necrosis factor-alpha. Mechanistically, these effects are associated with an increase in SOCS-3 levels and a decrease in NF-kappa B. Our results point to a potential therapeutic modality, in which a compound derived from an endogenous CNS-resident molecule, known for its destructive role in CNS recovery, might be helpful in overcoming inflammation-induced neurodegenerative conditions.

With the increasing use of the rat as an animal model for glaucoma and for the evaluation of neuroprotective treatments, there is a need for a sensitive test of retinal ganglion cell (RGC) function in this species. The aims of this study were to detect functional abnormalities of the inner retina in a rat model of high intraocular pressure (IOP) using the pattern electroretinogram (PERG), and to correlate them with morphometric analysis of RGC survival and the functional integrity of the inner retina. Unilateral ocular hypertension was induced in 17 Lewis rats through laser photocoagulation. Pattern ERGs were recorded prior to lasering and 3 weeks later, using a series of shifting patterns of decreasing spatial frequency projected directly onto the animals' fundus. IOP was measured at the same intervals, and the number of surviving RGCs estimated. Low amplitude PERG signals could be recorded in response to a narrow grating of 0.368 cycles per degree (cpd), and increased with stimulus size. Lasering caused mean (+/- S.D.) IOP to increase significantly from 18.3 +/- 4.5 (baseline) to 29.8 8.8 mmHg within 3 weeks (p

Object. A Phase I, open-label nonrandomized study was conducted to assess the safety and tolerability of incubated autologous macrophages administered to patients with acute complete spinal cord injury (SCI). Methods. This therapy was first tested in rat models of spinal cord transection and contusion, in which it was shown to promote motor recovery. The procedure developed for clinical use consists of isolating monocytes from patient blood and incubating them ex vivo with autologous dermis. The resulting incubated autologous macrophages were injected into the patient's spinal cord immediately caudal to the lesion within 14 days of injury. Patients underwent preoperative and follow-up neurological assessment (American Spinal Injury Association [ASIA] standards), electrophysiological monitoring (motor evoked and/or somatosensory evoked potentials), magnetic resonance imaging, and safety monitoring. Before macrophage administration, complete neurological functional loss (ASIA Grade A) was confirmed in all patients. Of the eight patients in the study, three recovered clinically significant neurological motor and sensory function (ASIA Grade C status). During the study period, some adverse events were encountered, the most serious of which involved two cases of pulmonary embolism and one case of osteomyelitis that were treated and resolved without further complication. These and other adverse events appear to be similar to those encountered in other spinal cord-injured patients and are not considered a consequence of the experimental therapy. Conclusions. It is concluded that incubated autologous macrophage cell therapy is well tolerated in patients with acute SCI. Further clinical evaluation is warranted.

Protective e autoimmunity refers to a well-controlled anti-self response that helps the body resist neurodegeneration. The response is mediated by autoimmune T cells, which produce cytokines and growth factors. Using an in vitro assay of hippocampal slices, we show that the cytokines interferon-gamma and (especially) interleukin-4, characteristic of pro-inflammatory and anti-inflarnmatory T cells, respectively, can make microglia neuroprotective. Aggregated beta-amyloid, like bacteria cell wall-derived lipopolysaccharide, rendered the microglia cytotoxic. Cytotoxicity was correlated with a signal transduction pathway that down-regulates expression of class-II major histocompatibility proteins (MHC-II) through the MHC-II-transactivator and the invariant chain. Protection by interleukin-4 was attributed to down-regulation of tumor necrosis factor-alpha, and up-regulation of insulin-like growth factor I. These findings suggest that beneficial or harmful expression of the local immune response in the damaged CNS depends on how microglia interpret the threat, and that a well-regulated T-cell-mediated response enables microglia to alleviate rather than exacerbate stressful situations in the CNS. (c) 2005 Published by Elsevier Inc.

The trigger that leads to the pathogenesis of type 1 diabetes is currently unknown. It is well established that the pathophysiology of the disease is biphasic. In the first stage, leukocytes infiltrate the pancreatic islets in a response that does not cause damage. In the second phase, which occurs only in diabetes-prone individuals and strains, autoreactive T cells acquire aggressive potential and destroy the majority of the pancreatic islets. Rodents and humans exhibit a physiological ripple of apoptotic beta-cell death shortly after birth, which induces an adaptive autoimmune response towards islet-antigens, both in diabetes-prone non-obese diabetic (NOD) mice and in mice that do not develop diabetes. Here, we propose that the early T cell-mediated autoimmune response towards islet-antigens is physiological, purposeful and beneficial.

The failure of the spinal cord to recover after injury has been associated with the immune privilege mechanism that suppresses immune activity throughout the central nervous system. Primed macrophages and dendritic cells were shown to promote neurological recovery in preclinical models of spinal cord injury. A cell therapy consisting of autologous incubated macrophages is now being tested on spinal cord injury patients in clinical trials.

Chondroitin sulphate proteoglycan (CSPG) inhibits axonal regeneration in the central nervous system (CNS) and its local degradation promotes repair. We postulated that the enzymatic degradation of CSPG generates reparative products. Here we show that an enzymatic degradation product of CSPG, a specific disaccharide (CSPG-DS), promoted CNS recovery by modulating both neuronal and microglial behaviour. In neurons, acting via a mechanism that involves the PKCalpha and PYK2 intracellular signalling pathways, CSPG-DS induced neurite outgrowth and protected against neuronal toxicity and axonal collapse in vitro. In microglia, via a mechanism that involves ERK1/2 and PYK2, CSPG-DS evoked a response that allowed these cells to manifest a neuroprotective phenotype ex vivo. In vivo, systemically or locally injected CSPG-DS protected neurons in mice subjected to glutamate or aggregated beta-amyloid intoxication. Our results suggest that treatment with CSPG-DS might provide a way to promote post-traumatic recovery, via multiple cellular targets.

Autoimmune T cells have been viewed for decades as an outcome of immune system malfunction, and specifically as a failure to distinguish between components of self and non-self. The need for discrimination between self and non-self as a way to avoid autoimmunity has been repeatedly debated over the years. Recent studies suggest that autoimmunity, at least in the nervous system, is the body's defense mechanism against deviations from the normal. The ability to harness neuroprotective autoimmunity upon need is evidently allowed by naturally occurring CD4+CD25+ regulatory T cells, which are themselves controlled by brain-derived compounds. These findings challenge widely accepted concepts of the need for discrimination between self and non-self, as they suggest that while such discrimination is indeed required, it is needed not as away to avoid an anti-self response but to ensure its proper regulation. Whereas a response to non-self can be self-limited by a decreased presence of the relevant antigen, the response to self needs a mechanism for strict control, such as that provided by the naturally occurring regulatory T cells.

Fighting off neuronal degeneration requires a well controlled T-cell response against self-antigens residing in sites of the CNS damage. The ability to evoke this response is normally suppressed by naturally occurring CD4(+) CD25(+) regulatory T-cells (Treg). No physiological compound that controls Treg activity has yet been identified. Here, we show that dopamine, acting via type 1 dopamine receptors ( found here to be preferentially expressed by Treg), reduces the suppressive activity and the adhesive and migratory abilities of Treg. Treg activity was correlated with activation of the ERK1/2 (extracellular signal-regulated kinase 1/2) signaling pathway. Systemic injection of dopamine or an agonist of its type 1 receptors significantly enhanced, via a T-cell-dependent mechanism, protection against neuronal death after CNS mechanical and biochemical injury. These findings shed light on the physiological mechanisms controlling Treg and might open the way to novel therapeutic strategies for downregulating Treg activity (e.g., in neuronal degeneration) or for strengthening it (in autoimmune diseases).

The effects of the adaptive immune system on the cognitive performance and abnormal behaviors seen in mental disorders such as schizophrenia have never been documented. Here, we show that mice deprived of mature T cells manifested cognitive deficits and behavioral abnormalities, which were remediable by T cell restoration. T cell-based vaccination, using glatiramer acetate (copolymer-1, a weak agonist of numerous self-reactive T cells), can overcome the behavioral and cognitive abnormalities that accompany neurotransmitter imbalance induced by (+)dizocilpine maleate (MK-801) or amphetamine. The results, by suggesting that peripheral T cell deficit can lead to cognitive and behavioral impairment, highlight the importance of properly functioning adaptive immunity in the maintenance of mental activity and in coping with conditions leading to cognitive deficits. These findings point to critical factors likely to contribute to age- and AIDS-related dementias and might herald the development of a therapeutic vaccination for fighting off cognitive dysfunction and psychiatric conditions.

