脂多糖刺激人乳腺上皮细胞SYK及炎症因子的表达变化
Study on the expression of spleen tyrosine kinase and inflammatory factors stimulated by lipopolysaccharide in human mammary epithelial cells
张铭 1仝洁 1刘赛赛 2卢兴浩 3丁倩 3陈云帆 3张小楠 2赵士弟 2杨帆 1陈传好3
作者信息
- 1. 蚌埠医科大学 人体解剖学教研室,安徽 蚌埠 233030;蚌埠医科大学 心脑血管疾病基础与临床重点实验室,安徽 蚌埠 233030
- 2. 蚌埠医科大学 心脑血管疾病基础与临床重点实验室,安徽 蚌埠 233030
- 3. 蚌埠医科大学 人体解剖学教研室,安徽 蚌埠 233030
- 折叠
摘要
目的:探讨脂多糖(LPS)刺激人乳腺上皮细胞中脾酪氨酸激酶(spleen tyrosine kinase,SYK)表达变化及对炎症因子分泌的影响.方法:不同浓度LPS诱导人乳腺上皮细胞产生炎症反应,制备急性乳腺炎细胞模型;通过CCK-8 法检测细胞增殖活性;采用实时荧光定量PCR技术确定LPS最适刺激时间、浓度及该条件下SYK、炎症因子mRNA表达水平;免疫荧光法检测LPS刺激人乳腺上皮细胞后SYK蛋白表达的变化.结果:CCK-8 结果显示,25 μg/mL LPS刺激1h,人乳腺上皮细胞增殖活性无明显变化(P>0.05);实时荧光定量PCR结果显示,25 μg/mL LPS刺激 1h后乳腺上皮细胞SYK基因表达最显著(P<0.05);LPS刺激后,白细胞介素(IL)-1β、IL-6、IL-8、肿瘤坏死因子α和NLRP3 表均增高(P<0.05),siRNA敲低SYK基因后,IL-1β、IL-6、肿瘤坏死因子α和NLRP3 表达均降低(P<0.05).结论:抑制SYK基因可减少LPS诱导的乳腺炎细胞模型炎症因子释放,对于乳腺上皮细胞具有一定保护作用.
Abstract
Objective:To investigate the expression levels of spleen tyrosine kinase(SYK)induced by LPS in human mammary epithelial cells,and its effects on the gene expression of inflammatory cytokine.Methods:The inflammatory response in human mammary epithelial cells were induced by LPS to establish the acute lactation mastitis cell model.The cell proliferation activity was detected by CCK-8 assay.The real-time quantitative PCR was used to obtain the optimal stimulation time and concentration of SYK,and detected the expression levels of related inflammatory cytokine genes.The expression levels of SYK genes in human mammary epithelial cells before and after LPS stimulation were detected by immunofluorescence.Results:The results of CCK-8 showed that there was no significant change in the proliferative activity of human mammary epithelial cells stimulated by 25 μg/mL LPS for 1 h(P>0.05).The results of real-time fluorescence quantitative PCR showed that the expression of SYK gene in mammary epithelial cells was the most significant after 25 μg/mL LPS stimulation for 1 h(P<0.05).After LPS stimulation,the expression levels of interleukin(IL)-1β,IL-6,IL-8,tumor necrosis factor α and NLRP3 increased(P<0.05),and the expression levels of IL-1β,IL-6,tumor necrosis factor α and NLRP3 decreased after the SYK gene knockdown by siRNA(P<0.05).Conclusions:Inhibition of SYK gene can reduce the release of inflammatory factors in LPS-induced mastitis cell models,and has a certain protective effect on mammary epithelial cells.
关键词
乳腺炎/脾酪氨酸激酶/脂多糖/炎症因子Key words
mastitis/spleen tyrosine kinase/lipopolysaccharide/inflammatory cytokine引用本文复制引用
基金项目
蚌埠医学院自然科学重点项目(2020byzd031)
博士科研启动基金资助项目(bsdq202005)
出版年
2024
NETL
NSTL
维普
万方数据