TNF-α and IFN-γ render microglia sensitive to Fas ligand-induced apoptosis by induction of Fas expression and down-regulation of Bcl-2 and Bcl-xL

The immune response in the central nervous system (CNS) involves microglial cells which represent intraparenchymal antigen-presenting cells (APC). To control immune effector mechanisms it may be required to induce apoptosis of APC and thereby limit reactivation of T cells that have invaded the CNS. In the present study we investigated the susceptibility of primary murine microglia and of the murine microglial cell line BV-2 to undergo Fas-mediated apoptosis. Whereas resting microglia are resistant to Fas ligand (FasL) treatment, induction of FasL-mediated apoptosis was achieved by treatment with TNF-α or IFN-γ. The effect of these cytokines was paralleled by up-regulation of Fas expression and down-regulation of Bcl-2 and Bcl-xL but not Bax. Activation of microglia by TNF-α and IFN-γ was also accompanied by increased amounts of mRNA for the apoptosis inhibit FLIP, an effect which did not protect the cells from FasL-induced apoptosis. The FasL-induced cell death pathway in microglia involves reactive oxygen intermediates because the antioxidants N-acetyl-cysteine and glutathione interfere with induction of apoptosis. Surprisingly, microglia constitutively express FasL on the cell surface. However, blocking of endogenous Fas-FasL inter action with Fas-Fc fusion protein did not enhance the survival of microglia, excluding the possibility of suicide or fratricide mechanisms. By their expression of FasL and their TNF-α / IFN-γ-dependent sensitivity to the pro-apoptotic effect of exogenous FasL, microglial cells may influence the course of T cell-mediated diseases of the CNS.     

B cell apoptosis in the central nervous system in experimental autoimmune encephalomyelitis: roles of B cell CD95, CD95L and Bcl-2 expression

The role and fate of B cells in the central nervous system (CNS) in experimental autoimmune encephalomyelitis (EAE) are unknown. Using enzyme-linked immunospot assays we now show that B cells reactive to myelin basic protein (MBP) accumulate in the CNS of Lewis rats with acute EAE induced by immunization with MBP and adjuvants. We also report that B cells are eliminated from the CNS by apoptosis during spontaneous recovery from this disease. Apoptotic B cells were identified by flow cytometry of inflammatory cells extracted from the spinal cord and by histological sections of the spinal cord using light and electron microscopic immunocytochemistry. B cell apoptosis occurred preferentially in the CNS rather than in the peripheral lymphoid organs and was maximal just prior to the onset of spontaneous clinical recovery. Three colour flow cytometry indicated that B cells expressing CD95 (Fas) or CD95 ligand (CD95L) were highly vulnerable to apoptosis, whereas B cells expressing Bcl-2 were relatively protected from apoptosis. We propose that B cells are eliminated from the CNS by the interaction of CD95L and CD95 on the same B cell and that this contributes to the spontaneous resolution of CNS inflammation and clinical recovery in acute EAE.     

Parenchymal microglia of naïve adult C57BL/6J mice express high levels of B7.1, B7.2, and MHC class II

In this study, we addressed B7.1, B7.2, and MHC class II expression on microglia of normal adult C57BL/6J mice, which are susceptible to MOG35-55 peptide-induced experimental autoimmune encephalomyelitis (EAE). We showed that there are two distinct major populations of CD11b(+) cells in the central nervous system (CNS) of na?ve mice: CD45 low (CD45(lo); parenchymal microglia) and CD45 intermediate (CD45(int); CNS-associated macrophages). These two populations compose CNS microglia. There is a rare CD45 high (CD45(hi)) population. By contrast, splenic CD11b(+) cells (macrophages) are CD45(int) and CD45(hi), but rarely CD45(lo). CD45(lo)CD11b(+) cells constitutively express much higher levels of B7.1, B7.2, and MHC class II compared to CD45(int) CD11b(+) cells. A shift of CD11b(+) cells from CD45(lo) to CD45(int) was observed in the CNS of EAE mice. Our study provides evidence that (1) CD45(lo) and CD45(hi), but not CD45(int), could be unique markers to differentiate parenchymal microglia from infiltrating macrophages in EAE; (2) the level of CD45 expression on parenchymal microglia (CD45(lo)) was upregulated in EAE; and (3) parenchymal microglia in normal CNS could be potent APCs by expressing high levels of B7.1, B7.2, and MHC class II molecules and could therefore play an important role in inflammation and autoimmunity in the CNS.     

