缺氧、高氧对肝星形细胞基质金属蛋白酶Ⅱ表达及活性的影响

Effects of hypoxia and hyperoxia on the regulation of the expression and activity of matrix metalloproteinase-2 in hepatic stellate cell

目的 研究缺氧、高氧对肝星形细胞基质金属蛋白酶Ⅱ (MMP 2 )表达及其活性影响。方法 分离大鼠肝星形细胞 ,在缺氧或高氧条件下培养 ,以免疫细胞化学标记链霉素卵白素生物素(LSAB)法及酶联免疫吸附 (ELISA)法分别检测细胞内MMP 2、基质金属蛋白酶组织抑制因子Ⅱ(TIMP 2 )、膜型基质金属蛋白酶Ⅰ (MT1 MMP)的表达 ,和培养上清中MMP 2、TIMP 2的相对含量 ,并以酶谱法测培养上清MMP 2活力。结果 (1)缺氧培养 12h ,MMP 2表达增高 (缺氧组阳性指数 :5 7±2 0 ;对照组 :3 2± 1 0 ,;P <0 0 1) ,TIMP 2表达则降低 (缺氧组阳性指数 :2 5± 0 7;对照组 :3 6±1 0 ;P <0 0 5 ) ;培养上清中MMP 2酶活性明显降低 (缺氧组总吸光度值 :7 334± 1 92 2 ;对照组 :17 2 77± 7 4 2 4 ;P <0 0 1)。缺氧不同时段 (6、12、2 4h)比较 ,变化以 6h段最明显。 (2 )高氧培养 12h ,上清中MMP 2、TIMP 2蛋白相对含量均高于对照组 ,TIMP 2更明显 (高氧组A450 :0 0 5 0± 0 0 14 ;对照组 :0 0 2 2± 0 0 10 ;P <0 0 1) ,酶活性亦高于对照组 (高氧组总吸光度值 :5 2 5 2± 0 771;对照组 :4 30 4± 1 0 83;P <0 0 5 ) ,高氧组MT1 MMP表达增强。结论 肝星形细胞对氧敏感。缺氧使肝星形细胞MMP 2表达增加 ,该影响在?

Objective To study the effects of hypoxia and hyperoxia on the expression and activity regulation of matrix metalloproteinase 2 (MMP 2) of the hepatic stellate cell (HSC) Methods The expression of MMP 2, tissue inhibitor of matrix metalloproteinase 2 (TIMP 2) and membrane type matrix metalloproteinase 1(MT1 MMP)in cultured rat HSC under hypoxic or hyperoxic conditions were detected with immunocytochemistry(LSAB method), the contents of MMP 2, TIMP 2 in culture supernatant with ELISA and the activity of MMP 2 in supernatant with zymography Results (1)In the situation of hypoxia for 12 h, the expression of MMP 2 increased (hypoxia group positive indexes: 5 7±2 0; control: 3 2±1 0; P <0 01), while TIMP 2 decreased (hypoxia group positive indexes: 2 5±0 7; control: 3 6±1 0; P <0 05) in HSC, and the activity of MMP 2 in supernatant declined obviously (hypoxia group: 7 334±1 922; control: 17 277±7 424; P <0 01) At the different time courses of hypoxia, the change of expression and activity of MMP 2 was most notable at 6 h (2) In the situation of hyperoxia for 12 h, the protein contents of MMP 2, TIMP 2 in supernatant were both higher than those of the control, especially the TIMP 2 (hyperoxia group A 450 : 0 050±0 014; control:0 022±0 010; P <0 01), and so was the activity of MMP 2 (hyperoxia group total A: 5 252±0 771; control: 4 304±1 083; P < 0 05) The expression of MT1 MMP was also increased Conclusions The HSC is sensitive to the oxygen Hypoxia accelerates the expression of MMP 2 and the effect is more marked at the early stage Hyperoxia increases the activity of MMP 2