Protective autoimmunity was only recently recognized as a mechanism for attenuating the progression of neurodegeneration. Using a rat model of optic nerve crush or contusive spinal cord injury, and a mouse model of neurodegenerative conditions caused by injection of a toxic dose of intraocular glutamate, we show that a single low dose of whole-body or lymphoid-organ gamma-irradiation significantly improved the spontaneous recovery. Animals with severe immune deficiency or deprived of mature T cells were unable to benefit from this treatment, suggesting that the irradiation-induced neuroprotection is immune mediated. This suggestion received further support from the findings that irradiation was accompanied by an increased incidence of activated T cells in the lymphoid organs and peripheral blood and an increase in mRNA encoding for the pro-inflammatory cytokines interleukin-12 and interferon-gamma, and that after irradiation, passive transfer of a subpopulation of suppressive T cells (naturally occurring regulatory CD4(+)CD25(+) T cells) wiped out the irradiation-induced protection. These results suggest that homeostasis-driven proliferation of T cells, induced by a single low-dose irradiation, leads to boosting of T cell-mediated neuroprotection and can be utilized clinically to fight off neurodegeneration and the threat of other diseases in which defense against toxic self-compounds is needed.

Glaucoma,is a neurodegenerative disease of the optic nerve, which continues to progress even if the primary cause of degeneration is identified and alleviated. At any given time the affected optic nerve contains fibers that are amenable to neuroprotection, and will escape degeneration provided that the proper pharmacological or other intervention is applied. Autoimmunity has long been viewed as. a deleterious phenomenon that should be terminated or at least minimized in order to preserve health. We recently demonstrated, however, that T cells specific to self-proteins residing in the site of CNS insult can be protective. With the aim of boosting autoimmunity for neuroprotection without risking the induction of an autoimmune disease, we developed the use of Cop-1 (an FDA-approved drug for the treatment of multiple sclerosis) as an active vaccination for neuroprotection. Cop-1 is a synthetic polymer that weakly cross-reacts with a wide range of self-reacting T cells. Vaccination with Cop-1 resulted in significant neuroprotection in rat models of optic nerve crush and chronic glaucoma. Thus,boosting of a T cell-based mechanism, which we have termed 'protective autoimmunity', promotes recovery of the damaged optic nerve. Current studies in our laboratory are aimed at translating this treatment into a clinical therapy. (C) 2003 Elsevier Inc. All rights reserved.

Injury-induced self-destructive processes cause significant functional loss after incomplete spinal cord injury (SCI). Cellular elements of both the innate (macrophage) and the adaptive (T-cell) immune response can, if properly activated and controlled, promote post-traumatic regrowth and protection after SCI. Dendritic cells (DCs) trigger activation of effector and regulatory T-cells, providing a link between the functions of the innate and the adaptive immune systems. They also initiate and control the body's response to pathogenic agents and regulate immune responses to both foreign and self-antigens. Here we show that post-injury injection of bone marrow-derived DCs pulsed with encephalitogenic or nonencephalitogenic peptides derived from myelin basic protein, when administered (either systemically or locally by injection into the lesion site) up to 12 d after the injury, led to significant and pronounced recovery from severe incomplete SCI. No significant protection was seen in DC recipients deprived of mature T-cells. Flow cytometry, RT-PCR, and proliferation assays indicated that the DCs prepared and used here were mature and immunogenic. Taken together, the results suggest that the DC-mediated neuroprotection was achieved via the induction of a systemic T-cell-dependent immune response. Better preservation of neural tissue and diminished formation of cysts and scar tissue accompanied the improved functional recovery in DC-treated rats. The use of antigen-specific DCs may represent an effective way to obtain, via transient induction of an autoimmune response, the maximal benefit of immune-mediated repair and maintenance as well as protection against self-destructive compounds.

Glutamate, a key neurotransmitter, is pivotal to CNS function. Alterations in its concentration can be dangerous, as seen for example in acute injuries of the CNS, chronic neurodegenerative disorders and mental disorders. Its homeostasis is attributed to the efficient removal of glutamate from the extracellular milieu by reuptake via local transport mechanisms. Our recent studies suggest that glutamate, either directly or indirectly, elicits a purposeful systemic T-cell-mediated immune response directed against immunodominant self-antigens that reside at the site of glutamate-induced damage. We suggest that the harnessed autoimmunity (which we have termed 'protective autoimmunity') helps the resident microglia in their dual function as antigen-presenting cells (serving the immune system) and as cells that clear the damaged site of potentially harmful material (serving the nervous system). The interplay between glutamate and an adaptive immune response illustrates the bidirectional dialog between the immune and nervous systems, under both physiological and pathological conditions. These results point to the possible development of a therapeutic vaccination with self-antigens, or with antigens cross-reactive with self-antigens, as a way to augment autoimmunity without inducing an autoimmune disease, thus providing a safe method of limiting degeneration. This approach, which boosts a physiological mechanism for the regulation of glutamate, and possibly also that of other self-compounds, might prove to be a feasible strategy for therapeutic protection against glutamate-associated neurodegenerative or mental disorders.

Inflammation has been widely perceived as participating in the etiology of acute and chronic neurodegenerative conditions. Accordingly, in the context of traumatic injuries or chronic neurodegenerative diseases in the central nervous system (CNS), activated microglia have been viewed as detrimental and attempts have been made to treat both conditions by antiinflammatory therapy. Recent studies have suggested that microglia act as stand-by cells in the service of both the immune and the nervous systems. In the healthy CNS these cells are quiescent, but in the event of injury to axons or cell bodies they exercise their neural function by buffering harmful self-compounds and clearing debris from the damaged site, and their immune function by providing immune-related requirements for recovery. Proper regulation of the inflammatory (autoimmune) response to injury will arrest degeneration and promote regrowth, whereas inappropriate regulation will lead to ongoing degeneration. Regulation is achieved by the operation of a T cell-mediated response directed to abundant selfantigens residing in the damaged site. Since this immune-dependent mechanism was found to protect against glutamate toxicity (a major factor in neurodegenerative disorders), boosting of this response might constitute the basis for development of a therapeutic vaccination against neurodegenerative diseases, all of which exhibit similar pathways and patterns of progression.

The function of the adaptive immune response against exogenous (non-self) agents is to help the innate arm of the immune system (represented by phagocytic cells) to fight and eliminate these agents. We suggest that the body also protects itself against potentially harmful self components using mechanisms similar to those used for fighting and eliminating non-self agents, and that the protective immune activity against self-components competes with the activity of self-destructive compounds. Tolerance to self is thus not a lack of response to self, but the ability to tolerate an active defense response to self without developing an autoimmune disease.

Purpose: To evaluate the neuroprotective effect of memantine, an NMDA receptor channel blocker, in two retinal ganglion cell (RGC) injury models in rats. Methods: Neuroprotective effect of memantine was tested in partial optic nerve injury and chronic ocular hypertensive models. In the optic nerve injury model, memantine (0.1 - 30 mg/kg) was injected intraperitoneally immediately after injury. Two weeks later, optic nerve function was determined by measuring compound action potential and surviving RGC was determined by retrograde labeling with dextran tetramethyl rhodamine. Chronic ocular hypertension was attained by laser photocoagulation of episcleral and limbal veins. Memantine (5 or 10 mg/kg) was administered continuously each day with an osmotic pump, either immediately after or 10 days after first laser photocoagulation, for 3 weeks, after which RGC survival was determined. Results: Two weeks after partial optic nerve injury, there was approximate to80% reduction in RGC number. Memantine (5 mg/kg) caused a twofold increase in compound action potential amplitude and a 1.7-fold increase in survival of RGCs, respectively. In the chronic ocular hypertension model there was 37% decrease in RGCS after 3 weeks of elevated intraocular pressure. Memantine (10 mg/kg daily) reduced ganglion cell loss to 12% when applied immediately after first laser photocoagulation, and prevented any further loss when applied 10 days after first laser photocoagulation. Conclusion: The protective effect of memantine suggests that excessive stimulation of NMDA receptors by glutamate is involved in causing cell damage in these RGC injury models.

The ability of rats or mice to withstand the consequences of injury to myelinated axons in the CNS was previously shown to depend on the ability to manifest a T cell-mediated protective immune response, which is amenable to boosting by myelin-specific T cells. Here we show that this ability, assessed by retinal ganglion cell survival after optic nerve injury or locomotor activity after spinal cord contusion, is decreased if the animals were immunized as neonates with myelin proteins (resulting in their non responsiveness as adults to myelin proteins) or injected with naturally occurring regulatory CD4(+)CD25(+) T cells immediately after the injury, and is improved by elimination of these regulatory T cells. In nude BALB/c mice replenished with a splenocyte population lacking CD4(+)CD25(+) regulatory T cells, significantly more neurons survived after optic nerve injury than in nude mice replenished with a complete splenocyte population or in matched wild-type controls. In contrast, neuronal survival in wildtype BALB/c mice injected with CD4(+)CD25(+) regulatory T cells immediately after injury was significantly worse than in noninjected controls. These findings suggest that the ability to cope with the sequelae of a CNS insult is affected unfavorably by non responsiveness to myelin self-antigens and favorably by conditions allowing rapid expression of an autoimmune response. The regulatory T cells might represent an evolutionary compromise between the need to avoid autoimmune diseases and the need for autoimmunity on alert for the purpose of tissue maintenance.

