Antigen-specific T cell trafficking into the central nervous system

The initiation step of cell-mediated immune responses in the central nervous system (CNS) involves the trafficking of the antigen-specific T cells into the brain. To study this trafficking, we developed an in vivo system for studying antigen-specific responses in the CNS. In this assay, T cell receptor (TCR) transgenic mice having 95% of T cells specific for a defined antigen-pigeon cytochrome c (PCC) were cannulated intraventricularly for PCC antigen infusion and cerebrospinal fluid (CSF) sampling. Upon PCC infusion into the CNS, the number of alpha/beta TCR(+) Vbeta3(+) Mac1(-) cells in the CSF was characterized. We found that infusion of antigen into the CSF induced an increased number of antigen-specific T cells in the CNS and activation of antigen-specific T cells in the peripheral blood. Hence, the drainage of CNS antigen into the periphery might play an important role in sustaining autoimmune reactivity in CNS inflammatory diseases.     

Stroke affects intestinal immune cell trafficking to the central nervous system

Stroke is an acute neurological disease with a strong inflammatory component that can be regulated by the intestinal microbiota and intestinal immune cells. Although stroke has been shown to alter immune cell populations in the gut, the dynamics of cell trafficking have not been elucidated. To study the trafficking of gut-derived immune cells after stroke, we used mice expressing the photoconvertible protein Kikume Green-Red, which turns form green to red when exposed to violet light. Mice underwent laparotomy and the small intestine was exposed to violet laser light. Immune cells were isolated from the small intestine immediately after photoconversion and 2 days later. Percentage of immune cells (CD45⁺KikR⁺) that expressed the red variant of the protein (KikR) was higher immediately after photoconversion than 2 days later, indicating cell egress from the small intestine. To investigate whether intestinal immune cells traffic to the periphery and/or the central nervous system (CNS) after stroke, we analyzed KikR⁺ immune cells (2 days after photoconversion) in peripheral lymphoid organs, meninges and brain, 3 and 14 days after transient occlusion of the middle cerebral artery (tMCAo) or sham-surgery. Although migration was observed in naïve and sham animals, stroke induced a higher mobilization of gut KikR⁺ immune cells, especially at 3 days after stroke, to all the organs analyzed. Notably, we detected a significant migration of CD45^hi immune cells from the gut to the brain and meninges at 3 days after stroke. Comparison of cell trafficking between organs revealed a significant preference of intestinal CD11c⁺ cells to migrate from the small intestine to brain and meninges after stroke. We conclude that stroke increases immune cell trafficking from the small intestine to peripheral lymphoid organs and the CNS where they might contribute to post-stroke inflammation.     

Effect of morphine and SIV on dendritic cell trafficking into the central nervous system of rhesus macaques

Recruitment of immune cells such as monocytes/macrophages and dendritic cells (DCs) across the blood–brain barrier (BBB) has been documented in diseases involving neuroinflammation. Neuroinvasion by HIV leads to neurocognitive diseases and alters the permeability of the BBB. Likewise, many HIV patients use drugs of abuse such as morphine, which can further compromise the BBB. While the role of monocytes and macrophages in neuroAIDS is well established, research demonstrating the presence and role of DCs in the CNS during HIV infection has not been developed yet. In this respect, this study explored the presence of DCs in the brain parenchyma of rhesus macaques infected with a neurovirulent form of SIV (SIV mac239 R71/17E) and administered with morphine. Cells positive for DC markers including CD11c (integrin), macDC-SIGN (dendritic cell-specific ICAM-3 grabbing nonintegrin), CD83 (a maturation factor), and HLA-DR (MHC class II) were consistently found in the brain parenchyma of SIV-infected macaques as well as infected macaques on morphine. Control animals did not exhibit any DC presence in their brains. These results provide first evidence of DCs’ relevance in NeuroAIDS vis-à-vis drugs of abuse and open new avenues of understanding and investigative HIV-CNS inflictions.     

