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目的:利用细胞瞬时转染技术,探讨过表达PDK1和Akt后化瘀解毒方对人胃癌SGC-7901细胞增殖的抑制作用。方法:采用Elisa检测化瘀解毒方提取物对PDK1激酶活力的影响,先后将SGC-7901细胞瞬时转染PDK1质粒和Akt质粒,采用Western blot检测PI3K/Akt通路关键分子p-Akt,Akt活性变化;MTT法检测化瘀解毒方提取物对过表达PDK1或者Akt细胞增殖抑制作用。结果:化瘀解毒方提取物剂量依赖性的抑制PDK1蛋白激酶活性。免疫印迹结果显示过表达PDK1或者Akt可以阻止化瘀解毒方提取物或者LY294002抑制SGC-7901细胞中Akt蛋白的磷酸化。MTT结果显示,化瘀解毒方提取物剂量依赖性的抑制转染空质粒组中SGC-7901细胞增殖。结论:化瘀解毒方通过PI3K/Akt通路抑制胃癌增殖作用。
OBJECTIVE: To investigate the inhibitory effect of Huayu Jiedu Decoction over-expressing PDK1 and Akt on the proliferation of human gastric cancer cell line SGC-7901 by transient transfection. Methods: The effect of Huayu Jiedu extract on PDK1 kinase activity was tested by Elisa. PDK1 plasmid and Akt plasmid were transiently transfected into SGC-7901 cells. The changes of p-Akt and Akt activities of key PI3K / Akt pathway molecules were detected by Western blot MTT assay was used to detect the inhibitory effect of Huayu Jiedu Fang on the proliferation of over-expressed PDK1 or Akt cells. Results: Huayu Jiedu Fang extract dose-dependently inhibited PDK1 protein kinase activity. Western blot results showed that overexpression of PDK1 or Akt could prevent Huayu Jiedu Fang extract or LY294002 to inhibit the phosphorylation of Akt protein in SGC-7901 cells. MTT results showed that Huayujiedu extract dose-dependently inhibited the proliferation of SGC-7901 cells transfected with empty plasmid group. Conclusion: Huayu Jiedu Decoction can inhibit the proliferation of gastric cancer through PI3K / Akt pathway.