Vaccination with peptides derived from interphotoreceptor retinoid-binding protein (a self-Ag that can cause experimental anto-immune uveoretinitis) resulted in protection of retinal ganglion cells from glutamate-induced death or death as a consequence of optic nerve injury. In the case of glutamate insult, no such protection was obtained by vaccination with myelin Ags (self-Ags associated with an autoimmune disease in the brain and spinal cord that evokes a protective immune response against consequences of injury to myelinated axons). We suggest that protective autoimmunity is the body's defense mechanism against destructive self-compounds, and an autoimmune disease is the outcome of a failure to properly control such a response. Accordingly, the specific self-Ag (although not necessarily its particular epitopes) used by the body for protection against potentially harmful self-compounds (e.g., glutamate) can be inferred from the specificity of the autoimmune disease associated with the site at which the stress occurs (irrespectively of the type of stress) and is in need of help.

Myelin-specific encephalitogenic T cells, when passively transferred into rats of mice, cause an experimental autoimmune disease. Previous studies by our group have shown that (a) the same cells also significantly reduce post-traumatic degeneration in these animals after injury to the central nervous system, (b) this beneficial autoimmunity is a physiological response, and (c) animals differ in their ability to resist injurious conditions, and the ability to resist post-traumatic degeneration correlates with resistance to the development of an autoimmune disease. Here we show that optic nerve neurons in both resistant and susceptible rat strains can be protected from secondary degeneration after crush injury by immunization with myelin basic protein emulsified in complete or incomplete Freund's adjuvant. We provide evidence that potentially destructive autoimmunity (causing autoimmune disease) and beneficial autoimmunity (causing improved neuronal survival) both result from activity of the same myelin-specific, proinflammatory Th1 cells. We further show that following passive transfer of such Th1 cells, the expression of their beneficial potential depends on the activity of an additional T cell (CD4(+)) population. By identifying the additional cellular component of autoimmune neuroprotection, we may be able to take meaningful steps toward achieving neuroprotection without risk of accompanying Autoimmune disease. (C) 2002 Elsevier Science B.V. All rights reserved.

PURPOSE. Glaucoma is widely accepted as a neurodegenerative disease in which retinal ganglion cell (RGC) loss is initiated by a primary insult to the optic nerve head, caused, for example, by increased intraocular pressure (IOP). In some cases, the surviving RGCs, despite adequate IOP control, may continue to degenerate as a result of their heightened susceptibility to self-destructive processes evoked by the initial damage. In animal models of mechanical or biochemical injury to the optic nerve or retina, a T-cell-mediated immune response evoked by the insult helps to reduce this ongoing loss. The current study was conducted to find out whether the ability to resist the IOP-induced loss of RGCs in a rat model is affected by the immune system. METHODS. The ocular veins and limbal plexus of rats of two strains differing in their resistance to experimental autoimmune encephalomyelitis (EAE) and in their ability to manifest a beneficial autoimmune response were laser irradiated twice to induce an increase in IOP. The pressure was measured weekly, and RGC losses were assessed 3 and 6 weeks after the first irradiation. To verify the existence of a relationship between the immune system and RGC survival, we assessed neuronal survival in Sprague-Dawley (SPD) rats devoid of mature T cells as well as after transferring splenocytes from Fisher rats, an EA,E-resistant rat strain capable of manifesting T-cell-mediated neuroprotection, to rats of a major histocompatibility complex (MHC)-matched EAE-susceptible strain (Lewis), in which the ability to manifest such protective immunity is limited. RESULTS. Both 3 and 6 weeks after the increase in IOP was initiated, the number of surviving RGCs in SPD rats, a strain in which a beneficial autoimmune response can be evoked spontaneously, was significantly higher than in Lewis rats. Moreover, in SPD rats that were thymectomized at birth, the number of surviving RGCs after an increase in IOP as adults was significantly diminished. Pa

Protective autoimmunity is the body's defense mechanism against destructive self-compounds such as those commonly associated with neurodegenerative disorders. Autoimmune disease and neurodegenerative disorders can thus be viewed as two extreme manifestations of the same process. Therefore, when designing therapy, it is important to avoid an approach that will cure the one by invoking the other. One way to stop, or at least slow down, the progression of neurodegeneration without risking development of an autoimmune disease is by boosting protective autoimmunity in a well-controlled way. Copolymer 1 (Cop-1), an approved drug for the treatment of multiple sclerosis, can be used as a treatment for autoimmune diseases and as a therapeutic vaccine for neurodegenerative diseases. We propose that the protective effect of Cop-1 vaccination is obtained through a well-controlled inflammatory reaction, and that the activity of Cop-1 in driving this reaction derives from its ability to serve as a 'universal antigen' by weakly activating a wide spectrum of self-reactive T cells.

Neurodegenerative diseases, whatever their primary causes, are characterized by certain common features, one of which is their self-perpetuating nature. The ongoing progression of the disorder is due to the effects of destructive self-compounds, whose presence in the tissues is an outcome of the early phase of the disease and which gradually destroy remaining functional neurons. Studies in our laboratory have led to the recent formulation of a novel concept of protective autoimmunity as the body's mechanism of defense against these destructive self-compounds. This autoimmune response to central nervous system (CNS) insults is mediated by T-cells and presumably operates by activating and regulating local microglia and infiltrating macrophages (inflammatory response) to carry out their function of clearing destructive material from the tissue at risk. We suggest that a well-controlled autoimmunity counteracts and overcomes the destructive effects of the potentially harmful self-compounds, at the cost of some loss of tissue. An additional risk to the individual is the induction of an autoimmune disease, which is likely to occur if the autoimmune response is malfunctioning. An optimal balance of the various factors will lead to an outcome of maximal benefit at minimal cost to the tissue. A procedure for safely boosting the autoimmune response, by vaccination with a weak self-crossreactive antigen such as glatiramer acetate (also known as Cop-1) was found to protect rats from glutamate toxicity, a major mediator of the spread of damage and a well-known causative factor in neurodegenerative disorders. Cop-1, when administered according to a different regimen, is an FDA-approved drug for the treatment of multiple sclerosis. Different formulations of the same drug can therefore be used to treat two extreme manifestations of chronic degenerative diseases of the CNS. (C) 2002 Wiley-Liss, Inc.

The myelin-associated protein Nogo-A has received more research attention than any other inhibitor of axonal regeneration in the injured central nervous system (CNS). Circumvention of its inhibitory effect, by using antibodies specific to Nogo-A, has been shown to promote axonal regrowth. Studies in our laboratory have demonstrated that active or passive immunization of CNS-injured rats or mice with myelin-associated peptides induces a T-cell-mediated protective autoimmune response, which promotes recovery by reducing posttraumatic degeneration. Here, we show that neuronal degeneration after incomplete spinal-cord contusion in rats was substantially reduced, and hence recovery was significantly promoted, by posttraumatic immunization with p472, a peptide derived from Nogo-A. The observed effect seemed to be mediated by T cells and could be reproduced by passive transfer of a T cell line directed against the Nogo-A peptide. Thus, it seems that after incomplete spinal-cord injury, immunization with a variety of myelin-associated peptides, including those derived from Nogo-A, can be used to evoke a T cell-mediated response that promotes recovery. The choice of peptide(s) for clinical treatment of spinal-cord injuries should be based on safety considerations; in particular, the likelihood that the chosen peptide will not cause an autoimmune disease or interfere with essential functions of this peptide or other proteins. From a therapeutic point of view, the fact that the active cellular agents are T cells rather than antibodies is an advantage, as T cell production commences within the time window required for a protective effect after spinal-cord injury, whereas antibody production takes longer.

Functional loss after injury to the mammalian central nervous system (CNS) has been attributed not only to the immediate loss of neurons but also to secondary neuronal degeneration caused by the toxicity of physiological Compounds present in abnormally high amounts as a result of the injury. One such compound appears to be the protease thrombin. Here we show that the beneficial effect of T cells directed against myelin self-antigens can be attributed, at least in part, to the ability of these 'autoimmune' T cells to produce antithrombin III. Using transgenic mice lacking the thrombin receptor PAR-1, we also present molecular evidence indicating that down-regulation of PAR-I by genetic manipulation leads to increased post-traumatic survival of CNS neurons. We further show that the ability of autoimmnune T cells to produce thrombin inhibitors and to exert post-traumatic neuroprotection are both independent of their PAR-1 expression. These findings suggest that thrombin plays a key role in post-injury neuronal survival, and that its post-traumatic toxicity can be down-regulated by appropriate alteration of the amounts of PAR-1 receptors or of antithrombin III, the latter achieved by T cell-mediated autoimmunity. (C) 2001 Elsevier Science B.V. All rights reserved.

