CD4 T cell loss and Th2 and Th17 bias are associated with the severity of severe fever with thrombocytopenia syndrome (SFTS).

Severe fever with thrombocytopenia syndrome (SFTS) is a newly emerging infectious disease caused by a novel bunyavirus with high mortality. Immune suppression is thought to be crucial in disease progression. However, data on immune responses during SFTS are scarce. This study aimed to evaluate the changes in CD4 T-cell subsets throughout the entirety of infection and analyse their relationships with disease severity in SFTS patients. In parallel with CD4 T-cell depletion, decreased Th1, Th2 and Treg numbers, but comparable Th17-cell numbers, were observed in deceased patients compared with those in surviving patients. Additionally, increased Th2 and Th17-cell percentages in the residual CD4 T-cell population led to aberrant Th2/Th1 and Th17/Treg ratios, which were positively correlated with disease severity. Collectively, our data indicated that CD4 T-cell deficiency, Th2 and Th17 bias were closely correlated with the severity of SFTS, indicating therapeutic potential of early immune interventions to ameliorate disease severity.

Betulin from Hedyotis hedyotidea ameliorates concanavalin A-induced and T cell-mediated autoimmune hepatitis in mice.

Hedyotis hedyotidea has been used in traditional Chinese medicine for the treatment of autoimmune diseases. However, the mechanisms underlying for the effect remain unknown. We previously showed that, among 11 compounds extracted from H hedyotidea, betulin produced the strongest suppressive effect on T cell activation. Here, we examined the hepatoprotective effects of betulin against acute autoimmune hepatitis in mice and the mechanisms underlying the effects. Freshly isolated mouse splenocytes were stimulated with concanavalin A (Con A, 5 µg/mL) in the presence of betulin, the cell proliferation was assessed with CSFE-dilution assay. Mice were injected with betulin (10, 20 mg·kg-1·d-1, ip) for 3 d. One hour after the last injection, the mice were injected with Con A (15 mg/kg, iv) to induce acute hepatitis. Blood samples and liver tissues were harvested at 10 h after Con A injection, and serum transaminase levels and liver histopathology were detected; serum levels of proinflammatory cytokines, hepatic T lymphocyte ratios, and functional statuses of conventional T and NKT cells were also analyzed. Betulin (16 and 32 µmol/L) dose-dependently suppressed the proliferation of Con A-stimulated mouse splenocytes in vitro. In Con A-challenged mice, preinjection with betulin (20 mg·kg-1·d-1) significantly decreased the levels of proinflammatory cytokines IFN-γ, TNF-α and IL-6, and ameliorated liver injury. Furthermore, pretreatment with betulin (20 mg·kg-1·d-1) significantly inhibited the Con A-induced activation of NKT and conventional T cells, and decreased production of proinflammatory cytokines IFN-γ, TNF-α and IL-6 in these two cell populations. Betulin has immunomodulatory effect on overly activated conventional T and NKT cells and exerts hepatoprotective action in mouse autoimmune hepatitis. The findings provide evidence for the use of H hedyotidea and its constituent betulin in the treatment of autoimmune diseases.

Induction of the Epstein-Barr virus latent membrane protein 2 antigen-specific cytotoxic T lymphocytes using human leukocyte antigen tetramer-based artificial antigen-presenting cells.

Cytotoxic T lymphocytes (CTLs) specific for the Epstein-Barr virus (EBV) latent membrane protein 2 (LMP2) antigen are important reagents for the treatment of some EBV-associated malignancies, such as EBV-positive Hodgkin's disease and nasopharyngeal carcinoma. However, the therapeutic amount of CTLs is often hampered by the limited supply of antigen-presenting cells. To address this issue, an artificial antigen-presenting cell (aAPC) was made by coating a human leukocyte antigen (HLA)-pLMP2 tetrameric complex, anti-CD28 antibody and CD54 molecule to a cell-sized latex bead, which provided the dual signals required for T cell activation. By co-culture of the HLA-A2-LMP2 bearing aAPC and peripheral blood mononuclear cells from HLA-A2 positive healthy donors, LMP2 antigen-specific CTLs were induced and expanded in vitro. The specificity of the aAPC-induced CTLs was demonstrated by both HLA-A2-LMP2 tetramer staining and cytotoxicity against HLA-A2-LMP2 bearing T2 cell, the cytotoxicity was inhibited by the anti-HLA class I antibody (W6/32). These results showed that LMP2 antigen-specific CTLs could be induced and expanded in vitro by the HLA-A2-LMP2-bearing aAPC. Thus, aAPCs coated with an HLA-pLMP2 complex, anti-CD28 and CD54 might be promising tools for the enrichment of LMP2-specific CTLs for adoptive immunotherapy.

[Prokaryotic expression of BirA enzyme and identification of the bioactivity of expressed product].

AIM: To construct the expression vector of biotin-protein ligase (BirA enzyme) gene and express the BirA enzyme with bioactivity in E.coli BL-21 (DE3). METHODS: The BirA gene was amplified from E.coli genome by PCR and cloned into pGEX-4T-2 to construct the recombinant plasmid pGEX-BirA. After being verified by DNA sequencing, the fusion protein was expressed under IPTG induction in the E.coli BL-21 (DE3). The expressed product was purified through Glutathione-agarose chromatography column. The enzyme activity of the expressed product was identified by ELISA and Western blot. RESULTS: The recombinant prokaryotic expression vector pGEX-BirA was constructed and the fusion protein GST-BirA was expressed successfully. The results of ELISA and Western blot showed that the purified BirA enzyme biotinylated HLA-A2 peptide complex. CONCLUSION: BirA enzyme with bioactivity is prepared successfully, which can be used for studying the interaction between protein and protein.