摘要
目的研究S100A6蛋白在无乳链球菌(Streptococcus agalactiae)所致新生儿脑膜炎中的作用及其分子机制。方法以人脑微血管内皮细胞(human brain microvascular endothelial cell,HBMEC)为体外试验模型,利用siRNA方法构建HBMEC的S100A6基因敲减株,慢病毒转染技术构建S100A6基因过表达细胞株,进行细菌侵袭试验;Western blot检测无乳链球菌感染后HBMEC内S100A6蛋白表达水平,以及敲低及过表达S100A6基因后细胞内炎症因子蛋白质水平变化;通过小脑延髓池穿刺注射无乳链球菌菌液构建新生SD大鼠细菌性脑膜炎模型,HE染色观察大脑组织病理改变,免疫组化检测脑组织S100A6蛋白表达及分布情况,Western blot及ELISA检测脑脊液中S100A6蛋白表达水平。结果与对照组相比,无乳链球菌感染后HBMEC内S100A6蛋白水平显著升高;S100A6基因敲减后,无乳链球菌对HBMEC的侵袭率显著降低(P<0.01),细胞内TNF-α及IL-6蛋白水平显著升高(P<0.01);而过表达S100A6基因后侵袭率则显著升高(P<0.01),TNF-α及IL-6蛋白水平显著下降(P<0.001)。在新生SD大鼠细菌性脑膜炎模型中,HE染色观察到无乳链球菌感染后脑组织有大量中性粒细胞浸润,免疫组化显示S100A6蛋白大量沉积在脑膜周围,脑脊液中也检测到S100A6蛋白质明显表达。结论S100A6蛋白在介导无乳链球菌感染所致的新生儿脑膜炎中具有重要作用,敲减S100A6基因后可促进细胞内炎症因子的产生,减少无乳链球菌对细胞的侵袭,从而减轻细菌对细胞的损伤;同时,感染后S100A6蛋白在脑组织及脑脊液中大量表达,提示其具有成为脑膜炎诊断标志物的可能。
Objective To explore the role and molecular mechanisms of S100A6 protein in neonatal meningitis caused by Streptococcus agalactiae.Methods Human brain microvascular endothelial cells(HBMECs)were used as an in vitro experimental model,and siRNA was employed to construct S100A6 gene knockdown HBMECs strain.The S100A6 gene overexpression cell line was established by lentiviral transfection method.Western blot was used to detect the expression level of S100A6 protein in HBMECs after Streptococcus agalactiae infection,and the change in intracellular inflammatory cytokine protein levels after S100A6 gene knockdown or overexpression.A neonatal bacterial meningitis model was established by injecting Streptococcus agalactiae suspension into the cisterna magna of neonatal Sprague-Dawley(SD)rats.HE staining was used to observe pathological changes in brain tissue;immunohistochemistry was used to detect the expression and distribution of S100A6 protein in brain tissue;Western blot and ELISA were used to measure S100A6 protein levels in cerebrospinal fluid(CSF).Results Compared with the control group,the intracellular S100A6 protein level in HBMECs increased significantly following Streptococcus cgalactiae infection.After S100A6 gene knockdown,the invasion rate of Streptococcus agalactiae into the HBMECs was significantly reduced(P<0.01),while intracellular TNF-α and IL-6 protein levels were elevated markedly(P<0.01).In contrast,overexpression of S100A6 gene increased the invasion rate(P<0.01)and notably decreased TNF-α and IL-6 protein levels(P<0.001).In the neonatal SD rat bacterial meningitis model,HE staining revealed substantial neutrophil infiltration in brain tissue after Streptococcus agalactiae infection.Immunohistochemistry showed extensive deposition of S100A6 protein around the meninges,and significant expression of S100A6 protein was also detected in CSF.Conclusions S100A6 protein is crucial in mediating neonatal meningitis caused by Streptococcus agalactiae infection.S100A6 gene knockdown promotes the production of intracellular inflammatory cytokines and reduces Streptococcus agalactiae invasion into cells,thereby alleviating bacteria-induced cellular damage.Additionally,the increased expression of S100A6 protein in brain tissue and CSF after Streptococcus agalactiae infection suggests its potential as a diagnostic biomarker for bacterial meningitis.
作者
肖承东
张沐捷
田晓燕
梁嘉欣
苏诗雨
黄愉程
彭亮
Xiao Chengdong;Zhang Mujie;Tian Xiaoyan;Liang Jiaxin;Su Shiyu;Huang Yucheng;Peng Liang(Guangdong Key Laboratory of BioTargeted Diagnosis,Treatment and Rehabilitation,the Fifth Affiliated Hospital of Guangzhou Medical University,Guangzhou 510700,China;Jinyu Laboratory College,Guangzhou Medical University,Guangzhou 510180,China)
出处
《中华微生物学和免疫学杂志》
北大核心
2025年第8期657-663,共7页
Chinese Journal of Microbiology and Immunology
基金
广东省基础与应用基础研究基金企业联合基金面上项目(2023A1515220222)。