IFN-gamma regulates Fas ligand expression in human CD4+ T lymphocytes and controls their anti-mycobacterial cytotoxic functions

Fas and Fas Ligand (FasL) expression, activation-induced cell death (AICD) and mycobacterial antigen-specific cytotoxicity of peripheral T cells from patients with complete inherited IFN-gamma receptor 1 binding chain deficiency (IFN-gammaR1-/-) were investigated. Fas was equally expressed in both normal and deficient T lymphoblasts and they underwent apoptosis when stimulated with agonist anti-Fas mAb. By contrast, T lymphoblasts and CD4+ T cell clones (TCC) from deficient patients displayed a reduced surface FasL expression and resistance to AICD. CD8+ TCC from healthy and deficient patients displayed similar high level of FasL and susceptibility to AICD. In Jurkat CD4+ T cells competent to transduce IFN-gamma signaling, IFN-gamma induced surface FasL export and their Fas-dependent apoptosis. Effector T cells generated from a patient with a dominant negative mutation of IFN-gammaR1 (IFN-gammaR1DN) following stimulation with mycobacterial antigens were unable to kill MHC class II-matched, mycobacterial antigen-pulsed macrophages. Normal Fas expression in T cells and FasL in CD8+ cells may account for the absence of autoimmune disorders in these patients. Conversely, defective FasL expression on IFN-gammaR1DN CD4+ T cells impairs their cytotoxic functions and highlights a novel role for IFN-gamma signaling in the control of mycobacterial infection in humans.     

TGF-beta induces the expression of the FLICE-inhibitory protein and inhibits Fas-mediated apoptosis of microglia

During inflammatory reactions in the central nervous system (CNS), resident macrophages, the microglia, are exposed to Th1 cell-derived cytokines and pro-apoptotic Fas ligand (FasL). Despite the presence of TNF-alpha and IFN-gamma, both being capable of sensitizing microglia to FasL, apoptosis of microglia is not a hallmark of inflammatory diseases of the CNS. In the present study, TGF-beta is found to counteract the effect of TNF-alpha and IFN-gamma to sensitize microglia to FasL-mediated apoptosis. Resistance to Fas-mediated apoptosis by TGF-beta does not correlate with a down-regulation of Fas expression. As a key inhibitor of Fas-mediated apoptosis, we found expression of the cellular FLICE-inhibitory protein (c-FLIP) to be induced by TGF-beta in resting as well as in activated microglia. Induction of FLIP was found to depend on a mitogen-activated protein kinase kinase (MKK)-dependent pathway as shown by the use of the specific MKK-inhibitor PD98059. The presence of FLIP strongly interfered with FasL-induced activation of caspase-8 and caspase-3 preventing subsequent cell death. The presented data provide the first evidence for a TGF-beta-mediated FLIP in macrophage-like cells and suggest a mode of action for the anti-apoptotic role of TGF-beta in the CNS.     

Enhanced T cell apoptosis in common variable immunodeficiency: negative role of the fas/fasligand system and of the Bcl-2 family proteins and possible role of TNF-RS

CVI is a primary immunodeficiency characterized by a failure of B cell differentiation associated with an array of T cell defects, such as enhanced T cell apoptosis. In this study we investigated the mechanisms underlying CVI enhanced T cell death. We analysed both the expression of Fas using flow cytometry techniques and the expression of FasL mRNA using RT-PCR in CVI T cells. We could not find any significant differences between CVI and normal subjects with regard to Fas expression, although there was a subgroup of CVI patients with very high Fas expression which was accompanied by an up-regulation of FasL mRNA. However, attempts to induce Fas-mediated apoptosis in these high Fas expressing cells, as evaluated by propidium iodide staining and APO2.7 staining, were unsuccessful. We also investigated intracellular levels of Bcl-2, bcl-xl and bax in CD4(+) and CD8(+) CVI T cells, as well as the bax/Bcl-2 ratio, using flow cytometry techniques but could not detect any differences between CVI and normal subjects. Finally we analysed TNF-RI and TNF-RII mRNA expression in CD4(+) and CD8(+) CVI T cells using semiquantitative RT-PCR and found a significant increase in expression of both TNF-Rs in CD4(+) T cells from CVI patients. Our data suggest that the increased expression of both TNF-Rs on T cells may be one of the mechanisms responsible for the accelerated T cell apoptosis in CVI.     

