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目的:探讨健脾化瘀方对肝癌细胞Smad7蛋白及上皮间质转移标志蛋白E-cadherin/N-cadherin表达水平及侵袭转移能力的影响。方法:分别以不同浓度(20%,高浓度组;10%,低浓度组)健脾化瘀法中药含药血清及空白对照血清(空白对照组)培养肝癌细胞MHCC97-H,q PCR检测细胞Smad7、Snail、Zeb1、Zeb2 m RNA水平,Western blot检测Smad7、Snail、E-cadherin和N-cadherin蛋白表达水平。划痕法和Transwell小室检测肝癌细胞侵袭/迁移能力。结果:高浓度组Smad7 m RNA水平高于空白对照组(P<0.05),Snail、Zeb2 m RNA水平低于空白对照组(P<0.05)。与空白对照组比较,高浓度组48h的Snail、N-cadherin蛋白表达量明显降低,Smad7、E-cadherin蛋白表达量明显升高(P<0.05)。低浓度组48h的E-cadherin升高,N-cadherin降低(P<0.05)。划痕法结果显示,与空白对照组比较,高、低浓度组肝癌细胞的划痕愈合较慢(P<0.05,P<0.01)。Transwell显示,与空白对照组比较,高、低浓度组MHCC97-H细胞的迁移受抑制(P<0.05)。结论:健脾化瘀法中药能抑制肝癌细胞侵袭转移,可能与上调Smad7蛋白含量及抑制上皮间质转化有关。
Objective: To investigate the effect of Jianpi Huayu Recipe on Smad7 protein and E-cadherin / N-cadherin expression and invasion and metastasis of hepatocellular carcinoma cells. Methods: Liver cancer cells MHCC97-H were cultured in different concentration (20%, high concentration, 10%, low concentration) Chinese medicated serum of Jianpi Huayu method and blank control serum Smad7, Snail, Zeb1 and Zeb2 m RNA were detected by Western blot. The protein expressions of Smad7, Snail, E-cadherin and N-cadherin were detected by Western blot. Scratch assay and Transwell chambers to detect hepatocellular carcinoma cell invasion / migration ability. Results: The levels of Smad7 mRNA in high concentration group were higher than those in blank control group (P <0.05). The levels of Snail and Zeb2 mRNA were lower than those in blank control group (P <0.05). Compared with the blank control group, the expression of Snail and N-cadherin in high concentration group decreased significantly, and the expression of Smad7 and E-cadherin protein increased significantly (P <0.05). E-cadherin increased and N-cadherin decreased in low concentration group at 48h (P <0.05). Scratch assay showed that the wound healing of hepatoma cells in high and low concentration group was slower than that in blank control group (P <0.05, P <0.01). Transwell showed that compared with the blank control group, the migration of MHCC97-H cells in high and low concentration groups was inhibited (P <0.05). Conclusion: Chinese herbal medicine for invigorating the spleen and removing blood stasis can inhibit the invasion and metastasis of hepatocellular carcinoma cells, which may be related to up-regulating Smad7 protein content and inhibiting epithelial-mesenchymal transition.