缺氧对口腔扁平苔藓角质形成细胞增殖及缺氧诱导因子1α、血管内皮生长因子、基质金属蛋白酶9表达的影响  

Effect of hypoxia on the proliferation and expressions of hypoxia-inducible factor-1α, vascular endothelial growth factor and matrix metalloproteinase-9 in keratinocytes obtained from oral lichen planus lesions

作　　者：王夏夏[1] 唐国瑶[2] 孙红英[1] 

机构地区：[1]复旦大学附属华山医院口腔科,上海200040 [2]上海交通大学医学院附属第九人民医院口腔医学院黏膜科上海市口腔医学研究所上海市口腔医学重点实验室

出　　处：《中华口腔医学杂志》2015年第2期89-94,共6页Chinese Journal of Stomatology

基　　金：国家自然科学基金（81170967、81470736）;上海市科学技术委员会科研计划（13ZR1405300）

摘　　要：目的 探讨缺氧对人口腔扁平苔藓（oral lichen planus,OLP）角质形成细胞增殖及缺氧诱导因子1α（hypoxia-inducible factor-1α,HIF-1 α）、血管内皮生长因子（vascular endothelial growth factor,VEGF）及基质金属蛋白酶9（matrix metalloproteinase-9,MMP-9）表达的影响,以期为OLP发病机制的研究提供新思路.方法 体外分离培养OLP角质形成细胞;使用密闭培养箱模拟缺氧环境,分为常氧组和缺氧组,两组细胞分别培养12、24、36及48 h,采用细胞计数试剂盒8的方法监测缺氧各时点细胞增殖状况,并确定进一步实验干预时间点,每组实验均重复3次;SYBR Green Ⅰ荧光染料实时定量PCR法和蛋白质印迹法分别检测各组细胞HIF-1α、VEGF、MMP-9 mRNA和蛋白表达水平,并采用相关性分析研究VEGF、MMP-9与HIF-1α的相关性.结果 24、36、48 h缺氧组OLP角质形成细胞增殖力（A值）（分别为0.340±0.002、0.415±0.006、0.546±0.006）均显著低于同时间点常氧组（分别为0.431 ±0.001、0.620±0.004、1.022±0.005）,P＜0.01; 24、36、48 h缺氧组VEGF（2.087±0.291、3.189±0.573、5.402±0.563）及MMP-9（2.936±0.500、4.083±0.300、6.374±0.858）的mRNA表达量均显著高于常氧组（P＜0.05）,HIF-1α （0.414±0.093、0.751±0.056、0.875±0.040）、VEGF （0.393±0.046、0.557±0.078、0.767±0.045）及MMP-9 （0.250±0.053、0.384±0.038、0.611 ±0.092）的蛋白表达量均显著高于常氧组（P＜0.05）.HIF-1α与VEGF（r =0.905,P=0.000）及MMP-9（r =0.881,P=0.000）的蛋白表达水平均呈显著正相关.结论 缺氧在一定时间内可抑制OLP角质形成细胞增殖,上调细胞中HIF-1α蛋白、VEGF及MMP-9 mRNA和蛋白的表达;缺氧环境下HIF-1α可能通过调控下游靶基因参与OLP的病情进展.Objective To investigate the effect of hypoxia on the proliferation and expressions of hypoxia-inducible factor-1α（HIF-1α）,vascular endothelial growth factor（VEGF） and matrix metalloproteinase-9（MMP-9） in keratinocytes obtained from oral lichen planus（OLP） lesions.Methods Hypoxia environment was induced by a airtight incubator.Five groups were included,normoxia control group,hypoxia control group（12,24,36,48 h）.The effect of different treatment time of hypoxia on cellular proliferation was determined with cell counting kit-8（CCK-8）.The mRNA and protein expressions of HIF-lα,VEGF and MMP-9 were analyzed respectively by quantitative real-time polymerase chain reaction with SYBR Green Ⅰ and Western blotting.Results The growth activity of keratinocytes obtained from OLP lesions in the hypoxia groups（0.340±0.002,0.415±0.006,0.546±0.006） was reduced than that in control （0.431±0.001,0.620±0.004,1.022±0.005）（P〈0.01）.The mRNA levels of VEGF（2.087±0.291,3.189±0.573,5.402±0.563） and MMP-9（2.936±0.500,4.083±0.300,6.374±0.858） were elevated by hypoxia（P〈0.05）.The protein levels of HIF-lα（0.414±0.093,0.751 ±0.056,0.875±0.040）,VEGF（0.393 ±0.046,0.557±0.078,0.767±0.045） and MMP-9（0.250±0.053,0.384±0.038,0.611±0.092） were all remarkably elevated by hypoxia （P〈O.05）.However,hypoxia had no effect on HIF-1α mRNA expression.The mRNA expression of HIF-1α after hypoxia exposure for 36 h（1.412±0.094） and 48 h（1.417±0.446） was higher than that of control group,however,there was no significant difference.A positive correlation was noted between HIF-1α and VEGF in protein level（r=0.905,P=O.O00）,and the same correlation found between HIF-1α and MMP-9（r=0.881,P=0.000）.Conclusions Hypoxia conditions may inhibit the proliferation of keratinocytes obtained from OLP lesions.Hypoxia conditions can promote the protein expressions of HIF-1α and both the mRNA and protein expression of VEGF and MMP-9 in keratinocytes obtained from

关 键 词：扁平苔藓 口腔 缺氧诱导因子1 Α亚基 基质金属蛋白酶9 角质形成细胞 

分 类 号：R781.5[医药卫生—口腔医学]