Upregulation of group 1 CD1 antigen presenting molecules in guinea pigs with experimental autoimmune encephalomyelitis: an immunohistochemical study

In humans, group 1 CD1 glycoproteins present foreign and self lipid and glycolipid antigens to T-cells. Homologues of these molecules are not found in mice or rats but are present in guinea pigs (GPs). We examined CD1 and MHC class II expression in the central nervous system (CNS) of GPs sensitized for experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis. In normal GPs and the uninflamed CNS, low-level MHC class II (MHC II) immunoreactivity occurred on vascular elements, meningeal macrophages and parenchymal microglial cells, whereas immunoreactivity for CD1 was absent. In the inflamed CNS, the majority of infiltrating cells were MHC II+ and microglia showed increased expression. CD1 immunoreactivity was detected on astrocytes and subsets of inflammatory cells Including B cells and macrophages. Minimal CD1 and MHC II co-expression was noted on inflammatory cells or glia. We conclude that group 1 CD1 molecules are strongly upregulated in the inflamed CNS on subsets of cells distinct from the majority of MHC II bearing cells. The expression of CD1 proteins in such lesions broadens the potential repertoire of antigens recognized at these sites and highlights the value of the GP as a model for studies of the relevance of CD1 molecules in host defense and autoimmune diseases.     

Soluble antigen therapy induces apoptosis of autoreactive T cells preferentially in the target organ rather than in the peripheral lymphoid organs

The administration of soluble myelin proteins is an effective way of down-regulating the inflammation in the central nervous system (CNS) in experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis. To shed more light on the mechanism of this antigen-specific therapy, we determined the effect of the intraperitoneal (i.p.) injection of soluble myelin basic protein (MBP) on T cell apoptosis in the CNS and peripheral lymphoid organs of Lewis rats with EAE induced by inoculation with MBP and complete Freund's adjuvant. In particular we assessed the level of apoptosis of Vβ8.2⁺ T cells, which constitute the predominant encephalitogenic MBP-reactive T cell population in the Lewis rat. The daily i.p. injection of MBP for 3 days from the onset of neurological signs inhibited the further development of neurological signs of EAE. Using two-color flow cytometry we found that a single i.p. injection of MBP increased the level of apoptosis of the Vβ8.2⁺ T cell population in the CNS to 26.2 % compared to 7.4 % in saline-treated rats and 7.6 % in ovalbumin-treated rats. In contrast, treatment with MBP did not increase the level of apoptosis of the Vβ8.2⁺ population in the popliteal lymph node draining the inoculation site (1.4 %) or in the spleen (1.6 %) above that occurring in saline-treated rats (1.6 % and 1.1 %, respectively). Limiting dilution analysis revealed that the frequency of T cells reactive to the major encephalitogenic epitope, MBP_{72 – 89}, was decreased in the CNS but not in the popliteal lymphnode by this treatment. Three-color flow cytometry in MBP-treated rats demonstrated that CNS Vβ8.2⁺ T cells expressing Fas (CD95) and Fas ligand were highly vulnerable to apoptosis compared to Vβ8.2⁺ T cells not expressing these proteins. We conclude that the i.p. injection of MBP increases the spontaneously occurring Fas-mediated activation-induced apoptosis of auto reactive T cells in the CNS in EAE and that this contributes to the therapeutic effect of the injection.     

