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目的:通过研究黄芩素对人类早孕期绒毛滋养细胞株HTR-8/SVneo的增殖活力、侵袭能力的影响,探讨中药黄芩作用于滋养细胞侵袭相关疾病的可能作用机制。方法:体外培养早孕期绒毛滋养细胞株HTR-8/SVneo细胞,以不同浓度(0.01,0.05,0.1,0.5,1μmol·L~(-1))黄芩素作用于细胞,设黄芩素0μmol·L~(-1)为空白组,采用细胞增殖、毒性活力实验法(cell counting kit-8,CCK-8)检测黄芩素对HTR-8/SVneo细胞增殖活力的影响,细胞侵袭实验(Transwell)检测黄芩素对HTR-8/SVneo细胞侵袭作用的影响。提取黄芩素处理后的细胞总RNA,蛋白,分别运用实时荧光定量PCR(Real-time PCR),蛋白免疫印迹法(Western blot)检测滋养细胞侵袭相关的重要因素-基质金属蛋白酶(MMP)-2,MMP-9的mRNA,蛋白表达水平;收集细胞上清,培养基采用明胶酶谱法检测黄芩素对MMP-2,MMP-9的分泌及酶活性的影响。结果:与空白组比较,0.01,0.05,0.1,0.5μmol·L~(-1)黄芩素能明显诱导增强滋养细胞HTR-8/SVneo的侵袭能力(P<0.05),且侵袭率对剂量呈依耐性增长。0.01,0.05,0.1,0.5μmol·L~(-1)黄芩素可明显上调MMP-9蛋白和mRNA表达水平,增强其酶活性(P<0.05)。结论:黄芩素能诱导增强滋养细胞HTR-8/SVneo的侵袭能力,可能是通过增强MMP-9 mRNA和蛋白表达及酶活性来发挥作用。
OBJECTIVE: To investigate the effects of baicalein on the proliferation and invasion of HTR-8 / SVneo cells in human first trimester of pregnancy, and to explore the possible mechanism of Scutellaria baicalensis Georgi on the diseases associated with trophoblast invasion. Methods: HTR-8 / SVneo cells were cultured in vitro. Cells were treated with different concentrations of baicalein (0, 0, 0, 1, ~ (-1) as blank group, cell proliferation and toxicity assay (cell counting kit-8, CCK-8) were used to detect the effect of baicalein on proliferation of HTR-8 / SVneo cells. Transwell assay Effect of baicalein on invasion of HTR-8 / SVneo cells. Total RNA and protein were extracted from the cells treated with baicalein. Real-time PCR and Western blot were used to detect the expression of matrix metalloproteinase (MMP) -2 , The mRNA and protein expression of MMP-9. The supernatant of the cells was collected and the culture medium was used to detect the effect of baicalein on the secretion and enzyme activity of MMP-2 and MMP-9 by gelatin zymography. Results: Compared with the blank group, baicalein 0.01, 0.05, 0.1 and 0.5μmol·L -1 could significantly enhance the invasiveness of HTR-8 / SVneo cells (P <0.05) By patience growth. Baicalein 0.01, 0.05, 0.1 and 0.5μmol·L -1 significantly up-regulated the protein and mRNA expression of MMP-9 and increased the activity of MMP-9 (P <0.05). CONCLUSION: Baicalein can induce the invasiveness of HTR-8 / SVneo cells, which may be related to the enhancement of MMP-9 mRNA and protein expression and enzyme activity.