Effects of Brain-Derived IL-2 Deficiency and the Development of Autoimmunity on Spatial Learning and Fear Conditioning.

Interleukin-2 (IL-2) has been implicated in neurological disorders including multiple sclerosis and Alzheimer's disease. Peripheral IL-2 deficiency in gene-deleted mice results in T cell mediated autoimmunity that begins to develop slowly after weaning and progressively increases through adulthood. Loss of brain-derived IL-2 results in neurobiological and behavioral abnormalities, and may contribute to the development of CNS autoimmunity by modifying the neuroimmunological milieu of the brain. We have shown previously that IL-2 knockout (KO) mice have altered learning acquisition in the Morris water-maze. Hypothesizing that the learning acquisition deficits in IL-2KO would be associated largely with the loss of brain-derived IL-2, the present study sought to determine if these cognitive alterations are due to the loss the IL-2 gene in the brain and/or autoimmunity resulting from loss of the gene in the peripheral immune system. We found that SCID congenic mice (mice free of IL-2 deficiency induced peripheral autoimmunity) without brain IL-2 (two IL-2KO alleles) did not differ from SCID congenic mice with normal brain IL-2 (two WT IL-2 alleles); thus, contrary to our hypothesis, loss of brain-derived IL-2 did not affect learning acquisition in the water-maze. Compared to adult WT littermates (9 weeks), adult IL-2KO mice with autoimmunity exhibited alterations in learning acquisition in the Morris water-maze whereas younger pre-autoimmune IL-2KO mice (5 weeks) had performance comparable to younger WT littermates, suggesting that the water-maze learning deficits in IL-2KO mice were associated with the development of peripheral autoimmunity. As IL-2KO mice have cytoarchitectural alterations in the dentate gyrus, circuitry involved in the differentiation of contexts (versus places), we also compared IL-2KO mice and littermates in a contextual fear discrimination paradigm. IL-2KO mice were found to have reduced conditioned fear discrimination that was not related to age-associated autoimmunity. Together, these findings suggest that complex interactions between IL-2 deficiency in the brain and immune system may modify brain processes involved in different modalities of learning and memory.

Loss of Neuronal Phenotype and Neurodegeneration: Effects of T Lymphocytes and Brain Interleukin-2.

Loss of neuronal phenotype and reversal of neuronal atrophy have been demonstrated in different models of central nervous system (CNS) injury. These processes may be generalizable to different types of brain neurons and circuitry. The idea that some injured neurons may lose their phenotype and/or atrophy with the potential to rejuvenate is a remarkable and potentially promising form of neuronal plasticity that is not well understood. In this paper, we present some of our laboratory's basic neuroimmunology research showing that peripheral T cells entering the CNS, and brain-derived interleukin-2 (IL-2), play significant roles in these intriguing processes. Our findings suggest, for example, that T cell immunosenesence could be involved in related processes of brain aging and contribute to neurodegenerative disease. Neuroimmunological approaches may provide new insights into yet undiscovered factors and brain mechanisms that regulate changes in neuronal integrity associated with aging and disease. Such findings could have important implications for discovering more effective strategies for treating patients with neurotrauma and neurodegenerative diseases (e.g., Alzheimer's disease).

Motor Neurons Exhibit Sustained Loss of Atrophy Reversal in Immunodeficent Mice.

