沉默GCN5或SOX9基因对Thy-1肾炎大鼠肾组织中TGF-β1生成的影响

Effects of GCN5 or SOX9 gene knockdown on TGF-β1 production in the renal tissue of rats with Thy-1 nephritis

目的:研究沉默大鼠肾组织中通用控制核苷酸合成5(general control nonderepressible 5,GCN5)基因、性别决定区Y框蛋白9(SRY related HMG box-9,SOX9)基因对Thy-1肾炎(Thy-1 nephritis,Thy-1N)大鼠肾组织内转化生长因子-β1(transforming growth factor-β1,TGF-β1)生成的影响。方法:分别用慢病毒(lentivirus,LV)包装GCN5和SOX9发夹状小干扰RNA(shRNA),即制备LV-shGCN5和LV-shSOX9重组病毒。然后行大鼠肾动脉灌注术将LV-shGCN5和LV-shSOX9分别导入大鼠肾脏,再经尾静脉注射兔抗大鼠胸腺细胞抗血清(anti-thymocyte serum,ATS)复制Thy-1N模型。在注射ATS后3 h,取大鼠肾组织,用RTPCR和Western blot检查各组大鼠肾组织中GCN5和SOX9的mRNA及蛋白表达水平,以验证干扰效果及GCN5、SOX9对TGF-β1产生的影响。结果:利用肾动脉灌注术将LV-shGCN5或LV-shSOX9重组病毒导入大鼠肾组织后,不仅能有效沉默相应的靶基因,而且还能下调TGF-β1的表达。结论:沉默大鼠肾组织中GCN5或SOX9基因后能显著抑制Thy-1N大鼠肾内TGF-β1的生成。

Objective:This Study aims to study the effect of silencing general control nonderepressible 5(GCN5)or SRY related HMG box-9(SOX9)gene on TGF-β1 production in the renal tissue of Thy-1 nephritis(Thy-1 N)rats.Methods:The plasmids of GCN5 and SOX9 shRNA were packaged with lentiviral(LV)to form LV-shGCN5 and LV-shSOX9.Then,the LV-shGCN5 and LV-shSOX9 vectors were separately perfused into rat kidneys by renal artery perfusion,and next,the rat Thy-1 N model was replicated by injection of rabbit anti-rat-thymocyte serum(ATS)through rat tail vein.Thereafter,rat renal tissues mentioned-above were taken at 3 h after ATS injection,and m RNA and protein levels of GCN5,SOX9 and TGF-β1 were examined by RT-PCR and Western blot to verify the effects of GCN5 or SOX9 gene interference on TGF-β1 expression in the renal tissues of rats with Thy-1 N.Results:LV-shGCN5 or LVshSOX9 vectors could be perfused successfully into rat kidneys through renal artery,and LV-shGCN5 or LV-shSOX9 not only effectively silenced GCN5 or SOX9 gene,but also downregulated TGF-β1 synthesis in the renal tissue of Thy-1 N rats.Conclusion:Knockdown of GCN5 or SOX9 gene can significantly inhibit renal TGF-β1 production of rats with Thy-1 N.