异基因造血干细胞移植术后CD4^+T细胞中缺乏SIRT1诱发急性移植物抗宿主病

SIRT1 deficiency in CD4^+T cells induces acute graft-versus-host disease after allogeneic hematopoietic stem cell transplantation

目的:研究异基因造血干细胞移植患者外周血CD4^+T细胞中SIRT1基因表达水平与急性移植物抗宿主病(acute graft-versus-host disease,a GVHD)的关系。方法:收集40例行同胞全相合异基因造血干细胞移植患者的血液样本,采用实时定量PCR和Western印迹检测各组患者SIRT1的表达水平;采用Western印迹检测各组患者STAT3乙酰化及磷酸化水平;采用免疫共沉淀和Western印迹检测各组患者SIRT1和STAT3结合水平;a GVHD患者CD4^+T细胞过表达SIRT1后采用Western印迹检测STAT3乙酰化及磷酸化水平,采用实时定量PCR检测Th17相关基因RORγt,IL-17A,IL-17F的表达。结果:与未发生a GVHD患者比较,a GVHD患者外周血CD4^+T细胞中SIRT1表达水平明显降低;STAT3表达水平、乙酰化及磷酸化水平均明显升高,且STAT3乙酰化与磷酸化水平呈明显正相关(r=0.69,P<0.01);STAT3与SIRT1结合显著减少。a GVHD患者外周血CD4^+T细胞中过表达SIRT1后,STAT3乙酰化及磷酸化水平均明显降低,RORγt,IL-17A,IL-17F的表达明显减少(P<0.05)。结论:CD4^+T细胞中SIRT1表达不足是异基因造血干细胞移植术后患者STAT3高度乙酰化及磷酸化,介导Th17的相关基因表达升高,进而诱发a GVHD的重要因素。

Objective： To study the relationship between acute graft-versus-host disease （aGVHD） and the SIRT1 expression in peripheral blood CD4^＋T cells from patients after allogeneic hematopoietic stem cell transplantation （allo-HSCT）. Methods： We collected 40 patients who underwent allo-HSCT from human leukocyte antigen （HLA）-identical sibling donors. SIRT1 expression level in CD4^＋T cells was measured by real-time PCR and Western blot. Acetylation and phosphorylation of STAT3 in CD4^＋T cells were detected by Western blot. 1he binding level between SIRT1 and STAT3 in CD4^＋T cells was analyzed by coimmunoprecipitation and Western blot. Over-expression of SIRT1 in aGVHD CD4^＋T cells, as well as STAT3 acetylation and phosphorylation were measured by Western blot. The mRNA levels of RORyt, IL-17A, IL-17F related to Th17 were detected by real-time PCR. Results： SIRT1 expression was significantly down-regulated, while STAT3 expression, acetylation and phosphorylation levels were significantly up-regulated in patients with aGVHD compared with patients without aGVHD. The STAT3 acetylation was positively correlated with STAT3 phosphorylation （r=0.69, P〈0.01）. Less SIRT 1-STAT3 complexes were found in CD4^＋T cells from patients with aGVHD compared with patients without aGVHD. After SIRT1 over-expression in aGVHD CD4^＋T cells, the STAT3 acetylation and phosphorylation, and the expression of RORyt, IL-17A, and IL-17F related to Th17 were significantly down-regulated （P〈0.05）. Conclusion： SIRT1 deficiency in CD4^＋T cells plays a crucial role in up-regulation of STAT3 acetylation and phosphorylation, the increase of Th17 related gene expression, and induction of aGVHD after allogeneic hematopoietic stem cell transplantation.