Characterization of the expanded T cell population in infectious mononucleosis: apoptosis, expression of apoptosis-related genes, and Epstein-Barr virus (EBV) status

Infectious mononucleosis (IM), a manifestation of primary infection with EBV, is characterized by a massive expansion of the T cell population. In this study we examined this expanded T cell population regarding its EBV status, its proliferative and apoptotic activity, and its expression of apoptosis-related genes. Whereas previous studies were performed on ex vivo cultures or on peripheral blood, our investigations included in vivo analysis of IM tonsillectomy specimens (14 cases) by in situ hybridization for viral RNA (EBERs) combined with immunohistochemistry (IHC; CD3, CD45RO, CD20, CD79a, Ki-67, Bcl-2, Bax, Fas, FasL) and the TUNEL method. Of the EBER+ cells 50-70% showed expression of the B cell markers CD20/CD79a. The remainder of the EBER+ cells expressed neither B nor T cell antigens. No co-expression of EBERs and T cell antigens was detected in any of the specimens. In accordance with a high rate of apoptosis (up to 2.37%) within the expanded T cell population, Bcl-2 expression was drastically reduced and FasL expression remarkably increased. The levels of Bax and Fas expression showed no or moderate up-regulation. In conclusion, the massive expansion of IM T cells is not caused by EBV infection of these cells but merely represents an intense immune reaction. Through altered expression of Bcl-2/Bax and Fas/FasL, the activated T cells are subject to enhanced apoptosis while residing within the lymphoid tissue, which eventually allows the efficient silencing of this potentially damaging T cell response.     

Decreased T Cell Stimulatory Capacity of Monocyte-Derived Human Macrophages following Herpes simplex Virus Type 1 Infection

Macrophages play a central role in establishing a specific immune response by acting as professional antigen presenting cells (APC) for T cells leading to a vigorous immune response. In order to analyze if Herpes simplex Virus (HSV) type 1 infection might affect the macrophage APC-function, monocyte-derived human macrophages were infected with HSV-1 strain F in vitro. Cocultures with allogeneic T cells revealed a strongly impaired stimulatory capacity of HSV-infected macrophages compared to uninfected controls which was not owing to a productive viral infection in macrophages. An increased expression of Fas ligand (FasL/CD95L) was detected in HSV-infected macrophages by FACS analysis. Although the majority of the macrophages expressed high levels of Fas (CD95/Apo-1), the HSV-induced upregulation of FasL did not result in an increased autocrine apoptosis of macrophages which might be related to endogenous expression of the apoptosis inhibitor FLICE inhibitory protein (FLIP). However, substantial apoptosis occurred in peripheral T cells as well as Fas-sensitive Jurkat T cells when cocultured with HSV-infected macrophages. These findings suggest that the paracrine killing of activated T cells by FasL expressing APC might be a novel strategy of immune evasion by HSV.     

CD4+ T cell-mediated cytotoxicity toward thyrocytes: the importance of Fas/Fas ligand interaction inducing apoptosis of thyrocytes and the inhibitory effect of thyroid-stimulating hormone