Axonal injury initiates a process of neuronal degeneration, with resulting death of neuronal cell bodies. We show here that in C57BL/6J mice, previously shown to have a limited ability to manifest a post-traumatic protective immunity, the rate of neuronal survival is increased if IL-6 is deficient during the first 24 hours after optic nerve injury. Immunocytochemical staining preformed 7 days after the injury revealed an increased number of activated microglia in the IL-6-deficient mice compared to the wild-type mice. In addition, IL-6-deficient mice showed an increased resistance to glutamate toxicity. These findings suggest that the presence of IL-6 during the early post-traumatic phase, at least in mice that are susceptible to autoimmune disease development, has a negative effect on neuronal survival. This further substantiates the contention that whether immune-derived factors are beneficial or harmful for nerve recovery after injury depends on the phenotype of the immune cells and the timing and nature of their dialog with the damaged neural tissue. (C) 2001 Elsevier Science B.V. All rights reserved.

Injury to the CNS is often followed by a spread of damage (secondary degeneration), resulting in neuronal losses that are substantially greater than might have been predicted from the severity of the primary insult. Studies in our laboratory have shown that injured CNS neurons can benefit from active or passive immunization with CNS myelin-associated antigens. The fact that autoimmune T-cells can be both beneficial and destructive, taken together with the established phenomenon of genetic predisposition to autoimmune diseases, raises the question: will genetic predisposition to autoimmune diseases affect the outcome of traumatic insult to the CNS? Here we show that the survival rate of retinal ganglion cells in adult mice or rats after crush injury of the optic nerve or intravitreal injection of a toxic dosage of glutamate is up to twofold higher in strains that are resistant to the CNS autoimmune disease experimental autoimmune encephalomyelitis (EAE) than in susceptible strains. The difference was found to be attributed, at least in part, to a beneficial T-cell response that was spontaneously evoked after CNS insult in the resistant but not in the susceptible strains. In animals of EAE-resistant but not of EAE-susceptible strains devoid of mature T-cells (as a result of having undergone thymectomy at birth), the numbers of surviving neurons after optic nerve injury were significantly lower (by 60%) than in the corresponding normal animals. Moreover, the rate of retinal ganglion cell survival was higher when the optic nerve injury was preceded by an unrelated CNS (spinal cord) injury in the resistant strains but not in the susceptible strains. It thus seems that, in normal animals of EAE-resistant strains (but not of susceptible strains), the injury evokes an endogenous protective response that is T-cell dependent. These findings imply that a protective T-cell-dependent response and resistance to autoimmune disease are regulated by a common mechanism. The result

Primary damage caused by injury to the CNS is often followed by delayed degeneration of initially spared neurons. Studies in our laboratory have shown that active or passive immunization with CNS myelin-associated self-antigens can reduce this secondary loss. Here we show, using four experimental paradigms in rodents, that CNS trauma spontaneously evokes a beneficial T cell-dependent immune response, which reduces neuronal loss. (1) Survival of retinal ganglion cells in rats was significantly higher when optic nerve injury was preceded by an unrelated CNS (spinal cord) injury. (2) Locomotor activity of rat hindlimbs (measured in an open field using a locomotor rating scale) after contusive injury of the spinal cord (T8) was significantly better (by three to four score grades) after passive transfer of myelin basic protein (MBP)-activated splenocytes derived from spinally injured rats than in untreated injured control rats or rats similarly treated with splenocytes from naive animals or with splenocytes from spinally injured rats activated ex vivo with ovalbumin or without any ex vivo activation. (3) Neuronal survival after optic nerve injury was 40% lower in adult rats devoid of mature T cells (caused by thymectomy at birth) than in normal rats. (4) Retinal ganglion cell survival after optic nerve injury was higher (119 +/- 3.7%) in transgenic mice overexpressing a T cell receptor (TcR) for MBP and lower (85 +/- 1.3%) in mice overexpressing a T cell receptor for the non-self antigen ovalbumin than in matched wild types. Taken together, the results imply that CNS injury evokes a T cell-dependent neuroprotective response.

In glaucoma, as in other degenerative diseases of the central nervous system (CNS), there are some neurons which, although susceptible to degeneration, are amenable to neuroprotection. Until very recently, attempts: to attenuate the spread of damage after CNS trauma or in neurodegenerative diseases did not include recruitment of the immune system, because it was assumed that ill the C:NS any immune activity, particularly autoimmune activity, would be harmful. Using the injured optic nerve of the rat as a model, we recently showed, however, that this 'secondary degeneration' call be slowed down by a well-controlled adaptive immune response, which is mediated by T cells: against C:NS sells antigens, such as myelin basic protein (MBP), and can be achieved either by passive transfer of MBP-activated T cells or by active immunization with MBP. Accordingly, we suggested that;autoimmune T cells can be neuroprotective. We further showed that the neuroprotective autoimmunity is a physiological response to the injury, perhaps insufficient in its natural state, but amenable to boosting. The neuroprotective activity of these T cells probably depends on their reactivation by their specific antigen after they are targeted to the injured nerve. As nerve degeneration is initiated and sustained by many. factors, it would probably best be counteracted by a comprehensive type of therapy rather than treatment that addresses only some aspects of nerve damage. T cell therapy, being physiological rather than pharmaceutical in nature, may provide a global answer. However, since the neuroprotective immune response is directed against die self, it must be rigorously regulated to avoid inducing an autoimmune disease. We showed that the synthetic copolymer Cop-1 can passively or actively evoke T cell-mediated neuroprotection, probably by cross-reacting with MBP. Safe synthetic peptides that resemble self-antigens and are cross-activated by CNS-associated self antigens may be a useful starti

Our group recently demonstrated that autoimmune T cells directed against central nervous system-associated myelin antigens protect neurons from secondary degeneration. We further showed that the synthetic peptide copolymer 1 (Cop-1), known to suppress experimental autoimmune encephalomyelitis, can be safely substituted for the natural myelin antigen in both passive and active immunization for neuroprotection of the injured optic nerve. Here we attempted to determine whether similar immunizations are protective from retinal ganglion cell loss resulting from a direct biochemical insult caused, for example, by glutamate (a major mediator of degeneration in acute and chronic optic nerve insults) and in a rat model of ocular hypertension. Passive immunization with T cells reactive to myelin basic protein or active immunization with myelin oligodendrocyte glycoprotein-derived peptide, although neuroprotective after optic nerve injury, was ineffective against glutamate toxicity in mice and rats. In contrast, the number of surviving retinal ganglion cells per square millimeter in glutamate-injected retinas was significantly larger in mice immunized 10 days previously with Cop-1 emulsified in complete Freund's adjuvant than in mice injected with PBS in the same adjuvant (2,133 +/- 270 and 1,329 +/- 121, respectively, mean +/- SEM; P

A recent study in our laboratory showed, against all expectations, that macrophages and a particular type of T cell, by promoting regrowth and reducing the post-traumatic spread of damage in the injured rat optic nerve or spinal cord, have a beneficial effect on the injured CNS. Macrophages in the CNS have long been thought to have predominantly destructive effects. Autoimmunity in general, and in the CNS in particular, has never been documented as a purposeful physiological response of benign character. Our results suggest that after traumatic injury to the central nervous system (CNS), both of these immune cell types potentially have beneficial effects: macrophages can promote repair and T cells of a particular specificity can reduce the spread of damage. However, possibly because of the immune-privileged character of the CNS, the spontaneously evoked physiological activities of both macrophages and T cells in the CNS are restricted, and appear to need well-controlled boosting in order to be effective. It thus appears that (i) a stress signal transmitted from the traumatized tissue (in this case the CNS) for recruitment of the adaptive immune system does not have to be pathogen-related in order to evoke a response, (ii) a response to self is not necessarily a quirk of nature, and (iii) an autoimmune response, provided that it is well-regulated, helps the individual to cope with stress signals from the traumatized CNS, and thus plays a role in maintenance of the injured tissue without posing a threat to the organism. (C) 2001 Elsevier Science B.V. All rights reserved.

The physiological conditions under which beneficial autoimmunity is evoked have never been documented. We recently demonstrated that autoimmune T cells directed against myelin-associated self-proteins. when passively transferred into rats or mice, reduce the spread of damage after traumatic injury to central nervous system axons. This finding raised a fundamental question: does this beneficial effect represent a physiological neuroprotective response that normally is too weak to be effective and requires boosting, or is it simply the welcome result of an ex vivo manipulation? It appears from our studies that trauma, at least in the central nervous system. evokes a stress signal that activates a T cell dependent response directed against self antigens, and that this response is physiological in nature, beneficial in intent, and amenable to boosting by active or passive immunization.