Our lab showed previously that whereas a substantial portion of chronically resected facial motor neurons reside in an atrophied state that can be reversed at 14 days following reinjury in wild-type (WT) mice, atrophy reversal was altered in immunodeficient mice. It was unclear, however, if the abnormal response at day 14 post-reinjury in immunodeficient mice might be due to differences in the kinetics of the reversal response or impaired regeneration. We sought to address this question, and test our working hypothesis that the normal regeneration of atrophied motor neurons is dependent on normal adaptive immunity, by comparing WT and immunodeficient recombination activating gene-2 knockout (RAG2-KO) mice that lack a mature T and B lymphocytes, at 3 and 28 days following reinjury. In WT mice, facial motor neurons that were resected for 10 weeks and subsequently reinjured for 3 days were able to regain fully an apparent 40% loss of countable neurons, and nearly 45% of that robust increase in neurons was sustained at 28 days post-reinjury in the WT mice. By contrast, at both 3 and 28 days post-reinjury RAG2-KO mice failed to show any increase in neuronal number. Size measurements showed that the surviving neurons of WT and RAG2-KO mice exhibited substantial motor neuron hypertrophy at 3 days post-reinjury, and similar levels of normal size motor neurons by 28 days post-reinjury. Among the WT mice, small numbers of T lymphocytes where found in the reinjured facial motor nucleus (FMN), and were significantly higher at 3 days, but not 28 days, in the reinjury compared to sham-reinjury groups. No differences were seen between the WT and RAG2-KO mice in overall microglial cell activity using CD11b expression following reinjury. These data suggest that many resected motor neurons did not survive the initial resection in RAG2-KO mice, whereas in WT mice they atrophied and could be restimulated by reinjury to regenerate their phenotype. Moreover, they indicate that normal T cell function, or some yet unknown function of the RAG2 gene in the brain, is essential for activating regeneration programs of atrophied motor neurons - programs with therapeutic potential for modifying neuroplasticity.

Loss of cholinergic phenotype in septohippocampal projection neurons: relation to brain versus peripheral IL-2 deficiency.

In the peripheral immune system, IL-2 is essential for immune homeostasis, normal T regulatory cell function, and self-tolerance. IL-2 knockout (IL-2KO) mice develop spontaneous autoimmunity characterized by increased T cell trafficking to multiple organs. The IL-2 gene is also expressed in the brain, and in vitro studies have shown that IL-2 is a potent modulator of acetylcholine release from septohippocampal neurons and exerts trophic effects on septal neurons in culture. We previously described the apparent loss of cholinergic cell bodies in the medial septum of IL-2KO mice. Here we investigated if loss of brain-derived IL-2, or autoimmunity stemming from loss of peripheral IL-2, is responsible for the alteration in choline acetyltransferase (ChAT) expression in the medial septum of IL-2KO mice. To accomplish this objective, we compared ChAT-positive neurons between wild-type (WT) mice, IL-2KO mice, and congenic mice with a double gene deletion for the IL-2 gene and the recombinase activating gene-2 (RAG-2) which are referred to as IL-2KO/RAG-2KO mice (congenic mice which lack mature T and B cells as well as peripheral and brain-derived IL-2). We found that the loss of ChAT staining did not coincide with an overall loss of cells in the medial septum, suggesting that loss of brain IL-2 results in a change in cholinergic phenotype unrelated to cell death. No differences were noted in the endogenous expression of cytokines and chemokines tested in the medial septum. Evaluation of BDNF and NGF levels between WT and IL-2KO mice in medial septal homogenates revealed that IL-2KO mice have markedly higher levels of NGF in the medial septum compared to WT mice. Our findings suggest that brain-derived IL-2 plays an essential role in the maintainance of septohippocampal projection neurons in vivo.

Selective Neuronal and Brain Regional Expession of IL-2 in IL2P 8-GFP Transgenic Mice: Relation to Sensorimotor Gating.

