Adenovirus-mediated MHC class II transactivator mutant in gene therapy of mouse experimental autoimmune thyroiditis / 第二军医大学学报

ABSTRACT

Objective: To observe the therapeutic effect of major histocompatibility complex (MHC) class II transactivator mutant (C II TAm) for gene therapy of mouse experimental autoimmune thyroiditis (EAT) and explore the possible mechanisms. Methods: Thirty-one healthy female CBA/J mice were randomly divided into 4 groups, namely EAT model group(n= 8), C II TAm therapy group(n=9), GFP control group (n=9), and normal control group(n=5). Animals in the first 3 groups were immunized with porcine thyroglobulin (pTg) and complete or incomplete Freud's adjuvant (CFA/IFA) to establish EAT model; mice in the C II TAm therapy group and GFP control group were also treated by intravenous recombinant adenovirus Ad-CMV-C II TAm and Ad-GFP, respectively, while those in the EAT model group were injected with equal volume of normal saline. Mice in the normal control group received no special treatment. All mice were sacrificed on the 29th day after the first immunization. The thyroid pathological changes were examined using H-E staining; the expression of MHC II molecules in the thyroid was examined using immunohistochemical staining; the spleen lymphocyte proliferation and IFN-γ secretion stimulated by pTg were examined in their culture supernatant; the titer of plasma anti-pTg autoantibody was assayed by ELISA; and the expression of inducible costimulator (ICOS) on CD4+ T cells in both peripheral blood and spleen was analyzed by flow cytometry. Results: H-E staining showed that the infiltration index of thyroid lymphocyte in the C II TAm therapy group (0.3±0.5) was significantly lower than that in the EAT model group (1.4±0.4) and the GFP control group (1.5±0.2, both P<0.01). Immunohistochemical staining showed diffused expression of MHC II molecules in the thyroid of the EAT model group and GFP control group, compared to very weak expression in the C II TAm therapy group and the negative expression in the normal control group. The lymphocyte stimulation index (SI) against 80 μg/ml pTg in the C II TAm therapy group was significantly lower than that in the EAT model group and the GFP control group(P<0.05). The IFN-γ secretion in the culture supernatants showed a similar difference as SI in all the groups (P<0.01). The titer of plasma anti-pTg autoantibody in the C II TAm therapy group was significantly lower than those in the EAT model group and the GFP control group (both P<0.01). The positive rate of ICOS on CD4+ T cells in the C II TAm therapy group was significantly lower than that in the EAT model group and the GFP control group(both P<0.01). Conclusion: Ad-CMV-C II TAm recombinant adenovirus can inhibit the MHC II molecule expression in the thyroid of EAT mouse and the proliferation of self-reactive T cells, attenuate the inflammatory cells infiltration in the thyroid, and decrease the titer of plasma anti-pTg autoantibody, indicating that C II TA mutants might have therapeutic effect for EAT.