T-cell autoimmunity to myelin basic protein was recently shown to be neuroprotective in injured rat optic nerves. In the present study, using the mouse optic nerve, we examined whether active immunization rather than passive transfer of T-cells can be beneficial in protecting retinal ganglion cells (RGCs) from post-traumatic death. Before severe crush injury of the optic nerve, SJL/J and C3H. SW mice were actively immunized with encephalitogenic or nonencephalitogenic peptides of proteolipid protein (PLP) or myelin oligodendrocyte glycoprotein (MOG), respectively. At different times after the injury, the numbers of surviving RGCs in both strains immunized with the nonencephalitogenic peptides pPLP 190-209 or pMOG 1-22 were significantly higher than in injured controls treated with the non-self-antigen ovalbumin or with a peptide derived from beta -amyloid, a non-myelin-associated protein. Immunization with the encephalitogenic myelin peptide pPLP 139-151 was beneficial only when the disease it induced, experimental autoimmune encephalomyelitis, was mild. The results of this study show that survival of RGCs after axonal injury can be enhanced by vaccination with an appropriate self-antigen. Furthermore, the use of nonencephalitogenic myelin peptides for immunization apparently allows neuroprotection without incurring the risk of an autoimmune disease. Application of these findings might lead to a promising new approach for treating optic neuropathies such as glaucoma.

Neurotrophins (NTs) promote neuronal survival and maintenance during development and after injury. However, their role in the communication between the nervous system and the immune system is not yet clear. We observed recently that passively transferred activated T cells of various antigen specificities home to the injured central nervous system (CNS), yet only autoimmune T cells specific to a CNS antigen, myelin basic protein (MBP), protect neurons from secondary degeneration after crush injury of the rat optic nerve. Here we examined the involvement of NTs in T-cell-mediated neuroprotection, and the possible significance of the antigen specificity of the T cells in this activity. Analysis of cytokine and NT expression in various rat T cell lines showed that the T cells express mRNA for cytokines of Th1, Th2, and Th3 phenotypes. in addition, the T cells express mRNA and protein specific to nerve growth factor, brain-derived neurotrophic factor, NT-3, and NT-4/5. Antigen activation significantly increased NT secretion. Thus, reactivation of CNS autoimmune T cells by locally presented antigens to which they are specific can lead to enhanced secretion of NTs and possibly also of other factors in injured optic nerves. mRNA for TrkA, TrkB and p75 receptors was expressed in the injured nerve, suggesting that these specific receptors can mediate the effects of the T-cell-derived NTs. The neuroprotective effect of the passively transferred autoimmune anti-MBP T cells in injured optic nerves was significantly decreased after local applicaiton of a tyrosine kinase inhibitor known to be associated with NT-receptor activity. These results suggest that the neuroprotective effect of autoimmune T cells involves the secretion of factors such as NTs by the T cells reactivated by their specific antigen in the injured CNS. T cell intervention in the injured CNS might prove to be a useful means of promoting post-injury CNS maintenance and recovery, possibly via supply of NTs and o

Degenerative diseases of the central nervous system (CNS) are characterized by progressive degeneration, which continues even after the primary causative factor has been identified and neutralized or removed. The progressive degeneration is thought to be a self perpetuating process, attributable in part to factors derived from the degenerating nerves themselves. If this is so, the mediators of toxicity causing secondary degeneration in the various degenerative diseases are likely to be similar. Common mediators include excitatory amino acids, compounds that cause oxidative or metabolic stress, and factors that disturb the ionic balance of the nerve's extracellular milieu. It is proposed here, based on recent work by the author and by others on CNS trauma and autoimmunity, as well as on accumulated information about the professional role of the adaptive and the immune response in general, that active or passive T cell-mediated autoimmunity directed against self-antigens associated with the disorder will be beneficial in halting the spread of damage. Drug Dev. Res. 50:223-225, 2000. (C) 2000 Wiley-Liss, Inc.

We recently demonstrated that autoimmune T cells protect neurons from secondary degeneration after central nervous system (CNS) axotomy in rats. Here we show, using both morphological and electrophysiological analyses, that the neuroprotection is long-lasting and is manifested functionally. After partial crush injury of the rat optic nerve, systemic injection of autoimmune T cells specific to myelin basic protein significantly diminished the loss of retinal ganglion cells and conducting axons, and significantly retarded the loss of the visual response evoked by light stimulation. These results support our challenge to the traditional concept of autoimmunity as always harmful, and suggest that in certain situations T cell autoimmunity may actually be beneficial. It might be possible to employ T cell intervention to slow down functional loss in the injured CNS. (C) 2000 Published by Elsevier Science B.V. All rights reserved.

Autoimmunity is usually considered only as a cause of disease; nevertheless, human T-cell repertoires are filled naturally with autoimmune lymphocytes. Here, we review evidence that autoimmune T cells can help heal damaged tissues, indicating that natural autoimmunity could also be a cause of health.

Keywords: CENTRAL-NERVOUS-SYSTEM; SPINAL-CORD INJURY; PERIPHERAL-NERVE; T-CELLS; WALLERIAN DEGENERATION; MACROPHAGE RESPONSES; AXONAL REGENERATION; WOUND REPAIR; RECOVERY; INFLAMMATION

Traumatic injury to the optic nerve is accompanied by a process of degeneration that leads to apoptotic death of the corresponding cell bodies. In the traumatized nerves, a process of self-propagating degeneration takes place and is mediated by toxic substances, some of them well-documented and others not yet identified. Using the partially injured rat optic nerve as a model, we show that glutamate is in above-normal concentrations in the aqueous humor of eyes with impaired optic nerves. Neurons which had escaped the primary insult and were still viable were more susceptible to glutamate than normal uninjured nerves, and were likely to die as a result of exposure to glutamate even at low concentrations. A transient accumulation of autoimmune T cells was observed at the site of the optic nerve injury. We show that autoimmune T cells that react specifically with myelin basic protein and are known to be detrimental to the central nervous system (CNS) are beneficial in reducing the deficit caused by secondary degeneration. The protective effect of the T cells is demonstrated both morphologically and functionally. We suggest that these autoimmune T cells may induce a benign immune response to pathogen-free CNS lesions, and that it may be possible to boost this response therapeutically in a way that will allow the neuroprotective effect to be manifested without incurring the risk of autoimmune disease.

Neurodegenerative diseases are characterized by a relentless loss of specific groups of neuronal subtypes. Many of these diseases share similar molecular mechanisms and extracellular mediators of neuronal loss. We now suggest that neurodegeneration originating in the neuronal cell bodies (e.g. in Alzheimer's disease, Parkinson's disease and amyotrophic lateral sclerosis) should be distinguished from that originating in the axons (e.g. in glaucoma, certain peripheral neuropathies and spinal stenosis). We propose that the former group of diseases be defined as "somagenic" and the latter as 'axogenic'. Although axogenic disorders may share common symptoms and mediators of toxicity with somagenic disorders, they have distinct temporal, subcellular and signal-transduction features. We further suggest that, by adopting this classification of disorders based on pathophysiological processes, we will come go recognize additional diseases (in particular, those defined as axogenic) as being neurodegenerative and therefore possibly amenable to neuroprotective therapy.

The central nervous system (CNS), unlike the peripheral nervous system (PNS), is an immune-privileged site in which local immune responses are restricted. Whereas immune privilege in the intact CNS has been studied intensively, Little is known about its effects after trauma. In this study, we examined the influence of CNS immune privilege on T cell response to central nerve injury. Immunocytochemistry revealed a significantly greater accumulation of endogenous T cells in the injured rat sciatic nerve than in the injured rat optic nerve (representing PNS and CNS white matter trauma, respectively). Use of the in situ terminal deoxytransferase-catalyzed DNA nick end labeling (TUNEL) procedure revealed extensive death of accumulating T cells in injured CNS nerves as well as in CNS nerves of rats with acute experimental autoimmune encephalomyelitis, but not in injured PNS nerves, Although Fas ligand (FasL) protein was expressed in white matter tissue of both systems, it was more pronounced in the CNS, Expression of major histocompatibility complex (MHC) class II antigens was found to be constitutive in the PNS, but in the CNS was induced only after injury, Our findings suggest that the T cell response to central nerve injury is restricted by the reduced expression of MHC class II antigens, the pronounced Fast expression, and the elimination of infiltrating lymphocytes through cell death.

It is now widely accepted that injured nerves, like any other injured tissue, need assistance from their extracellular milieu in order to heal. We compared the postinjury activities of thrombin and gelatinases, two types of proteolytic activities known to be critically involved in tissue healing, in nonregenerative (rat optic nerve) and regenerative (fish optic nerve and rat sciatic nerve) neural tissue. Unlike gelatinases, whose induction pattern was comparable in all three nerves, thrombin-like activity differed clearly between regenerating and nonregenerating nervous systems. Postinjury levels of this latter activity seem to dictate whether it will display beneficial or detrimental effects on the capacity of the tissue for repair. The results of this study further highlight the fact that tissue repair and nerve regeneration are closely linked and that substances that are not unique to the nervous system, but participate in wound healing in general, are also crucial for regeneration or its failure in the nervous system.