Brain-derived interleukin-2 (IL-2) has been implicated in diseases processes that arise during CNS development (e.g., autism) to neurodegenerative alterations involving neuroinflammation (e.g., Alzheimer's disease). Progress has been limited, however, because the vast majority of current knowledge of IL-2's actions on brain function and behavior is based on the use exogenously administered IL-2 to make inferences about the function of the endogenous cytokine. Thus, to identify the cell-type(s) and regional circuitry that express brain-derived IL-2, we used B6.Cg-Tg/ IL2-EGFP17Evr (IL2p8-GFP) transgenic mice, which express green fluorescent protein (GFP) in peripheral immune cells known to produce IL-2. We found that the IL2-GFP transgene was localized almost exclusively to NeuN-positive cells, indicating that the IL-2 is produced primarily by neurons. The IL2-GFP transgene was expressed in discrete nuclei throughout the rostral-caudal extent of the brain and brainstem, with the highest levels found in the cingulate, dorsal endopiriform nucleus, lateral septum, nucleus of the solitary tract, magnocellular/gigantocellular reticular formation, red nucleus, entorhinal cortex, mammilary bodies, cerebellar fastigial nucleus, and posterior interposed nucleus. Having identified IL-2 gene expression in brain regions associated with the regulation of sensorimotor gating (e.g., lateral septum, dorsal endopiriform nucleus, entorhinal cortex, striatum), we compared prepulse inhibition (PPI) of the acoustic startle response in congenic mice bred in our lab that have selective loss of the IL-2 gene in the brain versus the peripheral immune system, to test the hypothesis that brain-derived IL-2 plays a role in modulating PPI. We found that congenic mice devoid of IL-2 gene expression in both the brain and the peripheral immune system, exhibited a modest alteration of PPI. These finding suggest that IL2p8-GFP transgenic mice may be a useful tool to elucidate further the role of brain-derived IL-2 in normal CNS function and disease.

5. T cell immunity and neuroplasticity.

The proneuronal effects of T cells that impact the brain occur from both T cells trafficking into the brain, and from signals in the periphery (e.g., cytokine release and regulation). Recent data indicates that neuroimmunological changes in the brain can modify intrinsic brain processes that are involved in regulating neuroplasticity (e.g., T-cell/microglial interactions, neurotrophins, neurogenesis). We describe: 1) work from our lab and others showing that injury-induced loss of neuronal phenotype and reversal of motor neuron atrophy are associated with normal T cell immunity, and; 2) research indicating that these and other neuroimmunological processes may be generalizable to mechanisms of neuroplasticity involved in cognitive and emotional behavior. These findings are discussed in relation to our lab's working hypothesis, that T cell immunosenesence may contribute to alterations in brain neuroplasticity related to aging. Greater understanding of the role of adaptive T cell immunity on neuroplasticity could have important clinical implications for developing novel treatment strategies for neurodegenerative diseases (e.g., Alzheimer's) and brain injury (e.g., stroke, trauma).

Interleukin-2 and the brain: dissecting central versus peripheral contributions using unique mouse models.

Although many studies have documented peripheral immune alterations in patients with psychiatric and neurological disorders, almost all these data in humans are correlative. The actions of IL-2 on neurodevelopment, function, and disease are the result of both IL-2's actions in the peripheral immune system and intrinsic actions in the CNS. Determining if, and under what conditions (e.g., development, acute injury) these different actions of IL-2 are operative in the brain is essential to make advances in understanding the multifaceted affects of IL-2 on CNS function and disease. Mouse models have provided ways to obtain new insights into how the complex biology of a cytokine such as IL-2 can have simultaneous, dynamic effects on multiple systems (e.g., regulating homeostasis in the brain and immune system, autoimmunity that can affect both systems). Here we describe some of the relevant literature and our research using different mouse models. This includes models such as congenic IL-2 knockout mice bred on immunodeficient backgrounds coupled with immune reconstitution strategies used to dissect neuroimmunological processes involved in the development of septohippocampal pathology, and test the hypothesis that dysregulation of the brain's endogenous neuroimmunological milieu may occur with the loss of brain IL-2 gene expression and be involved in initiating CNS autoimmunity. Use of animal models like these in the field of psychoneuroimmunology may lead to critical advances into our understanding of the role of brain cytokines and autoimmunity in neurodegenerative diseases (e.g., Alzheimer's disease), neurodevelopmental disorder (e.g., autism, schizophrenia), and autoimmune diseases including multiple sclerosis.

