Effect of silencing CTLA4 with siRNA on the Th1/Th2 cytokines in peripheral blood of chronic hepatitis B patient

Abstract

Abstract: AIM: To explore the inhibition of expression for cytotoxic T-lymphocyte antigen 4(CTLA-4) in peripheral blood lymphocytes in patients with CHB by chemically synthesized small interfering RNA (siRNA) and the secretion of IFN-γ, IL-2, IL-4, then to analyze the T cellular immune activation by the interference of CTLA-4 gene.METHODS: CTLA-4 of peripheral blood lymphocytes in patients with CHB was detected to observe the correlation with the HBV-DNA; according to human CTAL-4 gene sequence, CTAL-4 siRNA and the negative control (siRNA-co) were designed, synthesized, and then they were transfected into peripheral blood lymphocytes separated from CHB patients by electroporation. The expression of CTLA-4 mRNA was detected by real-time Q-PCR, the CTAL-4 protein was measured by Western Blot and the secretion of IFN-γ,IL-2 and IL-4 were assayed by enzyme-linked immunosorbent assay (ELISA).RESULTS: CTLA-4 of peripheral blood lymphocytes in patients with CHB was correlated to HBV-DNA (P＜0.05); after CTLA-4 were transfected into peripheral blood lymphocytes separated from CHB patients, the expression of CTLA-4 mRNA, CTAL-4 protein were suppressed and the production of IFN-γ, IL-2 were enhanced, compared with the negative control group (P＜0.01), while IL-4 concentrations did not show any significant differences (P＞0.05).CONCLUSION: CTLA-4 of peripheral blood lymphocytes in patients with CHB might contribute to the persistent infection of HBV. Inhibiting the expression of CTLA-4 mRNA in peripheral blood lymphocytes in patients with CHB could up-regulate the secretion of Th1-type cytokines IFN-γ, IL-2 except Th2-type cytokines IL-4. It suggested that inhibiting CTAL-4 is helpful to activating T cellular immunity of patients with CHB.

Key words: CHB, CTLA-4, siRNA, Th1/Th2

CLC Number:

R542.96

YU Yong-sheng, WU Hao, TANG Zheng-hao, ZANG Guo-qing. Effect of silencing CTLA4 with siRNA on the Th1/Th2 cytokines in peripheral blood of chronic hepatitis B patient[J]. Chinese Journal of Clinical Pharmacology and Therapeutics, 2011, 16(5): 553-557.

References

[1] Bocher WO, Galun E, Marcus H, et al. Reduced hepatitis B virus surface antigen-specific Th1 helper cell frequency of chronic HBV carriers is associated with a failure to produce antigen-specific antibodies in the trimera mouse [J]. Hepatology, 2000, 31(2): 480-487.
[2] Priimiagi LS, Tefanova VT, Tallo TG, et al. Th1-cytokines in chronic hepatitis B and C[J]. Voprosy Virusologii, 2002, 47(3): 23-27.
[3] June CH, Ledbetter JA, Linsley PS, et al. Role of the CD₂₈ receptor in T-cell activation[J]. Immunol Today, 1990, 11(6): 211-216.
[4] Sharpe AH, Freeman GJ. The B7-CD₂₈ superfamily[J]. Nat Rev Immunol, 2002, 2(2): 1l6-126.
[5] Alegre ML, Frauwirth KA, Thompson CB. T-cell regulation by CD₂₈ and CTLA-4[J]. Nat Rev Immunol, 2001, 1(3): 220-228.
[6] Reuben JM, Lee BN, Li C, et al. Biologic and immunomodulatory events after CTLA-4 blockade with ticilimumab in patients with advanced malignant melanoma[J]. Cancer, 2006, 106(11): 2437-2444.
[7] Annunziato F, Cosmi L, Liotta F, et al. Phenotype, localization, and mechanism of suppression of CD₄⁺CD₂₅⁺ human thymocytes[J]. J Exp Med, 2002, 196(30): 379-383.
[8] Nasta F, Corinti S, Bonura A, et al. CTLA-4 regulates allergen response by modulating GATA-3 protein level per cell [J]. Immunology, 2007, 121(1): 62-70.
[9] Das G, Augustine MM, Das J, et al. An important regulatory role for CD₄⁺CD₈ αα T cells in the intestional epithelial layer in the prevention of inflammatory bowel disease[J]. Proc Natl Acad Sci USA, 2003, 100(9): 5324-5329.
[10] Zhou C, Peng G, Jin X, et al. Vaccination with a fusion DNA vaccine encoding hepatitis B surface antigen fused to the extracellular domain of CTLA4 enhances HBV-specific immune responses in mice: implication of its potential use as a therapeutic vaccine[J]. Clin Immunol, 2010, 137(2): 190-198.
[11] Schurich A, Khanna P, Lopes AR, et al. Role of the co-inhibitory receptor CTLA-4 on apoptosis-prone CD₈ T cells in persistent HBV infection [J]. Hepatology, 2011; 25(5):1494-1503.