PURPOSE. The neurodegenerative progression of glaucoma is considered to be related not only to primary risk factors such as the elevation of intraocular pressure, but also to mediators of secondary neuronal degeneration. In the present study, the neuroprotective activity of the alpha 2-adrenoreceptor agonists brimonidine, AGN 191103, and clonidine were examined in an animal model that simulates secondary neuronal degeneration of the optic nerve in a way thought to be independent of elevation of intraocular pressure. The beta-blocker timolol, currently used clinically to decrease intraocular pressure, was also examined for neuroprotective activity at dosages corresponding to the effective antihypertensive dosage. METHODS. A Single dose of each of the tested compounds was administered intraperitoneally immediately after partial crush injury of the rat optic nerve. Secondary degeneration was measured by determining injury-induced deficits with and without the drug. This was achieved electrophysiologically by measurement of compound action potential amplitude, and morphometrically by counting the retrogradely labeled retinal ganglion cells, representing viable optic nerve axons, in wholemounted retinas. RESULTS. All three alpha 2-adrenoreceptor agonists, but not timolol, exhibited neuroprotective effects. Treatment immediately after injury with each of these agonists resulted in a dose-dependent attenuation of the injury-induced decrease in compound action potential amplitude. Moreover, after treatment with 100 mu g/kg brimonidine administered intraperitoneally, the loss of retinal ganglion cells 2 weeks after injury was three times lower than in saline-treated animals. CONCLUSIONS. In addition to their known effect of lowering intraocular pressure, alpha 2-adrenoreceptor agonists, unlike timolol, exert a neuroprotective effect. Use of the rat optic nerve model of partial crush injury can serve as a method of screening compounds that are potentially capable of allevi

Autoimmunity to antigens of the central nervous system is usually considered detrimental. T cells specific to a central nervous system self antigen, such as myelin basic protein, can indeed induce experimental autoimmune encephalomyelitis, but such T cells may nevertheless appear in the blood of healthy individuals. We show here that autoimmune T cells specific to myelin basic protein can protect injured central nervous system neurons from secondary degeneration. After a partial crush injury of the optic nerve, rats injected with activated anti-myelin basic protein T cells retained approximately 300% more retinal ganglion cells with functionally intact axons than did rats injected with activated T cells specific for other antigens. Electrophysiological analysis confirmed this finding and suggested that the neuroprotection could result from a transient reduction in energy requirements owing to a transient reduction in nerve activity. These findings indicate that T-cell autoimmunity in the central nervous system, under certain circumstances, can exert a beneficial effect by protecting injured neurons from the spread of damage.

GTP and Ca2+, two well-known modulators of intracellular signaling pathways, control a structural/functional switch between two vital and mutually exclusive activities, cross-linking and G(alpha) activity, in the same enzyme. The enzyme, a brain-derived tissue-type transglutaminase (TGase), was recently cloned by us in two forms, one of which (s-TGN) lacks a C-terminal region that is present in the other (1-TGN). Immunoreaction with antibodies directed against a peptide present in the C-terminus of 1-TGN but missing in s-TGN suggested that this site, which is located in the C-terminal fourth domain, undergoes conformational changes as a result of interaction between 1-TGN and GTP. Site-directed mutagenesis suggested that the third domain is involved in mediating the inhibition of the cross-linking activity. These results were supported by molecular modeling, which further suggested that domains III and IV both participate in conformational changes leading to the functional switch between the Ca2+-dependent cross-linking activity and the G(alpha) activity. (C) 1998 Academic Press.

Neuroprotective therapy is a relatively new development in the approach to the treatment of acute and chronic brain damage. Though initially viewed in the framework of acute CNS injuries, the concept was recently extended to include chronic injuries, in which at any given time there are some neurons in an acute phase of degeneration coexisting with others that are healthy, marginally damaged, or dead. The healthy neurons and those that are only marginally damaged are the potential targets for neuroprotection. For the development of neuroprotective therapies, it is essential to employ an animal model in which the damage resulting from secondary degeneration can be quantitatively distinguished from primary degeneration. This is of particular relevance when the site of the damage is in the white matter (nerve fibers) rather than in the gray matter (cell bodies). In the present work we reexamine the concepts of secondary degeneration and neuroprotection in white matter lesions. Using a partial crush injury of the adult rat optic nerve as a model, we were able to assess both primary and secondary nerve damage. We show that neurons whose axons were not damaged or only marginally damaged after an acute insult will eventually degenerate as a consequence of their existence in the degenerative environment produced by the injury. This secondary degeneration does not occur in all of the neurons at once, but affects them in a stepwise fashion related to the severity of the damage inflicted. These findings, which may be applicable to the progression of acute or chronic neuropathy, imply that neuroprotective therapy may have a beneficial effect even if there is a time lag between injury and treatment. (C) 1998 Academic Press.

Axons in the central nervous system (CNS) of adult mammals do not regenerate after injury. Mammalian CNS differs In this respect from other mammalian tissues, Including the peripheral nervous system (PNS), and from the CNS of lower vertebrates. In most parts of the body, including the nervous system, injury triggers an Inflammatory reaction involving macrophages. This reaction Is needed for tissue healing; when it Is delayed or insufficient, healing Is incomplete. The CNS, although needing an efficient inflammatory reaction resembling that In the periphery for tissue healing, appears to have lost the ability to supply it. We suggest that restricted CNS recruitment and activation of macrophages are linked to regeneration failure and might reflect the Immune privilege that characterizes the mammalian CNS. As macrophages play a critical role In tissue restoration, and because their recruitment and activation are among the most upstream of the events leading to tissue healing, overcoming the deficiencies In these steps might trigger a self-repair process leading to recovery after CNS Injury.

Postinjury recovery in most tissues requires an effective dialog with macrophages; however, in the mammalian central nervous system, this dialog may be restricted (possibly due to its immune-privileged status), which probably contributes to its regeneration failure. We circumvented this by implanting macrophages, pre-exposed ex vivo to peripheral nerve segments, into transected rat spinal cord. This stimulated tissue repair and partial recovery of motor function, manifested behaviorally by movement of hind limbs, plantar placement of the paws and weight support, and electrophysiologically by cortically evoked hind-limb muscle response. We substantiated these findings immunohistochemically by demonstrating continuity of labeled nerve fibers across the transected site, and by tracing descending fibers distally to it by anterograde labeling. In recovered rats, re-transection of the cord above the primary transection site led to loss of recovery, indicating the involvement of long descending spinal tracts. Injection of macrophages into the site of injury is relatively non-invasive and, as the cells are autologous, it may be developed into a clinical therapy.

Tissue-type transglutaminases (TGases) were recently shown to exert dual enzymatic activities; they catalyze the posttranslational modification of proteins by transamidation, and they also act as guanosine triphosphatase (GTPase). Here we show that a tissue-type TGase is expressed in rat brain astrocytes in vitro, and is induced by the inflammation-associated cytokines interleukin-1 beta and to a lesser extent by tumor necrosis factor-alpha. Induction is accompanied by overexpression and appearance of an additional shorter clone, which does not contain the long 3'-untranslated region and encodes for a novel TGase enzyme whose C terminus lacks a site that affects the enzyme's interaction with guanosine triphosphate (GTP), Expression of two clones revealed that the long form is inhibited noncompetitively by GTP, but the short form significantly less so, The different affinities for GTP may account for the difference in physiological function between these two enzymes.

In this study we present a method to achieve a complete transection of optic nerve axons in adult rat, while preserving the vasculature and retaining the continuity of the meninges. Under deep anesthesia, the optic nerve of adult rat is exposed. Using specially designed instruments built from disposable glass microsampling pipets, a small opening is created in the meninges of the optic nerve, 2-3 mm behind the eye globe. A glass dissector is introduced through the opening and is used to cut all the axons through the whole width of the nerve. Complete transfection of the optic nerve axons was achieved, while retaining the continuity of the meninges and avoiding damage to the nerve's vascular supply, Transection was confirmed by transillumination showing a complete gap in the continuity of the nerve axons, and by both morphological and electrophysiological criteria. Nerve transection performed by the conventional technique leads to neuroma formation and hampers regeneration. Crush injury may cause nerve ischemia, which is detrimental to axonal recovery. Both of these disadvantages are avoided by the method of transection presented here. The opening created in the 'meningeal tube' can be used to inject substances that may be of benefit in recovery, rescue and/or regeneration of the injured axons. The model is particularly suitable for in vivo studies on nerve regeneration, and especially for screening of putative therapeutic agents.

Macrophages have long been known to play a key role in the healing processes of tissues that regenerate after injury; however, the nature of their involvement in healing of the injured central nervous system (CNS) is still a subject of controversy, Here we show that the absence of regrowth in transected rat optic nerve (which, like all other CNS nerves in mammals, cannot regenerate after injury) can be overcome by local transplantation of macrophages preincubated ex vivo with segments of a nerve (e.g., sciatic nerve) that can regenerate after injury, The observed effect of the transplanted macrophages was found to be an outcome of their stimulated activity, as indicated by phagocytosis, Thus, macrophage phagocytic activity was stimulated by their preincubation with sciatic nerve segments but inhibited by their preincubation with optic nerve segments. We conclude that the inability of nerves of the mammalian CNS to regenerate is related to the failure of their macrophages recruited after injury to acquire growth-supportive activity. We attribute this failure to the presence of a CNS resident macrophage inhibitory activity, which may be the biochemical basis underlying the immune privilege of the CNS, The transplantation of suitably activated macrophages into injured nerves may overcome multiple malfunctioning aspects of the CNS response to trauma, and thus may be developed into a novel, practical, and multipotent therapy for CNS injuries.