Dissecting the effects of endogenous brain IL-2 and normal versus autoreactive T lymphocytes on microglial responsiveness and T cell trafficking in response to axonal injury.

IL-2 is essential for T-helper regulatory (Treg) cell function and self-tolerance, and dysregulation of both endogenous brain and peripheral IL-2 gene expression may have important implications for neuronal injury and repair. We used an experimental approach combining mouse congenic breeding and immune reconstitution to test the hypothesis that the response of motoneurons to injury is modulated by the combined effects of IL2-mediated processes in the brain that modulate its endogenous neuroimmunological milieu, and IL2-mediated processes in the peripheral immune system that regulate T cell function (i.e., normal versus autoreactive Treg-deficient T cells). This experimental strategy enabled us to test our hypothesis by disentangling the effect of normal versus autoreactive T lymphocytes from the effect of endogenous brain IL-2 on microglial responsiveness (microglial phagocytic clusters normally associated with dead motoneurons and MHC2(+) activated microglia) and T cell trafficking, using the facial nerve axotomy model of injury. The results demonstrate that the loss of both brain and peripheral IL-2 had an additive effect on numbers of microglial phagocytic clusters at day 14 following injury, whereas the autoreactive status of peripheral T cells was the primary factor that determined the degree to which T cells entered the injured brain and contributed to increased microglial phagocytic clusters. Changes in activated MHC2(+) microglial in the injured FMN were associated with loss of endogenous brain IL-2 and/or peripheral IL-2. This model may provide greater understanding of the mechanisms involved in determining if T cells entering the injured central nervous system (CNS) have damaging or proregenerative effects.

Interleukin-2 and the septohippocampal system: intrinsic actions and autoimmune processes relevant to neuropsychiatric disorders.

The effects of IL-2 on brain development, function, and disease are the result of IL-2's actions in the peripheral immune system and its intrinsic actions in the central nervous system (CNS). Determining whether, and under what circumstances (e.g., development, acute injury), these different actions of IL-2 are operative in the brain is essential to make significant advances in understanding the multifaceted affects of IL-2 on CNS function and disease, including psychiatric disorders. For several decades, there has been a great deal of speculation about the role of autoimmunity in brain disease. More recently, we have learned a great deal about the role of cytokines on neurobiological processes, and there have been many studies that have found peripheral immune alterations in patients with neurological and neuropsychiatric diseases. Despite a plethora of published literature, almost all of this data in humans is correlative and much of the basic research has understandably relied on simpler models (e.g., in vitro models). Good animal models such as our IL-2 knockout mouse model could provide valuable new insight into understanding how the complex biology of a cytokine such as IL-2 can have simultaneous, dynamic effects on multiple systems (e.g., regulating homeostasis in the brain and immune system, autoimmunity that can affect both systems). Animal models can also provide much needed new data elucidating neuroimmunological and autoimmune processes involved in brain development and disease. Such information may ultimately provide critical new insight into the role of brain cytokines and autoimmunity in prominent neurological and neuropsychiatric diseases (e.g., Alzheimer's disease, autism, multiple sclerosis, schizophrenia).

Loss of CNS IL-2 gene expression modifies brain T lymphocyte trafficking: response of normal versus autoreactive Treg-deficient T cells.

