Malek Kamoun

IL-12p35-deficient mice are susceptible to experimental autoimmune encephalomyelitis : evidence for redundancy in the IL-12 system in the induction of central nervous system autoimmune demyelination

Experimental autoimmune encephalomyelitis (EAE) serves as a model for multiple sclerosis and is considered a CD4+, Th1 cell-mediated autoimmune disease. IL-12 is a heterodimeric cytokine, composed of a p40 and a p35 subunit, which is thought to play an important role in the development of Th1 cells and can exacerbate EAE. We induced EAE with myelin oligodendrocyte glycoprotein (MOG) peptide 35–55 (MOG35–55) in C57BL/6 mice and found that while IL-12p40-deficient (−/−) mice are resistant to EAE, IL-12p35−/− mice are susceptible. Typical spinal cord mononuclear cell infiltration and demyelination were observed in wild-type and IL-12p35−/− mice, whereas IL-12p40−/− mice had normal spinal cords. A Th1-type response to MOG35–55 was observed in the draining lymph node and the spleen of wild-type mice. A weaker MOG35–55-specific Th1 response was observed in IL-12p35−/− mice, with lower production of IFN-γ. By contrast, a Th2-type response to MOG35–55 correlated with disease resistance in IL-12p40−/− mice. Production of TNF-α by microglia, CNS-infiltrating macrophages, and CD4+ T cells was detected in wild-type and IL-12p35−/−, but not in IL-12p40−/−, mice. In addition, NO production was higher in IL-12p35−/− and wild-type mice than in IL-12p40−/− mice. These data demonstrate a redundancy of the IL-12 system in the induction of EAE and suggest that p40-related heterodimers, such as the recently cloned IL-23 (p40p19), may play an important role in disease pathogenesis.

Induction of Experimental Autoimmune Encephalomyelitis in IL-12 Receptor-β2-Deficient Mice: IL-12 Responsiveness Is Not Required in the Pathogenesis of Inflammatory Demyelination in the Central Nervous System

IL-12 is thought to be involved in the susceptibility to experimental autoimmune encephalomyelitis (EAE), a Th1 cell-mediated autoimmune disorder of the CNS. IL-12 signals through a heterodimeric receptor (IL-12Rβ1/IL-12Rβ2), whose β2-chain is up-regulated on activated, autoreactive Th1 cells. Contrary to the expectation that the absence of IL-12Rβ2 would protect from EAE, we found that IL-12Rβ2-deficient mice developed earlier and more severe disease, with extensive demyelination and CNS inflammation. The inflammatory cells were mainly comprised of CD4+ T cells, monocyte/macrophages, and dendritic cells. Compared to wild-type mice, IL-12Rβ2-deficient mice exhibited significantly increased autoantigen-induced proliferative response and increased production of TNF-α, GM-CSF, IL-17, IL-18/IL-18Rα, and NO. In addition, we found significantly increased levels of IL-23p19 mRNA expression in spleen cells from immunized IL-12Rβ2−/− mice compared with wild-type mice. These findings indicate that IL-12 responsiveness is not required in the pathogenesis of inflammatory demyelination in the CNS, and that, in the absence of IL-12Rβ2, increased IL-23 and other inflammatory molecules may be responsible for increased severity of EAE.

Erythrophagocytic Tγ Lymphoma

CLINICOPATHOLOGICAL studies suggest that the T-cell lymphomas are heterogeneous with respect to clinical presentation, course, and morphology. Mycosis fungoides and the Sezary syndrome affect the s...

Race and Electronically Measured Adherence to Immunosuppressive Medications after Deceased Donor Renal Transplantation

Nonadherence to immunosuppressive medications may partly explain the worse allograft outcomes among black recipients of renal transplants. In a prospective cohort study of recipients of deceased donor renal transplants, microelectronic cap monitors were placed on bottles of one immunosuppressive medication to (1) measure average daily percentage adherence during the first posttransplantation year and (2) determine the factors associated with adherence. A total of 278 transplant recipients who provided sufficient microelectronic adherence data were grouped into four categories of average daily percentage adherence: 95 to 100% adherence (41.0% of patients), 80 to 95% adherence (32.4%), 50 to 80% adherence (12.9%), and 0 to 50% adherence (13.7%). In the unadjusted ordinal logistic regression model, black race was associated with decreased adherence (odds ratio [OR], 0.43; 95% confidence interval [CI], 0.26 to 0.72; P = 0.001). Cause of renal disease, Powerful Others health locus of control, transplant center, and dosing frequency were also associated with adherence. After adjustment for transplant center and dosing frequency, the association between black race and decreased adherence was substantially attenuated (OR, 0.65; 95% CI, 0.38 to 1.14, P = 0.13). Transplant center (P = 0.003) and increased dosing frequency (OR, 0.43; 95% CI, 0.22 to 0.86, for three or four times per day dosing; OR, 2.35; 95% CI, 1.01 to 5.45, for daily dosing; versus two times per day dosing; P = 0.003) remained independently associated with adherence. Other baseline demographic, socioeconomic, medical, surgical, and psychosocial characteristics were not associated with adherence. The transplant center and dosing frequencies of immunosuppressive medications are associated with adherence and explain a substantial proportion of the race-adherence relationship.

