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目的:探讨橙皮苷对转化生长因子-β1(transforming growth factor-β1,TGF-β1)诱导的人非小细胞肺癌细胞上皮-间质转化的影响。方法:以A549人非小细胞肺癌细胞系作为研究对象。用MTT法,分别检测橙皮苷对正常培养条件或TGF-β1刺激下细胞增殖的影响。应用2种方法评估橙皮苷对TGF-β1刺激条件下A549细胞的上皮间质转化:圆形度值定量评估细胞的形态变化;双抗体夹心酶联免疫吸附法(enzyme linked-immuno-sorbent assay,ELISA)检测上皮细胞钙黏蛋白(E-cadherin,E-cad)、ɑ-平滑肌肌动蛋白(ɑ-smooth muscle actin,ɑ-SMA)、基质金属蛋白酶抑制因子-1(tissue inhibitors of metalloproteinases-1,TIMP-1)和基质金属蛋白酶-9(metallopreoteinases-9,MMP-9)水平。结果:0~40μmol·L-1的橙皮苷对正常培养条件或TGF-β1刺激后A549细胞的增殖没有显著影响。5 ng·mL-1 TGF-β1能够诱导A549细胞的上皮间质转化:由上皮特征的铺路石状的细胞形态转化为梭形形态,圆形度值减少;E-cad分泌量减少和ɑ-SMA合成量增加。另外,MMP-9分泌量增加,MMP-9/TIMP-1比值增加。在细胞出现形态改变后,应用橙皮苷(20μmol·L-1或40μmol·L-1)能够降低ɑ-SMA、MMP-9水平和MMP-9/TIMP-1比值,增加E-cad水平,对细胞的圆形度值没有影响。结论:橙皮苷减轻TGF-β1诱导的非小细胞肺癌细胞的上皮间质转化程度,对非小细胞肺癌的转移和扩散有一定的抑制作用。
Objective: To investigate the effect of hesperidin on the epithelial-mesenchymal transition of human non-small cell lung cancer cells induced by transforming growth factor-β1 (TGF-β1). Methods: A549 human non-small cell lung cancer cell line as the research object. The effects of hesperidin on the cell proliferation under normal culture conditions or TGF-β1 stimulation were detected by MTT assay. Two methods were used to evaluate the transmetastasis of hesperidin on A549 cells under the stimulation of TGF-β1: the circularity value was used to evaluate the morphological changes of cells; the enzyme linked-immuno-sorbent assay , ELISA) were used to detect the expression of E-cadherin, inhibitors-smooth muscle actin (ɑ-SMA), tissue inhibitors of metalloproteinases- 1, TIMP-1) and metalloproteinase -9 (MMP-9). Results: Hesperidin with 0 ~ 40μmol·L-1 had no significant effect on the proliferation of A549 cells stimulated by TGF-β1 under normal culture conditions. 5 ng · mL-1 TGF-β1 could induce the epithelial-mesenchymal transition of A549 cells: the shape of the epithelium characterized by paving stones transformed into a spindle-shaped morphology with reduced circularity; decreased E-cad secretion and ɑ- SMA synthesis increased. In addition, MMP-9 secretion increased, MMP-9 / TIMP-1 ratio increased. Hesperidin (20μmol·L-1 or 40μmol·L-1) could reduce the ratio of ɑ-SMA, MMP-9 and MMP-9 / TIMP-1 and increase the level of E- No effect on cell circularity value. CONCLUSION: Hesperidin can reduce the degree of epithelial-mesenchymal transition in non-small cell lung cancer cells induced by TGF-β1 and inhibit the metastasis and proliferation of non-small cell lung cancer.