Emerging data from our lab and others suggested that dysregulation of the brain's endogenous neuroimmunological milieu may occur with the loss of brain IL-2 gene expression and be involved in initiating processes that lead to CNS autoimmunity. We sought to test our working hypothesis that IL-2 deficiency induces endogenous changes in the CNS that play a key role in eliciting T cell homing into the brain. To accomplish this goal, we used an experimental approach that combined mouse congenic breeding and immune reconstitution. In congenic mice without brain IL-2 (two IL-2 KO alleles) that were reconstituted with a normal wild-type immune system, the loss of brain IL-2 doubled the number of T cells that trafficked into the brain in all regions quantified (hippocampus, septum, and cerebellum) compared to mice with two wild-type brain IL-2 alleles and a wild-type peripheral immune system. Congenic mice with normal brain IL-2 (two wild-type IL-2 alleles) that were immune reconstituted with autoreactive Treg-deficient T cells from IL-2 KO mice developed the expected peripheral autoimmunity (splenomegaly) and had a comparable doubling of T cell trafficking into the hippocampus and septum, whereas they exhibited an additional twofold proclivity for the cerebellum over the septohippocampal regions. Unlike brain trafficking of wild-type T cells, the increased homing of IL-2 KO T cells to the cerebellum was independent of brain IL-2 gene expression. These findings demonstrate that brain IL-2 deficiency induces endogenous CNS changes that may lead to the development of brain autoimmunity, and that autoreactive Treg-deficient IL-2 KO T cells trafficking to the brain could have a proclivity to induce cerebellar neuropathology.

Age and facial nerve axotomy-induced T cell trafficking: relation to microglial and motor neuron status.

Following peripheral axotomy of the facial nerve in mice, T lymphocytes cross the blood-brain-barrier (BBB) into the central nervous system (CNS), where they home to the neuronal cell bodies of origin in the facial motor nucleus (FMN) and act in concert with microglial cells to support the injured motor neurons. Several lines of evidence suggested normal aging may alter the injury-related responses of T cells, microglia, and motor neurons in this model. In this study, we therefore sought to test the hypothesis that compared to 8-week-old mice (young adult), 52-week-old mice (advanced middle age) would exhibit more neuronal damage and increased T cell trafficking into the injured FMN following facial nerve resection. Comparison of 8- and 52-week-old mice at 7, 14, 21, and 28 days post-resection of the facial nerve, confirmed our hypothesis that age influences the kinetics of CD3(+) T lymphocyte trafficking in the axotomized FMN. The peak T cell response was significantly higher, occurred later, and remained elevated longer in the injured FMN of mice in the 52 week age group. Although the kinetics of motor neuron death (identified by quantifying CD11b(+) perineuronal microglial phagocytic clusters engulfing the dead neurons at 7, 14, 21, and 28 days post-resection) differed between the age groups, motor neuron profile counts at day 28 showed that levels of cumulative motor neuron loss did not differ between the age groups. Compared to 8-week-old mice, however, there was small reduction in the mean cell size of the surviving motor neurons in the 52 week age group. Since T lymphocyte function decreases with normal aging, it will be important to determine if increased T cell trafficking into the injured CNS is a compensatory response to the decreased function of older T cells, and if these and related neuroimmunological changes are more pronounced in mice in the late stages of the life cycle.

Interleukin-2 deficiency-induced T cell autoimmunity in the mouse brain.

Interleukin-2 (IL-2) has been implicated in the pathogenesis of neurodevelopmental and neurodegenerative disorders. Studies from our lab have shown that adult IL-2 knockout (KO) mice exhibit septohippocampal pathology and related behavioral deficits. Compared to IL-2 wild-type (WT) mice, IL-2 KO mice have a marked and selective loss of septal cholinergic neurons that occurs between the third postnatal week and adulthood. Given that the development of septal neurons is completed by embryonic day 17 and that IL-2 KO mice exhibit peripheral autoimmunity that develops progressively post-weaning, our data and others led us to postulate that the loss of septal neurons in adult IL-2 KO mice is due to selective autoimmune neurodegeneration that coincides with increasing levels of peripheral autoimmunity. Thus, the present study tested the hypotheses: (1) that T cells selectively target the septum, and; (2) that T lymphocyte infiltration to the septum would correlate with peripheral autoimmune disease. We quantified CD3(+) T cells in the septum, hippocampus, and cerebellum of IL-2 KO and IL-2 WT mice at ages ranging from 2 to 14 weeks. T cells infiltrated the brains of IL-2 deficient mice, but were not selective for the septum. Brain T lymphocyte levels in IL-2 KO mice correlated positively with the degree of peripheral autoimmunity. We did not detect CD19(+) B lymphocytes, IgG-positive lymphocytes or IgG deposition indicative of autoantibodies in the brains of IL-2 KO mice. Further study is needed to understand how IL-2 deficiency-induced autoimmune T lymphocytes interact with endogenous brain cells to alter function and promote disease.