The accumulation of activated CD4+ T cells and antigen (Ag)-dependent cellular interactions between thyrocytes and CD4+ T cells have been determined in thyroid gland from patients with Graves' disease. The Fas/Fas ligand (FasL) interaction between antigen-presenting cells and T cells regulates the apoptosis of the former cells triggered by the latter cells. The inhibition of Fas-mediated apoptosis in thyrocytes could be a underlying mechanism of hyperplasia of thyrocytes in patients with Graves' disease. We investigated the potential role of Fas/FasL interaction between thyrocytes and CD4+ T cells in the induction of Fas-mediated apoptosis of the former cells induced by the latter cells. The presence of only a few specific T cells responsive to a putative autoantigen has hampered the investigation of specific T cell activation toward antigen-presenting cells (APCs). Therefore, we used a superantigen, staphylococcal enterotoxin B (SEB), to examine specific T cell activation toward thyrocytes in vitro since it stimulates a large proportion of T cells with particular Vbeta elements. Spontaneous apoptosis of thyrocytes in culture was not found even in the presence of various kinds of cytokines. In contrast, a clear induction of Fas-mediated apoptosis by anti-Fas IgM was determined in interferon-gamma (IFN-gamma)-stimulated thyrocytes. In addition, a significant cytotoxicity of purified CD4+ T cells toward IFN-gamma-stimulated thyrocytes in the presence of SEB was induced, and the addition of anti-HLA-DR and -DQ monoclonal antibodies (mAbs) or blockade of the Fas/FasL interaction reduced this cytotoxicity. FasL expression of CD4+ T cells cocultured with IFN-gamma-stimulated thyrocytes in the presence of SEB was clearly induced. Furthermore, the addition of mAbs against CD54 and CD58 inhibited both cytotoxicity and FasL expression of CD4+ T cells. The cytotoxicity of CD4+ T cells toward IFN-gamma-stimulated, SEB-pulsed thyrocytes was markedly inhibited when we used thyrocytes cultured with IFN-gamma in the presence of thyroid-stimulating hormone (TSH) as target cells. Our results suggest that 1) CD4+ T cells were activated by thyrocytes expressing MHC class II molecules in an SEB-dependent manner and then expressed FasL. 2) These activated FasL+ CD4+ T cells killed thyrocytes by interacting with Fas on thyrocytes and FasL on activated CD4+ T cells. The presence of costimulating molecules such as CD54 and CD58 on thyrocytes was also necessary to generate activated FasL+ CD4+ T cells. 3) Since the actions of thyroid stimulating antibody (TSAb) toward thyrocytes are similar to those of TSH, one goitrogenic activity of TSAb may, in part, be due to the inhibitory effect on Fas-mediated apoptosis of thyrocytes triggered by activated CD4+ T cells.     

Apoptosis with FasL+ cell infiltration in the periphery and thymus of corrected autoimmune mice.

Fas (CD95) ligand (L) is a death factor that binds to its receptor, Fas, and induces apoptotic cell death, a crucial process in immunological tolerance. gld (generalized lymphoproliferative disorder) mice, which have a point mutation in the FasL gene, develop spontaneous systemic autoimmune syndromes characterized by hypergammaglobulinaemia and lymphoid hyperplasia owing to accumulation of abnormal B220+ CD3+ cells. Transplantation of wild-type (wt) bone marrow cells into old gld mice on the same strain background results in normalization of autoimmune syndromes. We characterized the cellular mechanisms (functionally and histologically) of the above phenomena in gld mice after bone marrow transplantation (BMT) to determine the role of apoptosis via Fas/FasL interactions in inducing and maintaining self-tolerance in vivo. Activated splenocytes from wt and BMT (wt to gld) mice showed significant cytotoxic activity against Fas transfectant cells while those from BMT (gld to gld) mice did not. Cells in the thymus, spleen and lymph nodes of gld mice uniformly upregulated Fas expression and were sensitive to Fas-mediated apoptosis compared with those in wt mice. Cells sensitive to Fas-mediated apoptosis in gld mice resided not only among abnormal B220+ CD3+ cells but also among conventional lymphocytes. More importantly, histological analysis revealed that cells in the spleen, lymph nodes and thymus frequently underwent apoptosis with infiltration of FasL+ cells in BMT (wt to gld) mice compared with BMT (gld to gld) mice. Our results indicated that apoptosis via Fas/FasL interactions can directly eliminate pathogenic cells responsible for autoimmunity in the periphery and possibly in the thymus in vivo.     

The central nervous system environment controls effector CD4+ T cell cytokine profile in experimental allergic encephalomyelitis

In experimental allergic encephalomyelitis (EAE), CD4⁺ T cells infiltrate the central nervous system (CNS). We derived CD4⁺ T cell lines from SJL/J mice that were specific for encephalitogenic myelin basic protein (MBP) peptides and produced both Th1 and Th2 cytokines. These lines transferred EAE to naive mice. Peptide-specific cells re-isolated from the CNS only produced Th1 cytokines, whereas T cells in the lymph nodes produced both Th1 and Th2 cytokines. Mononuclear cells isolated from the CNS, the majority of which were microglia, presented antigen to and stimulated MBP-specific T cell lines in vitro. Although CNS antigen-presenting cells (APC) supported increased production of interferon (IFN)-γ mRNA by these T cells, there was no increase in the interleukin (IL)-4 signal, whereas splenic APC induced increases in both IFN-γ and IL-4. mRNA for IL-12 (p40 subunit) was up-regulated in both infiltrating macrophages and resident microglia from mice with EAE. We have thus shown that a Th1 cytokine bias within the CNS can be induced by CNS APC, and that IL-12 is up-regulated in microglial cells within the CNS of mice with EAE. Microglia may therefore control Th1 cytokine responses within the CNS.     

