XiongBiao Wang

A CTLA-4 gene polymorphism at position -318 in the promoter region affects the expression of protein.

CTLA-4 is an important negative regulator of the immune system. The regulation of the CTLA-4 gene (Ctla-4) transcription is poorly understood. A single nucleotide polymorphism (SNP) at −318 in the Ctla-4 promoter region is associated with certain autoimmune diseases. Since the −318 SNP occurs in a potential regulatory region, it is conceivable that the C′ T transition may affect the expression of Ctla-4. In the present study, we show that the −318T allele is associated with a higher promoter activity than the −318C allele (8.13 ± 0.46 vs 6.87 ± 0.49). The presence of the −318T allele may thus contribute to up regulation of the expression of CTLA-4, and consequently represent one mechanism to inhibit exaggerated immune activity.

Regulation of Surface and Intracellular Expression of CTLA-4 on Human Peripheral T Cells

Cytotoxic T‐lymphocyte‐associated antigen (CTLA‐4) is an important downregulator of T‐cell activation. In order to analyze the expression and regulation of CTLA‐4 on human peripheral T cells, CTLA‐4 mRNA and protein expression were determined using analysis by reverse transcription–polymerase chain reaction (RT–PCR) and FACs, respectively. Intracellular CTLA‐4 was constitutively expressed in unstimulated CD4+ and CD8+ T cells. Interleukin (IL)‐2 induced a dose‐dependent increase of both intracellular and surface expression of CTLA‐4 (CD152). Most of the CD4+ and CD8+ cells expressing CTLA‐4 also expressed CD25. Interferon (IFN)‐γ induced the upregulation of CTLA‐4 expression via antigen‐presenting cells (APC) activation. The CTLA‐4delTM mRNA (550 bp) had a shorter half‐life than the full length CTLA‐4 mRNA and the expression was downregulated upon activation of the cells by treatment with IL‐2. Given an inhibitory role of CTLA‐4 and CD4+ CD25+ T cells in immune responses, the present findings suggest that IL‐2‐induced immunosuppression may result from its stimulatory effect of the CTLA‐4 expression.

CDS1 and promoter single nucleotide polymorphisms of the CTLA-4 gene in human myasthenia gravis.

The cytotoxic T lymphocyte associated protein 4 (CTLA-4) gene (Ctla-4) is a candidate gene for autoimmune disease. We here report results of two single nucleotide polymorphisms (SNPs) in the Ctla-4, a +49 A/G SNP in CDS1 and a C/T promoter SNP at position −318. There were no differences in these two SNPs between patients and healthy individuals. The frequency of allele G and genotype G/G at position +49 in CDS1 was increased in patients with thymoma when compared with patients with normal and hyperplastic thymic histopathology. Patients with the G/G genotype had signs of immune activation manifested as higher levels of serum IL-1β and higher percentage of CD28+ T lymphocytes. There was a strong linkage between the 86 bp allele in the 3′-UTR and the A+49 allele in CDS1. Our results suggest that the SNP at position +49 in CDS1 might be associated with the manifestations of MG.

Mitigated Tregs and Augmented Th17 Cells and Cytokines are Associated with Severity of Experimental Autoimmune Neuritis

Experimental autoimmune neuritis (EAN), an animal model of human Guillain–Barré syndrome, has long been considered as a T helper (Th) 1 cell–mediated autoimmune disorder. However, deficiency of IFN‐γ, a signature Th1 cytokine, aggravated EAN, with features of elevated production of IL‐17A, despite an alleviated systemic Th1 immune response. We hypothesized that Th17 cells and their cytokines might play a pathogenic role in EAN. To further clarify the roles of these Th and regulatory T cell (Treg) cytokines in the pathogenesis of EAN and their interrelationship, we investigated the expression of Th1/Th2/Th17/Treg cytokines in EAN in this study. We found that the levels of Th17 cells and IL‐17A in cauda equina (CE)‐infiltrating cells and splenic mononuclear cells (MNCs) as well as in serum paralleled the disease evolution, which increased progressively during the initiation stage and reached higher value at the peak of EAN. The same pattern was also noticed for the expression of IL‐22. The diverse expression profiles of FoxP3, IL‐17 receptors A and C were seen in CE‐infiltrating cells and splenic MNCs in EAN. These findings indicate a major pro‐inflammatory role of Th17 cells and IL‐17A in the pathogenesis of EAN. Therapeutic interventions may be focused upon inhibiting Th17 cells and their cytokines in the early phase of EAN, so as to delay and suppress clinical signs of the disease, which has relevance for future studies on pathogenesis and treatment of GBS in humans.