T cell memory in the injured facial motor nucleus: relation to functional recovery following facial nerve crush.

T cells have the ability to mount a memory response to a previously encountered antigen such that re-exposure to the antigen results in a response that is greater in magnitude and function. Following facial nerve transection, T cells have been shown to traffic to injured motor neurons in the facial motor nucleus (FMN) and may have the ability to promote neuronal survival and functional recovery. Previously, we demonstrated that early exposure to neuronal injury on one side of the brain during young adulthood elicited a T cell response that was greater in magnitude following exposure to the same form of injury on the contralateral side later in adulthood. Whether the T cell memory response to neuronal injury influenced functional recovery following nerve crush injury was unknown. In the current study, we tested the hypotheses that (1) transection of the right facial nerve in sensitized mice would result in faster recovery of the whisker response when the contralateral facial nerve is crushed 10 weeks later, and (2) the early recovery would be associated with an increase in the magnitude of the T cell response in the contralateral FMN following crush injury in sensitized mice. The onset of modest recovery in sensitized mice occurred between 3 and 5 days following crush injury of the contralateral facial nerve, approximately 1.5 days earlier than naïve mice, and was associated with more than a two-fold increase in the magnitude of the T cell response in the contralateral FMN following crush injury. There was no difference between groups in the number of days to full recovery. Further study of how T cell memory influences neuroregeneration may have important implications for translational research.

Influence of injury severity on the rate and magnitude of the T lymphocyte and neuronal response to facial nerve axotomy.

The temporal relationship between severity of peripheral axonal injury and T lymphocyte trafficking to the neuronal cell bodies of origin in the brain has been unclear. We sought to test the hypothesis that greater neuronal death induced by disparate forms of peripheral nerve injury would result in differential patterns of T cell infiltration and duration at the cell bodies of origin in the brain and that these measures would correlate with the magnitude of neuronal death over time and cumulative neuronal loss. To test this hypothesis, we compared the time course of CD3(+) T cell infiltration and neuronal death (assessed by CD11b(+) perineuronal microglial phagocytic clusters) following axonal crush versus axonal resection injuries, two extreme variations of facial nerve axotomy that result in mild versus severe neuronal loss, respectively, in the facial motor nucleus. We also quantified the number of facial motor neurons present at 49 days post-injury to determine whether differences in the levels of neuronal death between nerve crush and resection correlated with differences in cumulative neuronal loss. Between 1 and 7 days post-injury when levels of neuronal death were minimal, we found that the rate of accumulation and magnitude of the T cell response was similar following nerve crush and resection. Differences in the T cell response were apparent by 14 days post-injury when the level of neuronal death following resection was substantially greater than that seen in crush injury. For nerve resection, the peak of neuronal death at 14 days post-resection was followed by a maximal T cell response one week later at 21 days. Differences in the level of neuronal death between the two injuries across the time course tested reflected differences in cumulative neuronal loss at 49 days post-injury. Altogether, these data suggest that the trafficking of T cells to the injured FMN is dependent upon the severity of peripheral nerve injury and associated neuronal death.

Psychopharmacology in HIV-infected patients.