A dynamic in vitro BBB model for the study of immune cell trafficking into the central nervous system

Although there is significant evidence correlating overreacting or perhaps misguided immune cells and the blood–brain barrier (BBB) with the pathogenesis of neuroinflammatory diseases, the mechanisms by which they enter the brain are largely unknown. For this purpose, we revised our humanized dynamic in vitro BBB model (DIV-BBBr) to incorporate modified hollow fibers that now feature transmural microholes (2 to 4 μm ∅) allowing for the transendothelial trafficking of immune cells. As with the original model, this new DIV-BBBr reproduces most of the physiological characteristics of the BBB in vivo. Measurements of transendothelial electrical resistance (TEER), sucrose permeability, and BBB integrity during reversible osmotic disruption with mannitol (1.6 mol/L) showed that the microholes do not hamper the formation of a tight functional barrier. The in vivo rank permeability order of sucrose, phenytoin, and diazepam was successfully reproduced in vitro. Flow cessation followed by reperfusion (Fc/Rp) in the presence of circulating monocytes caused a biphasic BBB opening paralleled by a significant increase of proinflammatory cytokines and activated matrix metalloproteinases. We also observed abluminal extravasation of monocytes but only when the BBB was breached. In conclusion, the DIV-BBBr represents the most realistic in vitro system to study the immune cell trafficking across the BBB.     

Inhibition of Piezo1 attenuates demyelination in the central nervous system

Piezo1 is a mechanosensitive ion channel that facilitates the translation of extracellular mechanical cues to intracellular molecular signaling cascades through a process termed, mechanotransduction. In the central nervous system (CNS), mechanically gated ion channels are important regulators of neurodevelopmental processes such as axon guidance, neural stem cell differentiation, and myelination of axons by oligodendrocytes. Here, we present evidence that pharmacologically mediated overactivation of Piezo1 channels negatively regulates CNS myelination. Moreover, we found that the peptide GsMTx4, an antagonist of mechanosensitive cation channels such as Piezo1, is neuroprotective and prevents chemically induced demyelination. In contrast, the positive modulator of Piezo1 channel opening, Yoda-1, induces demyelination and neuronal damage. Using an ex vivo murine-derived organotypic cerebellar slice culture model, we demonstrate that GsMTx4 attenuates demyelination induced by the cytotoxic lipid, psychosine. Importantly, we confirmed the potential therapeutic effects of GsMTx4 peptide in vivo by co-administering it with lysophosphatidylcholine (LPC), via stereotactic injection, into the cerebral cortex of adult mice. GsMTx4 prevented both demyelination and neuronal damage usually caused by the intracortical injection of LPC in vivo; a well-characterized model of focal demyelination. GsMTx4 also attenuated both LPC-induced astrocyte toxicity and microglial reactivity within the lesion core. Overall, our data suggest that pharmacological activation of Piezo1 channels induces demyelination and that inhibition of mechanosensitive channels, using GsMTx4, may alleviate the secondary progressive neurodegeneration often present in the latter stages of demyelinating diseases.     

T-lymphocyte entry into the central nervous system

The entry of T-lymphocytes into the parenchyma of the central nervous system is a critical early feature in the pathogenesis of many experimental and spontaneously occurring immune-mediated illnesses. The physiological mechanisms controlling this entry have not been elucidated. This study reports that T-cell entry into the rat CNS appears to be primarily dependent upon the activation state of the lymphocytes; T-lymphoblasts enter the CNS (and all other tissues examined) in an apparently random manner while T cells not in blast phase are excluded. Antigen specificity, MHC compatibility, T-cell phenotype, and T-cell receptor gene usage do not appear related to the ability of cells to enter. This study demonstrates that when T-lymphoblasts are introduced into the circulation they rapidly appear in the CNS tissue. Their concentration in the CNS reaches a peak between 9 and 12 hr, and lymphocytes which have entered, exit within 1 to 2 days. Cells capable of reacting with a CNS antigen remain in the tissue or cyclically reenter to initiate inflammation if they are able to recognize their antigen in the correct MHC context. This observation also appears to pertain to the entry of activated T cells into many other tissues, although their concentrations in these non-CNS sites was not quantitated.     

Stem cell transplantation into the central nervous system and the control of differentiation

Recent advances in stem cell biology, including methods of cell amplification and control of differentiation in vitro, provide us with new and powerful tools with which to explore the cellular, molecular, and genetic factors affecting cell survival, proliferation, differentiation, and differentiation potential. Mitigating this vein of enthusiasm are the results of stem cell transplantation studies, which highlight our inability to control the fate of stem cell populations following transplantation to the central nervous system (CNS). Differentiation of transplanted cells is strongly influenced by the environmental signals and cellular deficiencies operating at the site of implantation, over which we can exert little or no control. Where stem cell transplantation-mediated repair of the injured CNS has been demonstrated most successfully, the transplant environments have invariably been simplistic, and transplantation into the complex and reactive environment of a CNS injury site generally results in migration from the site of implantation followed by glial cell differentiation. Together, these findings suggest that the most significant advances for the stem cell transplantation field will come from research strategies that include predifferentiation of stem cells prior to transplant and studies that further our understanding of the factors affecting stem cell differentiation in the complex environment of the CNS in vivo.     

