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目的观察肺瘤平膏含药血清对A549细胞与巨噬细胞共培养条件下上皮间质转化(epithelialmesenchymal transition,EMT)相关mRNA及蛋白表达的影响。方法制备肺瘤平膏含药血清,建立A549细胞与巨噬细胞的共培养体系,将细胞分为A549加空白血清组(A549+NRS组)、共培养加空白血清组(共培养+NRS组)及共培养加肺瘤平膏含药血清组(共培养+FLP组)。采用荧光定量PCR、免疫荧光法比较各组A549细胞EMT相关基因(SNAIL1、SNAIL2、ZEB1、CDH1、CDH2、VIM、TJP1、CLDN1、CTNNB1、FLRT1)及E-cadherin、N-cadherin、ZO-1、Vimentin蛋白表达水平的影响。结果共培养条件下,A549细胞变为间充质细胞样。加入肺瘤平膏含药血清后,部分A549细胞形态恢复为上皮细胞样。与A549+NRS组比较,共培养+NRS组A549细胞SNAIL1、ZEB1、CTNNB1、FLRT1、CDH2、VIM mRNA表达上调,TJP1 mRNA表达下调(均P<0.05);与共培养+NRS组比较,共培养+FLP组SNAIL2、TJP1、CLDN1、CDH1、VIM mRNA表达上调,CTNNB1、FLRT1及CDH2 mRNA表达下调(均P<0.05)。免疫荧光结果显示,E-cadherin表达于细胞膜和胞浆内,以细胞膜为主;N-cadherin表达于细胞膜和胞浆内,以细胞浆为主;Vimentin表达于胞浆内;ZO-1表达于细胞连接处为主,部分细胞膜染色亦呈阳性。与A549+NRS组比较,共培养+NRS组A549细胞E-cadherin蛋白表达下调,N-cadherin及Vimentin蛋白表达上调(P<0.05);肺瘤平膏含药血清可上调E-cadherin表达,下调N-cadherin及Vimentin蛋白表达(均P<0.05)。结论肺瘤平膏可抑制共培养条件下A549细胞EMT的形成。
Objective To observe the effect of HGF on the expression of mRNA and protein of epithelialmesenchymal transition (EMT) under co-culture of A549 cells and macrophages. Methods The drug-containing serum of Pneumocystis preparations was prepared and the co-culture system of A549 cells and macrophages was established. The cells were divided into A549 plus blank serum group (A549 + NRS group), co-culture plus blank serum group (co-culture + NRS group ) And co-cultured with lung tumor plaster containing serum group (co-culture + FLP group). The expression of EMT related genes (SNAIL1, SNAIL2, ZEB1, CDH1, CDH2, VIM, TJP1, CLDN1, CTNNB1, FLRT1) and E-cadherin, N-cadherin and ZO-1 in A549 cells were detected by real- Vimentin protein expression levels. Results Under co-culture conditions, A549 cells became mesenchymal-like. After adding lung cancer Ping cream containing drug-containing serum, some A549 cells recovered to epithelial cells. Compared with A549 + NRS group, the expression of SNAIL1, ZEB1, CTNNB1, FLRT1, CDH2 and VIM mRNA were up-regulated and TJP1 mRNA expression was down-regulated in A549 + NRS group (all P < The mRNA expressions of SNAIL2, TJP1, CLDN1, CDH1 and VIM in FLP group were up-regulated while the expressions of CTNNB1, FLRT1 and CDH2 mRNA were down-regulated (all P <0.05). Immunofluorescence results showed that E-cadherin was mainly expressed in the cell membrane and the cytoplasm, mainly in the cell membrane. N-cadherin was mainly expressed in the cytoplasm and the cytoplasm, mainly in the cytoplasm. Vimentin was expressed in the cytoplasm. ZO-1 was expressed in the Cell junctions mainly part of the cell membrane staining was also positive. Compared with A549 + NRS group, the expression of E-cadherin protein and N-cadherin protein and Vimentin protein in A549 cells were decreased in co-cultured + NRS group (P <0.05) N-cadherin and Vimentin protein expression (all P <0.05). Conclusion Feiliuping cream can inhibit the formation of EMT in A549 cells under co-culture conditions.