AKIP1对结直肠癌细胞凋亡的影响及其分子机制研究

Effect of AKIP1 on Apoptosis of Colorectal Cancer Cells and Its Molecular Mechanism

【目的】探究AKIP1对人结直肠癌细胞HCT-116凋亡的影响及其相关分子机制。【方法】设计AKIP1干扰片段siRNA及NC片段,转染至HCT-116细胞,Real-time PCR及Western Blot检测AKIP1 mRNA及蛋白的表达;通过MTT及流式细胞术检测细胞活力及凋亡情况;通过Real-time PCR检测细胞凋亡相关基因的表达变化,并通过Western Blot检测NF-κB p65在胞浆和胞核中的表达以及凋亡相关蛋白的表达。【结果】转染AKIP1 siRNA后,AKIP1 mRNA及蛋白的表达水平显著下调;MTT实验结果显示AKIP1 siRNA可显著抑制HCT-116细胞的活力;流式细胞术结果表明AKIP1 siRNA可显著促进HCT-116细胞凋亡;同时干扰AKIP1可上调Bax、cleaved-Caspase-3并下调Bcl-2的表达;另外,AKIP1 siRNA可下调细胞核NF-κB p65的表达,上调细胞浆中的NF-κB p65蛋白表达。【结论】AKIP1可能通过活化NF-κB p65抑制HCT-116细胞凋亡进而促进结直肠癌的疾病进展。

【Objective】To investigate the effect and related molecular mechanism of AKIP1 on the apoptosis of human colorectal cancer cells HCT-116.【Methods】AKIP1 siRNA were designed and transfected into HCT-116 cells. Real-time PCR and Western Blot were used to detect the expression of AKIP1 mRNA and protein, respectively. Cell viability and apoptosis were measured by MTT and flow cytometry. We detected the mRNA expression of apoptosis associated genes by Real-time PCR, and the protein expression of NF-κB p65 in the cytoplasm and nucleus and the apoptosis-related proteins were detected by Western Blot.【Results】After transfection with AKIP1 siRNA, the expression levels of AKIP1 mRNA and protein were significantly down-regulated. MTT assay showed that AKIP1 siRNA significantly inhibited the viability of HCT-116 cells. Flow cytometry showed that AKIP1 siRNA significantly promoted apoptosis of HCT-116 cells. Meanwhile, AKIP1 siRNA up-regulated Bax, cleaved Caspase-3, and down-regulated bcl-2 expression. In addition, AKIP1 siRNA down-regulated the expression of NF-κB p65 in the nucleus and up-regulated the expression of NF-κB p65 in the cytoplasm.【Conclusion】AKIP1 may inhibit the apoptosis of HCT-116 cells by activating NF-κB p65 and promote the progression of colorectal cancer.