Identification of CTLA-4 isoforms produced by alternative splicing and their association with myasthenia gravis.

Myasthenia gravis (MG) is an autoimmune disease characterized by muscle weakness induced by autoantibodies against the acetylcholine receptor. CTLA-4 (CD152) plays an inhibitory role in the immune response and has been suggested to be involved in the pathophysiology of MG. In this study, we focused on alternative CTLA-4 mRNA expression in PBMCs from MG patients. We defined two new isoforms of CTLA-4 mRNA that arise due to alternative splicing. By semi-quantitative RT-PCR analysis, we observed a lower expression of sCTLA-4 mRNA relative to the membrane form in MG patients. In addition, the MG patients had lower levels of sCTLA-4 mRNA in PBMCs compared to healthy controls, as assessed by real-time PCR. One of the spliced isoforms (LCTLA-4) was more prevalent in MG patients compared to healthy controls. The alternative splicing was not associated with sex, thymectomy, serum levels of anti-AChR, immunosuppressive treatment or the four CTLA-4 gene polymorphisms analyzed. This study reveals an abnormal spectrum of mRNA expression of CTLA-4 in MG patients, which marks the importance of studying gene expression of alternative splicing.

Characterization of the expanded T-cell populations in patients with Wegener's granulomatosis

Objectives.  Wegeners granulomatosis (WG) is a chronic inflammatory disease characterized by granulomatosis inflammation, systemic vasculitis and glomerulonephritis. In patients, the peripheral T cells are characterized by mono/oligoclonal CD4+/CD8+ T‐cell AV/BV receptor expansions, with aberrant expression of activation markers. This study was designed to characterize the phenotypic differences between the expanded and nonexpanded T‐cell populations. Expression of markers for activation, costimulation and adhesion molecules was examined. As earlier studies have shown aberrant expression of CD28/CD152, we also analysed the expression of another costimulatory system, the tumour necrotic factor receptor (TNFR) superfamily proteins.

Human Soluble CD80 is generated by alternative splicing, and recombinant soluble CD80 binds to CD28 and CD152 influencing T-cell activation.

CD80 is a costimulatory factor mainly expressed on the surface of activated monocytes, B cells and dendritic cells. In this study, we demonstrate that 24% of healthy individuals have soluble forms of CD80, sCD80, in their serum. The concentration of sCD80 ranged from 0 to 1 mg/l. At the mRNA level, we detected a spliced form s1CD80 (771 bp), in unstimulated monocytes and B cells, while another form named s2CD80 (489 bp) was expressed in activated T cells as well as in freshly isolated and activated monocytes. s1CD80 lacks the transmembrane domain, and the IgC‐like domain plus the transmembrane domain are spliced out of s2CD80. We also present data demonstrating that recombinant s1CD80 binds to recombinant CD152‐Ig and CD28‐Ig. It can also bind to T cells, preferentially to activated T cells. Recombinant sCD80 had immunomodulatory effects shown by its inhibition of the mixed lymphocyte reaction and inhibition of T‐cell proliferation. sCD80 in human serum adds a new member to the family of soluble receptors, implying a network of soluble costimulatory factors with functional relevance. The inhibitory effect of the recombinant protein on T‐cell activation makes it a possible candidate for treatment of diseases associated with hyperactivated T cells.

Single-nucleotide polymorphisms in the B7H3 gene are not associated with human autoimmune myasthenia gravis

Single-nucleotide polymorphisms in B7H3 gene are not associated with human autoimmune myasthenia gravis