Apoptosis of CD4+ T cells occurs in experimental autoimmune anterior uveitis (EAAU)

To investigate the spontaneous turning off mechanism of endogenous uveitis, EAAU was induced in Lewis rats. Immunohistochemical and terminal deoxynucleotidyl transferase-mediated dUTP nick end labelling (TUNEL) stains revealed that CD4+ T cells were predominant in the uveal tissue of EAAU and that the apoptosis of these cells had occurred and progressed throughout the inflammatory period in EAAU eyes. The immunohistochemistry and in situ hybridization for Fas ligand (FasL) expression showed that the expression of Fas ligand was increased in the EAAU eyes compared with control eyes. These results suggest that the apoptosis of CD4+ T cells may play a key role in the spontaneous turning off mechanism of intra-ocular inflammation and that the induction of apoptosis may be mediated by the Fas-FasL system in EAAU.     

Role of Fas/FasL signaling in regulation of anti-viral response during HSV-2 vaginal infection in mice

Fas receptor–Fas ligand (FasL) signaling is involved in apoptosis of virus-infected cells but increasing evidence accumulates on Fas receptor as a mediator of apoptosis-independent processes such as induction of activating and pro-inflammatory signals. In this study, we examined the role of Fas/FasL pathway in regulation of anti-viral response to genital HSV-2 infection using a murine model of HSV-2 infection applied to C57BL6/J, B6. MRL-Faslpr/J and B6Smn.C3-Faslgld/J mice. HSV-2 infection of Fas- and FasL-deficient mice led to decreased migration of IFN-γ expressing NK cells and CD4+ T cells, but not of γδ T cells, into the vaginal tissue. The vaginal tissues of HSV-2 infected Fas- and FasL-deficient mice showed increased production of IL-10, followed by low expression of the early CD69 activation marker on CD4+ and CD8+ T cells and increased numbers of regulatory T cells (Tregs). Experiments in co-cultures of CD4+ T cells and bone marrow derived dendritic cells showed that lack of bilateral Fas–FasL signaling led to expansion of Tregs and increased production of IL-10 and TGF-β1. Our results demonstrate that Fas/FasL can regulate development of tolerogenic dendritic cells and expansion of Tregs early during HSV-2 infection, which further influences effective anti-viral response.     

GADD45β参与类风湿关节炎发病机制研究(2)—GADD45β介导类风关CD4~+T细胞抗凋亡研究

在发现了GADD45β在类风湿关节炎(rheumatoid arthritis,RA)病灶局部组织细胞中、CD4+T细胞表达增高并参与介导IFN-γ产生的基础上,进一步探讨GADD45β参与RA的其他机制。用siRNA干扰人PBMC中GADD45β基因表达。Real-time PCR检测GADD45β和凋亡相关基因的表达格局。用RA患者滑膜液(RA synovial fluid,RA SF)刺激GADD45β基因敲除的健康人PBMC并用流式细胞仪检测细胞凋亡。结果RA患者的SFMC中T细胞凋亡减少。用SF刺激正常人PBMC时细胞凋亡减少。经siRNA干扰后PBMC中GADD45β的表达明显降低,同时细胞凋亡增加。同时检测与凋亡相关基因发现SF刺激后Bcl-2分子在凋亡的细胞中表达明显降低,而Fas-FasL、Bcl-2家族其他分子如Bcl-xl、Bax、Bim等变化不明显。GADD45β参与介导RA的炎症性T细胞凋亡,而Bcl-2分子参与了由GADD45β介导抗凋亡作用,而这一作用与RA患者滑膜液中炎症细胞能够长期存活并介导炎症可能有密切关系。     