HU-211 is a novel synthetic analog of tetrahydrocannabinol that was recently shown in animal models to be nonpsychotropic. In this study we show that HU-211 can potentially be used as a neuroprotective compound in the CNS. Using a calibrated crush injury of adult rat optic nerve, we show that HU-211 can reduce injury-induced metabolic and electrophysiological deficits. Energy metabolism was monitored by measuring the intramitochondrial nicotine-amine adenine dinucleotide redox state hourly for 6 h after injury and treatment. Electrophysiological activity was assessed by compound action potential and visual evoked potential response. Beneficial effects were dose-dependent, being optimal at 7 mg/kg, administered intraperitoneally. The time window during which treatment was effective was found to be from the time of injury for at least 5 h, with treatment most effective at the time of injury. These results strongly suggest that HU-211, given immediately after CNS injury at the optimal dosage, may possess neuroprotective activities.

Migration of astrocytes is thought to play a role in nerve regeneration and to be mediated, at least in part, by inflammation-associated cytokines, Plasminogen activators are secreted proteases that function in fibrinolysis and participate in cellular migration and invasion and, in some cases, are modulated by cytokines, Here, we show that two cytokines, tumor necrosis factor-alpha and interleukin-1 beta, can modulate plasminogen activation in astrocytes, each causing 90% reduction of total plasminogen activator activity. Direct and reverse zymography indicated that this reduction resulted from two simultaneous events, a pronounced decrease in tissue-type plasminogen activator activity and an induction of plasminogen activator inhibitor-1. Northern hybridization analysis indicated a 30-fold increase of the steady-state level of plasminogen activator inhibitor-1 mRNA following treatment with each of the two cytokines. Both of the cytokine-induced effects could be blocked by cycloheximide or actinomycin D. When signal transduction pathways were blocked, the results indicated the involvement of reduction in cyclic AMP levels, protein kinase activity, and arachidonic metabolites of the lipoxygenase pathway. The results thus show that the two cytokines reduce the ability of astrocytes to conduct fibrinolysis and extracellular proteolysis, and suggest that the effect of these cytokines on members of the plasminogen activation system is through a common signal transduction pathway.

Cytokines have been suggested to be involved in the cross talk between the immune and the nervous systems, under normal and pathological conditions, For example, the cytokine interleukin-2 was suggested to be involved in response to CNS trauma and spontaneous regeneration. Here, we examined whether mammalian CNS has an intrinsic potential to produce interleukin-2 and, if so, what its cellular origin is. mRNA sequences encoding for interleukin-2 were detected in brains of humans and rodents. Northern blot analysis revealed the presence of several interleukin-2 transcripts of different sizes in the brain, all recognized by lymphocyte-derived interleukin-2 cDNA probes. One of the transcripts, a high molecular weight form of similar to 5 kb, appeared to be unique to the brain. Reverse transcription and amplification by PCR of human fetal brain mRNA revealed one cDNA product that, upon sequence analysis, showed a high degree of homology with the human lymphocyte-derived interleukin-2 coding sequence, To identify the possible cellular source of the interleukin-2 transcripts within the mammalian brain, we similarly analyzed mRNA of rat brain cells in culture. Northern blot analysis revealed that astrocytes contain transcripts that hybridize with interleukin-2 cDNA probe. These findings point to the astrocytes as a possible source of brain interleukin-2.

A covalent dimer of interleukin (IL)-2, produced in vitro by the action of a nerve-derived transglutaminase, has been shown previously to be cytotoxic to mature rat brain oligodendrocytes. Here we report that this cytotoxic effect operates via programmed cell death (apoptosis) and that the p53 tumor suppressor gene is involved directly in the process. The apoptotic death of mature rat brain oligodendrocytes in culture following treatment with dimeric IL-2 was demonstrated by chromatin condensation and internucleosomal DNA fragmentation. The peak of apoptosis was observed 16-24 h after treatment, while the commitment to death was already observed after 3-4 h. An involvement of p53 in this process was indicated by the shift in location of constitutively expressed endogenous p53 from the cytoplasm to the nucleus, as early as 15 min after exposure to dimeric IL-2. Moreover, infection with a recombinant retrovirus encoding a C-terminal p53 minoprotein, shown previously to act as a dominant negative inhibitor of endogenous wild-type p53 activity, protected these cells from apoptosis.

An interleukin-2 dimer, produced enzymatically by a nerve-derived transglutaminase in vitro, is cytotoxic to oligodendrocytes, unlike the immune-derived monomeric interleukin-2. The abject of this study was to establish a way to produce a dimer of interleukin-2 in quantities, by means of genetic engineering, and to confirm that the structure of the resulting molecule is critical for its function. A defective herpes simplex virus vector was utilized for overproduction of a dimeric interleukin-2. The resulting linear dimer, which is a translational product, differs from the enzymatically produced dimer, which is a posttranslational modification of interleukin-2. The linear dimer, while retaining the known interleukin-2 activity of monomeric interleukin-2 with respect to mitogenicity on T cells, was not cytotoxic to oligodendrocytes. This finding suggests that the lack of cytotoxicity of the linear dimeric interleukin-2 is not caused by a loss of activity during its preparation but is related to its conformational structure, which evidently does not meet the requirements for cytotoxicity. This study opens the way to the design at the transcriptional level of modified proteins and their efficient production, provided that the new transcript encodes for the desired modification in the protein at the appropriate sites.

Laser irradiation at subthreshold energies exerts various effects on the eye and other parts of the body, mainly the skin and nervous system, through a mechanism that has yet to be adequately explained. The ocular bioeffects are manifested mostly in the retina, but also in other ocular tissues. This review outlines the reported effects of low-energy laser irradiation on nonophthalmological tissues and organs, including those of the nervous system, with special emphasis on the optic nerve. It also details the ophthalmic phenomena induced by low-energy laser irradiation and examines claims of its therapeutic efficacy in several eye diseases, such as keratitis, glaucoma and macular degeneration.

Recent results shed new light on how some nervous systems can regenerate after injury while others cannot. Until recently it was widely believed that the main difference between systems that regenerate and those that do not lies in the normal state of their permissiveness to the regenerating axons, Thus, while nonregenerative systems such as the rat optic nerve, were shown to contain myelin-associated growth inhibitors, regenerative systems such as the fish optic nerve, were thought to have no such inhibitors. However, it has now been demonstrated that spontaneously regenerating systems do contain growth inhibitors, though their levels seem to be lower than in nonregenerative systems. The main difference, however, appears to reside in the system's response to injury. This article discusses the involvement of myelin-associated growth inhibitors in the spontaneously regenerating nervous system of fish, traces the apparent discrepancy, and shows how it has been resolved recently.

This study shows that the fish optic nerve, which is able to regenerate after injury, contains myelin-associated growth inhibitors similar to the growth inhibitors present in mammalian central nervous system (CNS) myelin. The ability of nerves to regenerate was previously correlated with the ability of sections from these nerves to support neuronal attachment and axonal growth in vitro. Thus neuroblastoma cells or embryonic neurons became attached to and grew axons on sections of rat sciatic nerve or fish optic nerve, which are spontaneously regenerating systems, but not on sections of rat optic nerve, a nonregenerating system. Failure of the latter to support axonal growth has been attributed, at least in part, to growth inhibitors. Recently it was shown that adult neurons, which differ in their growth requirement from embryonic neurons, are unable to extend neurites on sections of normal sciatic nerve but are able to extend neurites on sections of sciatic nerve that was injured prior to its excision. We found a similar situation in the fish optic nerve, i.e., that the nerve is normally not permissive to growth of adult retinal axons but becomes growth permissive after injury. The nonpermissiveness of the normal fish optic nerve was found to correlate with the presence of myelin-associated growth-inhibitory molecules. This inhibitory activity of fish myelin was neutralized by IN-1 antibodies, known to neutralize rat myelin growth inhibitors. The results thus demonstrate that fish optic nerve myelin contains inhibitors apparently similar or even identical to those of rat, but possibly present in lower amounts than in the rat. Results are discussed with respect to the possibility that fish optic nerve, like the rat sciatic nerve and unlike the rat optic nerve, undergoes certain changes after injury that support regeneration of adult neurons. Such changes might include elimination or neutralization of growth inhibitors. (C) 1994 Wiley-Liss, Inc.

