重组人IL-17A在胃癌组织中的表达及其对胃癌BGC-823细胞增殖、侵袭、迁移和凋亡的影响

Expression of recombinant human IL-17A in gastric cancer tissue and its effects on proliferation, invasion, migration and apoptosis of gastric cancer BGC-823 cells

目的:检测人白细胞介素17A(IL-17A)重组蛋白在人胃癌组织中的表达及其对胃癌细胞增殖、侵袭、迁移和凋亡的影响,并初步探讨其作用机制。方法:收集胃癌患者术后石蜡标本30例、肠上皮化生标本20例和正常胃组织标本10例。免疫组织化学染色法检测各种组织中IL-17A和白细胞介素17受体A(IL-17RA)的阳性表达率,Pearson相关分析法分析IL-17A与IL-17RA表达的相关性。将对数生长期BGC-823细胞分为对照组和不同浓度(50、100和200μg·L-1)IL-17A组。MTT法检测各组胃癌BGC-823细胞增殖活性,流式细胞术检测各组胃癌BGC-823细胞凋亡率,Transwell实验检测各组胃癌BGC-823细胞侵袭细胞数,划痕实验检测各组胃癌BGC-823细胞迁移率,Western blotting法检测各组胃癌BGC-823细胞中血管内皮生长因子(VEGF)、基质金属蛋白酶9(MMP-9)、B细胞淋巴瘤2(Bcl-2)和Bcl-2相关X蛋白(Bax)表达水平。结果:肠上皮化生和胃癌组织中IL-17A和IL-17RA阳性表达率明显高于正常胃组织(P<0.01),胃癌组织中IL-17A与IL-17RA表达呈正相关关系(r=0.891,P<0.01)。IL-17A作用48和72 h后,与对照组比较,100和200μg·L^(-1)IL-17A组胃癌BGC-823细胞增殖活性升高(P<0.05)。与对照组比较,100和200μg·L-1IL-17A组胃癌BGC-823细胞凋亡率明显降低(P<0.01),侵袭细胞数和细胞迁移率升高(P<0.05);Western blotting法检测,与对照组比较,200μg·L^(-1)IL-17A组胃癌BGC-823细胞中VEGF、MMP-9和Bcl-2蛋白表达水平明显升高(P<0.05),Bax蛋白表达水平明显降低(P<0.05)。结论:重组人IL-17A可促进胃癌BGC-823细胞增殖、迁移和侵袭,抑制细胞凋亡,其作用机制可能与促进侵袭相关蛋白VEGF和MMP-9表达以及抑制凋亡相关蛋白Bcl-2和Bax表达有关。

Objective:To detect the expression of human interleukin-17A(IL-17A)recombinant protein in the human gastric cancer tissue and its effects on the proliferation,invasion,migration and apoptosis of gastric cancer cells,and to explore their mechanisms.Methods:Thirty cases of postoperative paraffin specimens of the patients with gastric cancer,20 cases of intestinal metaplasia specimens,and 10 cases of normal gastric tissue were collected.The positive expression rates of IL-17A and interleukin-17 receptor A(IL-17 RA)in different kinds of tissues were detected by immunohistochemical staining.The correlation between the expressions of IL-17A and IL-17 RA was analyzed by Pearson correlation analysis.The BGC-823 cells in logarithmic growth phase were divided into control group and different concentrations(50,100,and 200μg·L^(-1))of IL-17A groups.The proliferation activities of the gastric cancer BGC-823 cells in various groups were detected by MTT method.The numbers of invasion cells of the gastric cancer BGC-823 cells in various groups were detected by Transwell test.The migration rates of the gastric cancer BGC-823 cells in various groups were detected by scratch test,and the expression levels of vascular endothelial growth factor(VEGF),matrix metalloproteinase-9(MMP-9),B cell lymphoma-2(Bcl-2),and Bcl-2 associated X protein(Bax)in the gastric cancer BGC-823 cells in various groups were detected by Western blotting method.Results:The positive expression rates of IL-17A and IL-17 RA in the intestinal metaplasia and gastric cancer tissue were significantly higher than those in normal gastric tissue(P<0.01).There was positive correlation between IL-17A expression and IL-17 RA expression in the gastric cancer tissue(r=0.891,P<0.01).Compared with control group,the proliferation activities of the gastric cancer BGC-823 cells in 100 and 200μg·L^(-1) IL-17A groups were significantly increased after treated with IL-17A for 48 and 72 h(P<0.05).Compared with control group,the apoptotic rates of gastric cancer BGC-823 cells in 100 and 200μg·L^(-1) IL-17A groups were decreased significantly(P<0.01),the numbers of invasion cells and migration rates were increased(P<0.05).The Western blotting method results showed that compared with control group,the expression levels of VEGF,MMP-9 and Bcl-2 proteins in the gastric cancer BGC-823 cells in 200μg·L^(-1) IL-17A group were increased(P<0.05),and the expression level of Bax protein was decreased(P<0.05).Conclusion:The recombinant human IL-17A can promote the proliferation,migration,and invasion of the gastric cancer BGC-823 cells and inhibits the apoptosis,and its mechanism may be related to the promotion of expressions of invasion-related proteins VEGF,MMP-9,and the inhibition of apoptosis-related proteins Bcl-2 and Bax.