Direct demonstration of the infiltration of murine central nervous system by Pgp-1/CD44 CD45RB CD4 T cells that induce experimental allergic encephalomyelitis

In experimental allergic encephalomyelitis (EAE), autoimmune T cells infiltrate the central nervous system (CNS) and initiate demyelinating pathology. We have used flow cytometry to directly analyse the migration to the CNS of MBP-reactive CD4⁺ T cells labelled with a lipophilic fluorescent dye (PKH2), in SJL/J mice with passively transferred EAE. Labelled cells constituted about 45% of the CNS CD4⁺ population at the time of EAE onset. Almost all (>90%) of the PKH2-labelled CD4⁺ T cells from EAE CNS were blasts and were α/β T cell receptor (TCR)⁺, CD44(Pgp-1)^high, and the majority were CD45RB^low. By contrast, most PKH2-labelled CD4⁺ T cells in lymph nodes, although CD44^high, were CD45RB^high cells. The cells that were transferred to induce EAE were essentially similar to antigen-primed lymph node cell populations, containing less than 15% CD44^high cells, and most of them were CD45RB^high. The CD44^high CD45RB^low phenotype is characteristic of memory/effector T cells that have been activated by antigen recognition. The difference in CD45RB expression between CNS and LN could therefore reflect differential exposure and/or response to antigen. Consistent with this, PKH2-labelled CD4⁺ cells isolated from the CNS were responsive to MBP in vitro, whereas PKH2⁺ CD4⁺ cells from lymph nodes showed almost undetectable responses. In control experiments in which ovalbumin (OVA)-reactive T cells were transferred, a small number of fluorescent-labelled CD4⁺ T cells were also detected in CNS, but there were very few blasts, and these remained CD45RB^high. These results argue for induction of the memory/effector phenotype of CD4⁺ cells, their selective retention in the CNS, as a consequence of antigen recognition.     

The design of clinical trials for cell transplantation into the central nervous system

Although initial excitement was raised in the medical and scientific communities by pilot trials using adrenal autografts (1980s) and fetal allograft (1990s), two controlled trials in PD patients demonstrated negative results, with the conclusion that today, cell therapy cannot be recommended for PD. Nevertheless, trials are still in progress for diseases like HD, stroke, MS, cord transection, epilepsy, and others. Human knowledge about the biology of stem cells and precursor cells is growing and we have more and more evidence that cell therapy could be useful in “repairing” brain lesions, not only by survival and integration of grafted neurons, but also by stimulation of resident precursors already present within the CNS.     

Molecular genetic diagnosis of a primary central nervous system T cell lymphoma

Primary central nervous system T cell lymphomas are rare tumors. Histologically, they may be indistinguishable from other entities like inflammatory processes. In such cases, molecular genetic verification of clonal T cell receptor (TCR) gene rearrangements is an indispensable diagnostic tool. Here we present a case where identification of TCR beta and gamma gene rearrangements by polymerase chain reaction was used to differentiate between a vasculitis and a primary CNS T cell lymphoma, which has profound consequences for therapy management and outcome.     

PECAM-1 (CD31) expression in the central nervous system and its role in experimental allergic encephalomyelitis in the rat

The role of T cell activation associated adhesion molecules on lymphocyte traffic and the initiation of inflammation has received considerable attention. This study, using a new monoclonal antibody (mAb) TLD-3A12, describes the distribution of PECAM-1 (CD31), an Ig supergene family adhesion molecule thought to be important in leukocyte transmigration during inflammation, in rat lymphoid organs and spinal cord. PECAM expression within the CNS is confined to endothelial cells of the blood brain barrier (BBB). Induction of inflammation within the CNS using the adoptive transfer of myelin reactive CD4⁺ T cells results in the de novo expression of immune adhesion and accessory molecules in the spinal cord, while the level of PECAM appeared only mildly increased. The distribution of PECAM on CNS endothelial cells became more diffuse during EAE induction, possibly the result of endothelial cell activation. In vitro studies demonstrate a partial inhibition of antigen-specific CD4⁴ T cell proliferation following anti-PECAM mAb treatment. Treatment of Lewis rats with TLD-3A12 antibody prior to T cell injection and throughout EAE induction does not result in a delay in the onset of clinical signs or weight loss, nor does it decrease the incidence and severity of disease. These data suggest that the expression of PECAM by CNS endothelial cells is not a requirement for the initiation of inflammation and clinical signs of EAE following the adoptive transfer of encephalitogenic lymphocytes. Thus, cells requiring PECAM-1 to migrate and perform their pathogenic functions are not critical to the development of rat EAE. © 1996 Wiley-Liss, Inc.     