Differential expression and regulation of IL-23 and IL-12 subunits and receptors in adult mouse microglia

IL-23 and IL-12 are functionally related heterodimeric cytokines that share the IL-12p40 subunit. IL-23 and IL-12 function through heterodimeric receptors, which share the IL-12Rbeta1 subunit. Production of IL-23, a heterodimer of IL-12p40 and IL-23p19, by CNS antigen-presenting cells (APC) is critical for susceptibility to experimental autoimmune encephalomyelitis (EAE), the animal model for multiple sclerosis (MS). We report that the expression of IL-23p19 mRNA is highly induced by stimulation with IFN-gamma and LPS in adult mouse microglia and a microglia cell line, EOC13. Expression of the IL-12R subunits, IL-12Rbeta1 and IL-12Rbeta2, is upregulated in both microglia and splenic macrophages upon stimulation with LPS or IFN-gamma and LPS, whereas the IL-23R subunit is upregulated only in macrophages. In EAE, an early peak of IL-23p19 mRNA expression is found in CD11b(+) CNS APC, compared with peripheral macrophages. In contrast, IL-12p40 and IL-12p35 mRNA maximum levels in the CNS are detected at peak of disease. The expression of IL-12p35 mRNA is more sustained than that of IL-12p40 and IL-23p19. Thus, IL-23 produced by CNS microglia/macrophages may contribute to the early induction of EAE. In the CNS, IL-23 may preferentially target infiltrating mononuclear cells, which upregulate IL-23R, rather than parenchymal microglia.

Induction of HLA expression in Daudi cells after cell fusion

The Daudi cell line, established from a Burkitt lymphoma, has recently been found to be HLA- andΒ2-microglobulin-negative, although it expresses B lymphocyte alloantigens. This report is concerned with the reexpression of HLA-A10, B38, and B17 on the Daudi cell, after cell fusion with another human cell line (Raji) or with mouse fibroblasts. In the latter fusion, the same HLA specificities are re-expressed, but not humanΒ2-microglobulin while mouseΒ2-microglobulin andH-2 could be detected. No such reexpression was observed when Daudi was fused with the F9 mouse teratocarcinoma, which lacks mouseΒ2-m andH-2. No HLA activity (alloantigenic and xenogenic activity) was detected in the membrane or cytoplasm of Daudi, using salt extraction and sonication. Therefore we postulate thatΒ2-microglobulin could be necessary for the expression and possible synthesis of the HLA antigen.

Role of IL-12 Receptor β1 in Regulation of T Cell Response by APC in Experimental Autoimmune Encephalomyelitis

IL-12 was thought to be involved in the development of experimental autoimmune encephalomyelitis (EAE), a Th1 cell-mediated autoimmune disorder of the CNS. However, we have recently found that IL-12 responsiveness, via IL-12Rβ2, is not required in the induction of EAE. To determine the role of IL-12Rβ1, a key subunit for the responsiveness to both IL-12 and IL-23, in the development of autoimmune diseases, we studied EAE in mice deficient in this subunit of IL-12R. IL-12Rβ1−/− mice are completely resistant to myelin oligodendrocyte glycoprotein (MOG)-induced EAE, with an autoantigen-specific Th2 response. To study the mechanism underlying this Th2 bias, we cocultured purified CD4+ T cells and APCs of MOG-immunized mice. We demonstrate that IL-12Rβ1−/− APCs drive CD4+ T cells of both wild-type and IL-12Rβ1−/− mice to an Ag-induced Th2 phenotype, whereas wild-type APCs drive these CD4+ T cells toward a Th1 type. IL-12Rβ1−/− CD4+ T cells, in turn, appear to exert an immunoregulatory effect on the capacity of wild-type APCs to produce IFN-γ and TNF-α. Furthermore, decreased levels of IL-12p40, p35, and IL-23p19 mRNA expression were found in IL-12Rβ1−/− APCs, indicating an autocrine pathway of IL-12/IL-23 via IL-12Rβ1. IL-18 production and IL-18Rα expression are also significantly decreased in IL-12Rβ1−/− mice immunized with MOG. We conclude that in the absence of IL-12Rβ1, APCs play a prominent regulatory role in the induction of autoantigen-specific Th2 cells.