The expression of intermediate filaments is developmentally regulated. In the mammalian embryo keratins are the first to appear, followed by vimentin, while the principal intermediate filament of the adult brain is glial fibrillary acidic protein. The intermediate filaments expressed by a cell thus reflect its state of differentiation. The differentiation state of cells, and especially of glial cells, in turn determines their ability to support axonal growth. In this study we used three new antibodies directed against three fish intermediate filaments (glial fibrillary acidic protein, keratin 8 and vimentin), in order to determine the identity and level of expression of intermediate filaments present in fish glial cells in culture. We found that fish astrocytes and oligodendrocytes are both able to express keratin 8 and vimentin. We further demonstrate that under proliferative conditions astrocytes express high keratin 8 levels and most oligodendrocytes also express keratin 8, whereas under nonproliferative conditions the astrocytes express only low keratin 8 levels and most oligodendrocytes do not express keratin 8 at all. These results suggest that the fish glial cells retain characteristics of immature cells. The findings are also discussed in relation to the fish glial lineage.

The poor regenerative ability of neurons of the central nervous system in mammals, as compared with their counterpart in fish or amphibians, is thought to stem from differences in their immediate nonneuronal environment and its response to axonal injury. We describe one aspect of the environmental response to axonal injury in a spontaneously regenerating system-the fish optic nerve. The aspect under investigation was the reaction of glial cells at the injury site. This was examined by the use of antibodies that specifically recognize vimentin in fish glial cells. In the present study, affinity-purified vimentin antibodies were raised against a nonconserved N-terminal 14-amino acid peptide, which was predicted from the nucleotide sequence of vimentin. These antibodies were found to react specifically with glial cells in vitro. Moreover, the antivimentin antibodies stained both the optic nerve and the optic tract, but with different patterns. Specificity of the antibodies was verified by protein immunoblotting, tissue distribution, and labeling patterns. After injury, vimentin immunoreactivity initially disappeared from the site of the lesion due to cell death. Early signs of glial cell migration toward the injury site were evident a few days later. It is suggested that the reappearance of vimentin-positive glial cells at the site of injury is associated with axonal elongation across it, and that they contribute to the regenerative ability of the fish optic nerve. (C) 1994 Wiley-Liss, Inc.

The elements that control neuronal proliferation are largely unknown. Proliferating neurons in cultures of goldfish brain were studied in an attempt to identify the cell types involved. Neuronal proliferation was found to occur only when the neuronal stem cells were in direct contact with astrocytes, and never directly on the substrate. The regulation of neuronal proliferation thus appears to be mediated, at least in part, by contact with astrocytes. In addition, neurite extension was inhibited by medium conditioned by fish astrocytes. Since neurite extension and neuronal proliferation are mutually exclusive processes, inhibition of neurite extension by soluble substances derived from the astrocytes is probably one of the mechanisms controlling neuronal proliferation. The complex reciprocal relationship between neurons and astrocytes is also demonstrated by an observed inhibition of astrocytic proliferation by medium conditioned by differentiating fish neurons. This inhibition of astrocytic proliferation might be part of a mechanism through which interference with neuronal differentiation by astrocytes is avoided. The results of this study thus suggest that astrocytes, in addition to their known roles in controlling neuronal migration, neuronal differentiation and neurite elongation, may also play a role in the control of neuronal proliferation.

Regenerating optic nerves from fish produce a factor that is cytotoxic to oligodendrocytes. The cytotoxic factor is recognized by antibodies to interleukin-2 (IL-2) and has the apparent molecular size of a dimer of IL-2. An enzyme, identified as a nerve transglutaminase, was purified from regenerating optic nerves of fish and was found to catalyze dimerization of human IL-2. The dimerized IL-2, unlike monomeric IL-2, is cytotoxic to oligodendrocytes from rat brain in culture. The results suggest that posttranslational modification of a cytokine can alter its activity. Under conditions in which oligodendrocytes inhibit neuronal regeneration, dimerization of IL-2 might provide a mechanism to permit nerve growth.

1. The present review describes the results of a cloning that was successfully employed in the study of optic nerve regeneration in fish. 2. Three intermediate filaments (IFs) expressed by the glial fish optic nerve were cloned. 3. By the use of this approach it was possible to resolve the controversial question of whether glial fibrillary acidic protein (GFAP) is expressed in the fish optic nerve. 4. Moreover, as a result of the information that emerged from the cloning, it was possible to raise well-characterized monospecific antibodies and successfully exploit them in order to determine glial cell maturation, lineage and plasticity, monitor the glial cell response to injury of the fish optic nerve, and compare it to that of a non-regenerative system.

Axons of the central nervous system in adult mammals do not regenerate spontaneously after injury, partly because of the presence of oligodendrocytes that inhibit axonal growth. This is not the case in lower vertebrates (e.g., in fish), where regeneration of the optic nerve does occur spontaneously and has been correlated with the presence of factors cytotoxic to oligodendrocytes. The present study provides evidence that the substance originating from the fish optic nerves, which is cytotoxic to oligodendrocytes, is an interleukin 2-like substance.

Previous studies have suggested that L1, the cell adhesion molecule, is present in the regenerating fish optic nerve. The present study was undertaken in order to determine whether L1 is expressed by fish neurons and specifically by non-myelinated axons, using fish retinal explants in vitro and the developing fish visual system in vivo. In vitro, the nonmyelinated axons emerging from retinal explants showed L1 immunoreactivity and in vivo, L1 immunoreactive sites were found to be associated with areas rich in non-myelinated axons. At embryonic stage 23, as the eye developed and optic nerve axons began to elongate towards the tectum, L1-like immunoreactivity was seen both in optic nerve and in plexiform layers of the retina. At this stage and until hatching, the cellular layers within the retina showed little or no staining relative to the layers containing axons or dendrites. After hatching, L1 immunoreactivity was also observed in the ganglion cell layer, but upon maturation both the retina and the optic nerve lost most of their L1 immunoreactivity. We therefore suggest that non-myelinated axons of the fish visual system express L1 during development, which is lost after myelination and presumably reappears during regeneration.

Keywords: Biochemistry & Molecular Biology; Physiology

The high post-traumatic regenerative ability of fish central nervous system has been partially attributed to the hospitable nature of the surrounding non-neuronal cells and their appropriate response to injury. Uncovering the correlation between fish non-neuronal cell structure and behavior might yield a better understanding of what makes them supportive to axonal growth. Towards this goal, structural proteins expressed by fish non-neuronal cells need to be characterized. In the present study we isolated cDNA clones encoding fish intermediate filaments which are prominent structural proteins in astrocytes. Among the isolated clones, one was identified as fish vimentin and another was found identical to the cloned fish keratin 8. Results are discussed with respect to the use of these cDNAs for further understanding of fish non-neuronal cell plasticity.

The intermediate filament glial fibrillary acidic protein (GFAP) is the predominant cytoskeletal protein of mature glial cells in the mammalian nervous system. The nervous systems of lower vertebrates, such as fish, have been examined for the presence of GFAP and several investigators have shown that goldfish (Carassius auratus) brain contains GFAP-positive astrocytes. The same studies have demonstrated that, in contrast to the brain, the optic nerve of goldfish did not show any GFAP immunoreactivity, suggesting that this intermediate filament protein is not expressed in fish optic nerve astrocytes. The present study shows, however, that the monoclonal antibodies to porcine GFAP react with the optic nerve of carp (Cyprinus carpio), another member of the goldfish family. These antibodies to porcine GFAP cross react with rat brain and carp optic nerve, yielding a band of approximately 52 kDa in both species. Northern blot analysis using mouse GFAP DNA probe revealed that carp optic nerve RNA contains two transcripts, of 2.3 and 2.1 kb, which hybridize with the mouse GFAP probe. Injury to the carp optic nerve was followed by a decrease of GFAP immunoreactivity from neural tissue and a strong expression around blood vessels and connective tissues. On the basis of these observations and within the limitation of the techniques it is reasonable to conclude that the carp optic nerve expresses GFAP immunoreactivity and that the pattern of expression of this intermediate filament protein is altered after injury. Such an alteration might be relevant to the process of regeneration.

In the present study, we have developed an animal model for central nervous system (CNS) damage using the graded crush injury of the adult rat optic nerve. This injury model has the following characteristics: (a) the injuries are reproducible; (b) the injuries can be created with controlled severity; and (c) variations in the severity of the injuries correlate with definable electrophysiological and behavioral outcomes. Self-closing forceps, modified by attaching a screw to the handle, were used as a lesion-causing device. Rat optic nerves were crushed in vitro. The severity of the injury, calibrated by a measured applied force, was determined, electrophysiologically, in vitro. Compound action potential was assessed and recorded by suction electrodes connected to both sides of the nerve. In vivo studies correlated the calibrated crush injuries with behavioral outcome. After severe crush (applied force: 205 g or higher), the rats were unable to orient towards visual stimuli presented in the visual field of the damaged nerve and only minor transient recovery was observed. After moderate crush (applied force: 120 g), however, there was a significant recovery of orienting performance up to the 6th postoperative day, followed by a gradual, secondary loss of function. In the mildly injured group (applied force lower than 30 g), functional improvement continued for up to 10 days and remained stable thereafter, with only minor secondary loss. This CNS graded injury model may be valuable to study the molecular and anatomical mechanisms underlying secondary degeneration and the potency of various posttraumatic treatments in leading to recovery of function.