LPS激活TLR4抑制BMP9诱导iMEFs成骨分化

基础医学与临床 ›› 2017, Vol. 37 ›› Issue (1): 25-31.

LPS激活TLR4抑制BMP9诱导iMEFs成骨分化

郭杨柳¹,陈思成²,李亚¹,范梦恬¹,孙艳婷¹,李汪¹,施琼¹

1. 重庆医科大学临床检验诊断学教育部重点实验室
2. 新乡医学院

收稿日期:2016-08-25 修回日期:2016-09-23 出版日期:2017-01-05 发布日期:2016-12-30
通讯作者: 施琼 E-mail:anniesq878@aliyun.com
基金资助:
国家自然科学基金面上项目;教育部博士点基金;重庆市教育委员会科学技术研究项目;重庆市渝中区科委基金

TLR4 activation with LPS inhibits BMP9-induced osteogenic differentiation of immortalized mouse embryonic fibroblasts

Received:2016-08-25 Revised:2016-09-23 Online:2017-01-05 Published:2016-12-30
Supported by:
the National Natural Science Foundation of China

摘要/Abstract

摘要： 目的研究脂多糖（ lipopolysaccharide，LPS）激活的Toll样受体4（Toll-like receptor 4，TLR4）信号对骨形态发生蛋白9（bone morphogenetic proteins 9，BMP9）诱导永生化小鼠胚胎成纤维细胞(immortalized mouse embryonic fibroblasts，iMEFs)成骨分化的影响。方法细胞免疫荧光检测TLR4/NF-κB信号通路的激活；LPS，BAY11-7082和BMP9处理iMEFs，ALP染色和活性检测iMEFs早期成骨分化能力；茜素红S染色检测晚期成骨分化能力；半定量PCR和Western blot检测晚期成骨基因OCN和OPN表达；Western blot 检测Smad1/5/8磷酸化水平；半定量PCR和Western blot检测成骨关键转录因子Runx2和Dlx5的表达。结果 LPS成功激活TLR4/NF-κB信号通路；LPS抑制BMP9诱导的ALP染色和活性（P<0.01）、钙盐沉积、OCN的mRNA和蛋白质表达（P<0.05）、OPN的mRNA（P<0.01）和蛋白质（P<0.05）表达、Smad1/5/8信号通路激活（P<0.01）、Runx2的mRNA和蛋白质（P<0.05）、Dlx5的mRNA（P<0.01）和蛋白质（P<0.05）表达，BAY11-7082可以部分逆转LPS的抑制作用（P<0.05）。结论 LPS激活TLR4可以通过NF-κB信号通路抑制BMP9诱导的iMEFs成骨分化。

关键词: TLR4, 骨形态发生蛋白9, iMEFs, 成骨分化

Abstract: Objective To study the effect of TLR4 activation with LPS on BMP9-induced osteogenic differentiation of immortalized mouse embryonic fibroblasts(iMEFs). Methods The activation of TLR4/NF-κB signaling pathway was detected by ICC.iMEFs were treated with LPS,BAY11-7082,Adnovirus GFP and BMP9. The early osteogenic differentiation capability ofiMEFs was detected by ALP staining and quantitative assay. The later osteogenic differentiation capability was detected by alizarin red S staining. The expression of later osteogenic differentiation marker gene OCNand OPNwere detected by PCR and Western blot. The change of p-Smad1/5/8 was detected by Western blot. The expression of Runx2and Dlx5 were detected by PCR and Western blot. Results LPS can stimulate TLR4/NF-κB signaling pathway effectively. TLR4 activation inhibited BMP9-induced osteogenic differentiation. BMP9-induced osteogenic differentiation related gene and Smad1/5/8 signaling activation were inhibited by TLR4 activation. The inhibition effect could be partly reversed by BAY11-7082(P<0.05).Conclusions TLR4 activation with LPS can inhibit BMP9-induced osteogenic differentiation of iMEFs cellsvia NF-κB signaling pathway.

Key words: TLR4, BMP9, iMEFs , osteogenic differentiation

中图分类号:

Q254

郭杨柳陈思成李亚范梦恬孙艳婷李汪施琼. LPS激活TLR4抑制BMP9诱导iMEFs成骨分化[J]. 基础医学与临床, 2017, 37(1): 25-31.

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