Human brain parenchymal microglia express CD14 and CD45 and are productively infected by HIV-1 in HIV-1 encephalitis

Microglia are endogenous brain macrophages that show distinct phenotypes such as expression of myeloid antigens, ramified morphology, and presence within the neural parenchyma. They play significant roles in a number of human CNS diseases including AIDS dementia. Together with monocyte-derived (perivascular) macrophages, microglia represent a major target of HIV-1 infection. However, a recent report challenged this notion based on findings in SIV encephalitis. This study concluded that perivascular macrophages can be distinguished from parenchymal microglial cells by their expression of CD14 and CD45, and that macrophages, but not microglia, are productively infected in SIV and HIV encephalitis. To address whether parenchymal microglia are productively infected in HIV encephalitis, we analyzed expression of CD14, CD45 and HIV-1 p24 in human brain. Microglia were identified based on their characteristic ramified morphology and location in the neural parenchyma. We found that parenchymal microglia are CD14+ (activated), CD45+ (resting and activated), and constitute approximately two thirds of the p24+ cells in HIV encephalitis cases. These results demonstrate that microglia are major targets of infection by HIV-1, and delineate possible differences between HIVE and SIVE. Because productively infected tissue macrophages serve as the major viral reservoir, these findings have important implications for AIDS.     

The Fas death pathway controls coordinated expansions of type 1 CD8 and type 2 CD4 T cells in Trypanosoma cruzi infection

We investigated the role of the Fas ligand (FasL)/Fas death pathway on apoptosis and cytokine production by T cells in Trypanosoma cruzi infection. Anti-FasL, but not anti-TNF-alpha or anti-TRAIL, blocked activation-induced cell death of CD8 T cells and increased secretion of IL-10 and IL-4 by CD4 T cells from T. cruzi-infected mice. CD4 and CD8 T cells up-regulated Fas/FasL expression during T. cruzi infection. However, Fas expression increased earlier in CD8 T cells, and a higher proportion of CD8 T cells was activated and expressed IFN-gamma compared with CD4 T cells. Injection of anti-FasL in infected mice reduced parasitemia and CD8 T cell apoptosis and increased the ratio of CD8:CD4 T cells recovered from spleen and peritoneum. FasL blockade increased the number of activated T cells, enhanced NO production, and reduced parasite loads in peritoneal macrophages. Injection of anti-FasL increased IFN-gamma secretion by splenocytes responding to T. cruzi antigens but also exacerbated production of type 2 cytokines IL-10 and IL-4 at a late stage of acute infection. These results indicate that the FasL/Fas death pathway regulates apoptosis and coordinated cytokine responses by type 1 CD8 and type 2 CD4 T cells in T. cruzi infection.     

The role of the Fas/Fas ligand system in estrogen-induced thymic alteration

PROBLEM: Estrogen induces atrophy in the thymus by an unknown mechanism. Since the Fas/FasL system is one of the main pathways in T cell apoptosis, we tested the hypothesis that estrogen-induced thymic atrophy is mediated by the Fas/FasL system. METHODS OF STUDY: In vivo experiments were done using ovariectomized female rats treated with estrogen or saline. In vitro experiments were performed using isolated thymocytes. Estrogen receptor (ER) alpha and beta expression was characterized using flow cytometry, RT-PCR and immunofluorescence. Fas and FasL mRNA and protein expression was evaluated using RT-PCR and Western blot analysis respectively. RESULTS: ERalpha and ERbeta are present in thymocytes and stromal cells. ER expression is mainly localized in the Double Positive CD4+CD8+ thymocytes. Estrogen treatment decreases thymus size and increase FasL expression. CONCLUSION: CD4+CD8+ thymocytes and thymic stroma cells express ERalpha and ERbeta. In vivo and in vitro we showed that estrogen treatment increases FasL expression while decreasing thymus cell number. These findings support the hypothesis that estrogen-induced thymic atrophy occurs as a result of apoptosis and is mediated by estrogen-induced FasL expression.