Neuropsychiatric disorders and syndromes may be underdiagnosed and inadequately treated in individuals infected with HIV. Depression in particular is among the most prevalent diagnoses, and data from controlled clinical studies have shown that antidepressant medications are efficacious and safe for treating depression in HIV-infected persons. A significant shortcoming of this literature is that most of the available data are from studies conducted before the advent of highly active antiretroviral therapy. In addition, apart from antidepressant medications, controlled studies systematically assessing efficacy and safety issues for other classes of psychotropic drugs (e.g., antipsychotic and anxiolytic medications) in HIV-infected persons are lacking. This review summarizes essential findings pertaining to the use of psychotropic medications to treat depression and other neuropsychiatric disorders in the context of HIV. It includes a discussion of clinically relevant treatment considerations (e.g., side effects, drug-drug interactions) derived from the existing literature as well as judgments that clinicians face in the absence of research data. Despite some shortcomings of the existing literature, overall there is compelling evidence that the appropriate use of psychotropic medications (coupled with behavioral therapy) can improve the quality of life of mentally ill HIV-infected individuals.

Immunodeficiency impairs re-injury induced reversal of neuronal atrophy: relation to T cell subsets and microglia.

Following facial nerve resection in the mouse, a substantial number of neurons reside in an atrophied state (characterized by cell shrinkage and decreased ability to uptake Nissl stain), which can be reversed by re-injury. The mechanisms mediating the reversal of neuronal atrophy remain unclear. Although T cells have been shown to prevent neuronal loss following peripheral nerve injury, it was unknown whether T cells play a role in mediating the reversal of axotomy-induced neuronal atrophy. Thus, we used a facial nerve re-injury model to test the hypothesis that the reversal of neuronal atrophy would be impaired in recombinase activating gene-2 knockout (RAG-2 KO) mice, which lack functional T and B cells. Measures of neuronal survival were compared in the injured facial motor nucleus (FMN) of RAG-2 KO and wild-type (WT) mice that received a resection of the right facial nerve followed by re-injury of the same nerve 10 weeks later ("chronic resection+re-injury") or a resection of the right facial nerve followed by sham re-injury of the same nerve 10 weeks later ("chronic resection+sham"). We recently demonstrated that prior exposure to neuronal injury elicited a marked increase in T cell trafficking indicative of a T cell memory response when the contralateral FMN was injured later in adulthood. We examined if such a T cell memory response would also occur in the current re-injury model. RAG-2 KO mice showed no reversal of neuronal atrophy whereas WT mice showed a robust response. The reversal of atrophy in WT mice was not accompanied by a T cell memory response. Although the number of CD4(+) and CD8(+) T cells in the injured FMN did not differ from each other, double-negative T cells appear to be recruited in response to neuronal injury. Re-injury did not result in increased expression of MHC2 by microglia. Our findings suggest that T cells may be involved in reversing the axotomy-induced atrophy of injured neurons.

IL-15 and IL-15R alpha gene deletion: effects on T lymphocyte trafficking and the microglial and neuronal responses to facial nerve axotomy.

IL-15 is a potent T cell chemoattractant, and this cytokine and its unique alpha subunits, IL-15R alpha, can modify immune cell expression of several T cell chemokines and their receptors. Facial nerve axotomy in mice leads to T cell migration across an intact blood-brain-barrier (BBB), and under certain conditions T cells can provide neuroprotection to injured neurons in the facial motor nucleus (FMN). Although chemokines and chemoattractant cytokines are thought to be responsible for T cell migration to the injured cell bodies, data addressing this question are lacking. This study tested the hypothesis that T cell homing to the axotomized FMN would be impaired in knockout (KO) mice with the IL-15 and IL-15R alpha genes deleted, and sought to determine if microglial responsiveness and motoneuron death are affected. Both IL-15KO and IL-15R alpha KO mice exhibited a marked reduction in CD3(+) T cells and had fewer MHC2(+) activated microglia in the injured FMN than their respective WT controls at day 14 post-axotomy. Although there was a relative absence of T cell recruitment into the axotomized FMN in both knockout strains, IL-15R alpha KO mice had five times more motoneuron death (characterized by perineuronal microglial clusters engulfing dead motoneurons) than their WT controls, whereas dead neurons in IL-15KO did not differ from their WT controls. Further studies are needed to dissect the mechanisms that underlie these observations (e.g., central vs. peripheral immune contributions).