Activated/effector CD4+ T cells exacerbate acute damage in the central nervous system following traumatic injury

CD4(+) helper T cells (Th) have been demonstrated to participate in the chronic phase of traumatic injury repair in the central nervous system (CNS). Here, we show that CD4(+) T cells can also contribute to the severity of the acute phase of CNS traumatic injury.We compared the area of tissue damage and the level of cellular apoptosis in aseptic cerebral injury (ACI) sites of C57BL/6 wild type and RAG1(-/-) immunodeficient mice. We demonstrate that ACI is attenuated in RAG1(-/-) mice compared to C57BL/6 animals. Adoptive transfer of CD4(+)CD62L(low)CD44(high) activated/effector T cells 24 h prior to ACI into RAG1(-/-) mice resulted in a significantly enhanced acute ACI that was comparable to ACI in the C57BL/6 animals. Adoptive transfer of CD4(+)CD62L(high)CD44(low) naive/non-activated T cells did not increase ACI in the brains of RAG1(-/-) mice. T cell inhibitory agents, cyclosporin A (CsA) and FK506, significantly decreased ACI-induced acute damage in C57BL/6 mice. These results suggest a previously undescribed role for activated/effector CD4(+) T cells in exacerbating ACI-induced acute damage in the CNS and raise a novel possibility for acute treatment of sterile traumatic brain injury.     

Stem cell biology of the central nervous system

Neural stem cells (NSCs) are multipotential progenitor cells that have self-renewal activities. A single NSC is capable of generating various kinds of cells within the central nervous system (CNS), including neurons, astrocytes, and oligodendrocytes. Because of these characteristics, there is increasing interest in NSCs and neural progenitor cells from the aspects of both basic developmental biology and therapeutic applications to the damaged brain. This special issue, dedicated to understanding the nature of the NSCs present in the CNS, presents an introduction to several avenues of research that may lead to feasible strategies for manipulating cells in situ to treat the damaged brain. The topics covered by these studies include the extracellular factors and signal transduction cascades involved in the differentiation and maintenance of NSCs, the population dynamics and locations of NSCs in embryonic and adult brains, prospective identification and isolation of NSCs, the induction of NSCs to adopt particular neuronal phenotypes, and their transplantation into the damaged CNS.     

Monocyte-mediated entry of pathogens into the central nervous system

The origin of the microglia has long been a subject of debate. However it is now clear that monocytes enter the normal central nervous system and follow a series of morphological transformations as they differentiate into microglia. Thus, microglia are of monocytic origin. Since monocytes migrate into the normal CNS, they represent potential vehicles for the entry of pathogens into the nervous system and indeed may carry particulate matter into the CNS. Both viruses and bacteria use this 'Trojan horse' mechanism of entry in the pathogenesis of CNS disease.     

Transplantation of adrenal tissue into the central nervous system

Adrenal medullary tissue can survive transplantation to the central nervous system. Such survival has been obtained experimentally with grafts to the anterior eye chamber, to the brain and to the spinal cord, using medullary tissue from the recipient animal or unrelated animals of the same or, in some cases, different species. Appropriately placed grafts have been shown, under certain conditions, to interact with the host nervous system, exerting behavioral effects including amelioration of experimentally-induced parkinsonian symptoms. Such effects may be enhanced by administration of nerve growth factor to the grafts. On the basis of such findings, adrenal medullary tissue has been grafted to the brain of Parkinson's disease patients. Both animal and human experiments raise important questions about mechanisms of graft action and about factors that influence the outcome of these procedures.     

Transition from enhanced T cell infiltration to inflammation in the myelin-degenerative central nervous system

Myelin degeneration in the central nervous system (CNS) is often associated with elevated numbers of T cells in brain and spinal cord (SC). In some degenerative diseases, this T cell immigration has no clinical relevance, in others, it may precede severe inflammation and tissue damage. We studied T cells in the myelin-degenerative SC of transgenic (tg) Lewis rats overexpressing the proteolipid protein (PLP). These lymphocytes are T(H)1/T(C)1 cells and represent different T cell clones unique to individual animals. The SC-infiltrating CD8(+) T cell pool is more restricted than its CD4(+) counterpart, possibly due to constrictions in the peripheral CD8(+) T cell repertoire. Some SC-infiltrating T cells are highly motile and cover large distances within their target tissue, others are tethered to MHC class II(+) microglia cells. The activation of the tethered cells may trigger the formation of inflammatory foci and could pave the way for inflammation in degenerative CNS disease.