Effects of GCN5 or SOX9 gene knockdown on TGF⁃β1 production in the renal tissue of rats with Thy⁃1 nephritis
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摘要:
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RT-PCR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。
Abstract:
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-1N） 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-1N 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 mRNA 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-1N. Results：LV-shGCN5 or LV-shSOX9 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-1N rats. Conclusion：Knockdown of GCN5 or SOX9 gene can significantly inhibit renal TGF-β1 production of rats with Thy-1N.