IL-27 subunits and its receptor (WSX-1) mRNAs are markedly up-regulated in inflammatory cells in the CNS during experimental autoimmune encephalomyelitis

IL-27 (EBI3p28) is a recently discovered heterodimeric cytokine, which is functionally related to IL-23p40p19 and IL-12p40p35. IL-27 acts in synergy with IL-12 early during Th1 development from naive T cells. IL-27 functions through the WSX-1 and the gpl30 receptor subunits, which shares homology with the IL-12Rbeta2 subunit. We have previously reported that IL-23 is up-regulated in CD11b+ microglia/macrophages in the CNS during the early phase of experimental autoimmune encephalomyelitis (EAE), and thus may contribute to the early induction of EAE. In the present study, we examined the expression of IL-27 and its receptor in the CNS, spleen, and lymph nodes at different stages of EAE actively induced with myelin oligodendrocyte glycoprotein peptide(35-55). Our findings show that IL-27 EBI3 and p28 mRNA were up-regulated to a maximum level at the peak of disease in APC from the CNS and lymph nodes, but not in the spleen. Moreover, IL-27 receptor (WSX-1) expression was greatly up-regulated during the early stage of EAE in both the CNS and lymph nodes. Taken together, our data show that subunits of IL-27 and its receptor (WSX-1) mRNAs are markedly up-regulated in inflammatory cells in the CNS at the peak of disease. Thus, IL-27 produced by infiltrating cells in the CNS may regulate in a paracrine manner the Th1 response in EAE.

Positive and negative acute phase proteins in affective subtypes

BACKGROUND Patients with affective disorders show evidence of increased positive acute phase proteins (e.g., C-reactive protein [CRP], alpha-1-acid glycoprotein, haptoglobin) and decreased negative acute phase proteins (e.g., albumin, transferrin [TFN]). CRP reductions have been reported to be greater in patients who later respond to lithium augmentation, and these patients also demonstrate higher CRP levels on the failed antidepressant, prior to the addition of lithium. However, association of such systemic immune changes with affective subtypes, mood state, psychotropic medications, age and gender has not been extensively explored. METHODS The present study assessed levels of CRP and TFN in 79 bipolar I, 24 bipolar II, and 46 unipolar depressed outpatients in comparison to 22 healthy controls. RESULTS Patients on lithium monotherapy were significantly less likely to demonstrate elevated CRP, and a similar trend was noted in those patients taking lithium in combination with an antidepressant. The frequency of elevated CRP levels did not significantly vary for different psychotropic medications, affective subgroups, or mood states. TFN levels were not influenced by diagnosis, affective state or psychotropic medications. LIMITATIONS Due to the retrospective nature of this analysis, the affective subgroups were heterogeneous with regard to medications and affective state, and differed significantly in age. Due to limitations in subgroup sample size, significant effects of clinical variables may have been masked by interactions of medications, age, affective subtype, and mood state. CONCLUSIONS The results imply that lithium may play a role in normalizing systemic immune activation associated with depression. Whether such immune changes may be restricted to lithium-responsive subgroups deserves further evaluation.

Parallel purification of three catalytic subunits of the protein serine/threonine phosphatase 2A family (PP2AC, PP4C, and PP6C) and analysis of the interaction of PP2AC with alpha4 protein ☆

The protein serine/threonine phosphatase (PP) type 2A family consists of three members: PP2A, PP4, and PP6. Specific rabbit and sheep antibodies corresponding to each catalytic subunit, as well as a rabbit antibody recognizing all three subunits, were utilized to examine the expression of these enzymes in select rat tissue extracts. PP2A, PP4, and PP6 catalytic subunits (PP2A(C), PP4(C), and PP6(C), respectively) were detected in all rat tissue extracts examined and exhibited some differences in their levels of expression. The expression of alpha4, an interacting protein for PP2A family members that may function downstream of the target of rapamycin (Tor), was also examined using specific alpha4 sheep antibodies. Like the phosphatase catalytic subunits, alpha4 was ubiquitously expressed with particularly high levels in the brain and thymus. All three PP2A family members, but not alpha4, bound to the phosphatase affinity resin microcystin-Sepharose. The phosphatase catalytic subunits were purified to apparent homogeneity (PP2A(C) and PP4(C)) or near homogeneity (PP6(C)) from bovine testes soluble extracts following ethanol precipitation and protein extraction. In contrast to PP2A(C), PP4(C) and PP6(C) exhibited relatively low phosphatase activity towards several substrates. Purified PP2A(C) and native PP2A in cellular extracts bound to GST-alpha4, and co-immunoprecipitated with endogenous alpha4 and ectopically expressed myc-tagged alpha4. The interaction of PP2A(C) with alpha4 was unaffected by rapamycin treatment of mammalian cells; however, protein serine/threonine phosphatase inhibitors such as okadaic acid and microcystin-LR disrupted the alpha4/PP2A complex. Together, these findings increase our understanding of the biochemistry of alpha4/phosphatase complexes and suggest that the alpha4 binding site within PP2A may include the phosphatase catalytic domain.