化瘀消癥复方靶向MAPK通路诱导滋养细胞自噬性死亡的机制研究

Mechanism Study on Targeted MAPK Pathway Induced Autophagy Induced Death of Trophoblasts by Huayu Xiaozheng Compound

目的基于MAPK通路,探讨化瘀消癥复方对输卵管妊娠滋养细胞自噬性死亡的调控机制。方法选用不同浓度的MAPK信号通路抑制剂SB202190干预滋养细胞,MTS法及免疫印迹法检测MAPK通路抑制对滋养细胞增殖、自噬的影响;联合化瘀消癥复方含药血清干预滋养细胞,MTS法检测细胞增殖活力,免疫印迹法检测自噬、凋亡、MAPK通路相关蛋白的表达,流式细胞术检测细胞凋亡情况。结果与0μmol/L SB202190比较,5、10、20μμmol/L的药效浓度均可抑制HTR-8/SVneo细胞增殖,并呈现浓度依赖性;5、10μμmol/L的SB202190可上调LC3-Ⅱ/LC3-Ⅰ的表达,下调P62的表达(P<0.05)。化瘀消癥复方含药血清联合SB202190干预后,可显著抑制HTR-8/SVneo细胞增殖;诱导凋亡(P<0.05)。免疫印迹结果显示,联合用药组LC3-Ⅱ/LC3-Ⅰ、Bax表达上调,P62、Bcl-2表达下降,p-MAPK、p-ERK蛋白表达下调(P<0.05)。结论化瘀消癥复方含药血清通过抑制MAPK通路,抑制HTR-8/Svneo细胞的活力,诱导妊娠滋养细胞自噬性死亡,发挥治疗异位妊娠的效果。

Objective Based on the MAPK pathway,to explore the regulatory mechanism of Huayu Xiaozheng compound on auto-phagic death of tubal pregnancy trophoblast cells.Methods Different concentrations of MAPK signaling pathway inhibitor SB202190 were selected to intervene in trophoblast cells,and the effects of MAPK pathway inhibition on trophoblast cell prolifera-tion and autophagy were detected by MTS and immunoblotting;The combination of Huayu Xiaozheng compound containing medici-nal serum intervenes in nourishing cells,MTS method is used to detect cell proliferation activity,immunoblotting method is used to detect the expression of autophagy,apoptosis,and MAPK pathway related proteins,and flow cytometry is used to detect cell apoptosis.Results Compared to 0μComparison of M SB202190,5,10,20μThe therapeutic concentration of M can inhibit the proliferation of HTR-8/SVneo cells in a concentration dependent manner;5.10μSB202190 of M can upregulate the expression of LC3-Ⅱ/LC3-Ⅰand downregulate the expression of P62(P<0.05).After the intervention of blood stasis and syndrome elimination compound containing serum combined with SB202190,it can significantly inhibit the proliferation of HTR-8/SVneo cells;Induce apoptosis(P<0.05).The immunoblotting results showed that the expression of LC3-II/LC3-I and Bax was up-regulated in the combination therapy group,while the expression of P62 and Bcl-2 was decreased.The expression of p-MAPK and p-ERK proteins was down regulated(P<0.05).Conclusion The medicinal serum of Huayu Xiaozheng compound can in-hibit the MAPK pathway,inhibit the activity of HTR-8/Svneo cells,induce autophagic death of gestational trophoblast cells,and exert therapeutic effects on EP.