Toll样受体4与经典Wnt信号在破骨样细胞中作用机制的初步研究  

A preliminary study on the interaction between Toll-like receptor 4 and canonical Wnt signaling pathway in osteoclast-like cells

作　　者：白庆霞[1] 杨博[1] 张艳[1] 刘晓静[1] 陆群[1] 

机构地区：[1]第四军医大学口腔医学院,陕西西安710032

出　　处：《牙体牙髓牙周病学杂志》2013年第3期157-161,共5页Chinese Journal of Conservative Dentistry

基　　金：国家自然科学基金资助项目(31271030)

摘　　要：目的:观察Toll样受体4(TLR4)与经典Wnt信号在破骨样细胞中的相互作用。方法:体外分离培养大鼠破骨样细胞,经抗酒石酸酸性磷酸酶染色、形态学观察鉴定后,随机分为6组:空白对照组(正常培养液)、LPS组、Wnt3a组、DKK1组、LPS+DKK1组、LPS+Wnt3a组,分别与破骨样细胞共同培养24 h后;利用RT-PCR分别检测破骨样细胞中TLR4、破骨相关酶酒石酸酸性磷酸酶(TRAP)、基质金属蛋白酶9(MMP9)、组织蛋白酶K(cathin K)以及经典Wnt信号通路中主要信号分子β-catenin、LRP-6 mRNA表达水平。结果:体外分离培养的大鼠破骨样细胞数目多,状态良好。Wnt3a能下调细胞内破骨相关标志酶mRNA的表达水平、DKK1则上调其mRNA的表达水平(P<0.05);但两者对TLR4 mRNA的表达水平均无明显作用(P>0.05)。LPS可激活破骨样细胞中TLR4的表达,且可上调破骨样细胞内各破骨相关酶mRNA的表达水平;而下调Wnt信号分子β-catenin、LRP-6 mRNA的表达水平(P<0.05)。将Wnt3a、DKK1、LPS+Wnt3a、LPS+DKK1分别作用破骨样细胞24 h后,与LPS单独刺激相比,LPS联合Wnt3a作用后对相关酶表达的下调作用明显降低(P<0.05);相反,LPS联合DKK1作用后对相关酶表达的上调作用进一步增强(P<0.05)。结论:TLR4可通过抑制经典Wnt信号而促进破骨细胞内破骨相关酶的活性;同样,经典Wnt信号受到TLR4的调节后也能反向抑制TLR4的激活而降低破骨细胞内酶的活性。AIM： To study the interaction between Toll-like receptor 4 （TLR4） and canonical Wnt signa-ling pathway in osteoclasts-like cells. METHODS： Primary rat osteoclast-like cells were cultured and identified by tartrate resistant acid phosphatase（TRAP） staining and morphological observation. The cells were then stimulated with LPS （10 g/mL）, Wnt3a （100 ng/mL）, DKK1 （200 ng/mL）, LPS plus Wnt3a and LPS plus DKK1 respectively. Cells without stimulation served as the controls. 24 hours after stimulation, the mRNA expression levels of TRAP, MMP-9, cathin K, TLR4 and LRP6, β-catenin were examined by RT-PCR. RESULTS： Wnt3a stimulation de-creased MMP-9, TRAP and Cathin K mRNA expression （ P 〈 0.05 ）. DKK1 stimulation increased MMP - 9, TRAP and CathinK mRNA expression （P 〈 0.05 ）. However, neither Wnt3a nor DKK1 showed significant influence on TLR4 expression （P 〉 0.05 ）. LPS increased TLR4, TRAP, MMP-9 and cathin K mRNA expression （P 〈 0.05 ）, de-creased β-catenin and LRP6 mRNA expression （P 〈 0.05 ）. Co-stimulation of LPS plus Wnt3a attenuated the effects of LPS （P 〈 0.05）. Co-stimulation of LPS plus DKK1 significantly augmented the effects of LPS （P 〈 0.05）. CON- CLUSION： TLR 4 can promote osteoclastic activity of osteoclast-like cells via inhibition of Wnt signaling pathway.

关 键 词：破骨样细胞 脂多糖 TOLL样受体4 Β-CATENIN 

分 类 号：R780.2[医药卫生—口腔医学]