Cyclosporine A May Affect the Expression of Foxp3 by Regulation of Akt and Stat3 Phosphorylation in CD4+CD25+ Regulatory T Cells from Mouse Model of Severe Aplastic Anemia

Although Cyclosporine A(CsA) is an effective immunosuppressant in the treatment of aplastic anemia(AA), its affects on CD4+CD25+ regulatory T cells(Tregs) from patients with AA still remains controversial. We have first developed a novel mouse model of severe aplastic anemia(SAA) induced by administration with IFN-γ and busulphan. In this study, the SAA model female mice were intragastrically administered with CsA at daily dose of 5 mg/kg for 10 days(the CsA-treated group, n=60), the SAA group(n=60) and the un-treated group (n=60) were as control. All mice selected to set up the SAA model developed the typical clinical and pathological patterns of SAA from day 10 post dosing. Most of them presented bleedings in association with anemia and infections. At day 15 after treatment, a mortality of 35% was found in the SAA group, while a mortality of 20% was found in the CsA-treated group because of impaired liver function and infections. In order to explore the effect of CsA on CD4+CD25+ Tregs from the novel mouse SAA model, the mononuclear cells of the peripheral blood and spleen were subjected to assess the percentage of CD4+CD25+ Tregs and the intracellular Foxp3 expression in CD4+CD25+ Tregs by flow cytometry(FCM). In the SAA group, the FCM analysis showed decreased ratios of CD4+CD25+ Tregs in CD4+ T cells and down-expression of Foxp3 in CD4+CD25+ Tregs compared with the un-treated group(P<0.05, respectively). These results were in accordance with our previous observations. However, after treatment with CsA, the expression of Foxp3 in CD4+CD25+ Tregs was increased(P<0.05) but not the ratios of CD4+CD25+ Tregs in CD4+ T cells(P>0.05). In addition, after being pured by immunomagnetic beads, the splenic CD4+CD25+ Tregs was subjected to assess the Akt and Stat3 phosphorylation using Western blot. Compared with the un-treated group, CD4+CD25+ Tregs with increased Akt phosphorylation accompanied by increased Stat3 phosphorylation was found in SAA group(P<0.05, respectively). On the contrary, CsA-treated group exhibited CD4+CD25+ Tregs with a reduction in both Akt phosphorylation and Stat3 phosphorylation compared with the SAA group(P<0.05, respectively). These results indicated that CsA may increase the expression of Foxp3 by up-regulation the levels of Akt and Stat3 phosphorylation in CD4+CD25+ Tregs from the novel mice model of severe aplastic anemia.