Prior facial motor neuron injury elicits endogenous T cell memory: relation to neuroregeneration.

We tested the hypotheses that prior injury to the facial motor nucleus (FMN) would elicit a more robust T cell response in the opposite FMN when the contralateral facial nerve was injured later in life, and that this would result in improved neuroregeneration. Measures of T cell, neuronal and microglial status were compared in sensitized mice (right facial nerve transection followed by contralateral facial nerve transection 9.5 weeks later) and naïve mice (sham surgery of the right facial nerve followed by contralateral facial nerve transection 9.5 weeks later) following axotomy of the contralateral facial nerve. At day 14 post-axotomy, sensitized mice exhibited nearly a two-fold increase in T cells in the FMN compared to naïve mice. There were no differences between the groups in levels of dead neurons and NeuN expression by surviving motor neurons at day 14, or motor neuron survival and cell area at day 49 post-axotomy. Measures of microglial responsiveness did not differ between the groups. Further study is needed to delineate the role of endogenous T cell memory in neuronal injury and regeneration.

Endogenous T lymphocytes and microglial reactivity in the axotomized facial motor nucleus of mice: effect of genetic background and the RAG2 gene.

Following facial nerve axotomy in mice, peripheral T cells home to the injured facial motor nucleus (FMN) where they may influence the glial response. Interactions between T cells and microglia, which proliferate in response to axotomy, appear to confer neuroprotection to injured motoneurons. The primary objective of this study was to determine whether T lymphocytes could influence the microglial reaction to motoneuron injury. These experiments tested the hypotheses that (1) C57BL/6 (B6) and 129 mice, inbred strains which have high and low levels of astroglial reactivity in the axotomized FMN, respectively, would also exhibit high and low levels of T cell infiltration, and (2) that these differences would correspond with levels of microglial reactivity and neuronal regeneration. Thus, we compared the response to facial nerve axotomy in B6, 129, and immunodeficient RAG2 knockout (RAG2 KO) mice on these two backgrounds at 14 day post-axotomy for differences in levels of 1) CD3+ T cell infiltration; (2) major histocompatibility complex II (MHC2) expression by microglia; (3) perineuronal microglial phagocytic clusters, an indirect measure of neuronal death; and (4) overall microglial activity as assessed by CD11b expression. To examine the inheritance pattern of the abovementioned neuroimmune measures, we also made assessments in B6x129 F1 generation mice. B6 and 129 mice displayed high and low levels of T cell infiltration to the affected FMN and low and high MHC2 expression, respectively. Levels of microglial activity did not differ between the two strains. In immunodeficient RAG2 KO mice on both backgrounds, the number of MHC2+ microglia did not differ from their immunologically normal background controls. Moreover, deletion of either the RAG2 or RAG1 genes in B6 mice was not associated with increased neuronal death at day 14 post-axotomy, as we had previously found in B6 mice with the severe combined immunodeficiency (SCID) mutation. Contrary to our hypothesis, the paucity of T cells in the affected FMN of the 129 mice was associated with less neuronal death when compared to B6 mice, which showed a robust T cell response. Moreover, the data suggest that parameters of the central and peripheral immune responses to axotomy are independently regulated. Assessments in B6x129 F1 generation mice revealed dominant phenotypes for both T cell infiltration and neurodegeneration, whereas both strains contributed significantly to the phenotype for MHC2 expression. Our findings suggest that (1) T cells do not appear to modify measures of microglial reactivity in the axotomized FMN; and (2) the impact of T cells on injured motoneurons in immunologically intact mice and in immunodeficient mice grafted with T cells by adoptive transfer may be different. Further study is required to understand the role of T cells following motoneuron injury in immunologically intact mice and how the seemingly divergent effects of T cells in intact and immunodeficient mice might provide insight into their role in neuronal injury and repair.