水蛭提取液对体外培养RF/6A的细胞液中MMP-2、IV型胶原含量的影响

Effects of leech extract on MMP-2,type IV collagen expression in RF/6A cultured in vitro

目的初探水蛭提取液对新生血管生长的作用机制。方法采用酶联免疫法、放射免疫法检测不同时间点水蛭提取液对细胞培养上清液中MMP-2、IV型胶原表达的影响。结果 12 h各组间比较,空白组、水蛭组MMP-2的OD值分别为0.137±0.006、0.136±0.007,表达低于凝血酶组0.165±0.018,组间差异均有统计学意义(均为P<0.05)。空白组12 h的MMP-2含量要低于2 h、6 h,差异均有统计学意义(均为P<0.05);水蛭组2 h时MMP-2的分泌高于6 h、12 h,差异均有统计学意义(均为P<0.05)。2 h时空白组IV型胶原的含量(20.67±1.02)ng·mL-1高于凝血酶组(15.58±3.40)ng·mL-1、合药组(15.75±6.23)ng·mL-1,差异均有统计学意义(均为P<0.05);6 h时水蛭组IV型胶原的含量明显高于其他3组,差异均有统计学意义(均为P<0.05);12 h时观察,空白组IV型胶原的含量(21.33±2.75)ng·mL-1,明显高于合药组(16.49±1.59)ng·mL-1,差异有统计学意义(P<0.05),水蛭组IV型胶原的含量明显高于凝血酶组及合药组,差异有统计学意义(P<0.05)。空白组2 h、12 h IV型胶原的含量均较6 h高,差异均有统计学意义(均为P<0.05);凝血酶组IV型胶原的合成呈增加趋势,12 h与2 h相比,差异有统计学意义(P<0.05);水蛭组IV型胶原的合成明显增加,6 h、12 h IV型胶原的含量显著高于2 h,差异有统计学意义(P<0.05)。结论64 g·L-1水蛭提取液可以促进IV型胶原合成,抑制MMP-2的分泌,对新生血管的形成有抑制作用。

Objective To primary study the active mechanism of leech extracts on retinal neovascularization. Methods The euzymelinked immunosorbent assay and radio-immunity were used to study the effects of leech extracts on MMP-2 and type IV collagen expressions in RF/6A cultured in vitro at different time points. Results The OD value of MMP-2 at 12 hours in blank group and leech group were 0. 137±0.006 and 0. 136 ± 0. 007, respectively, which all lower than that in thrombin group （0. 165 ± 0. 018）, there were statistical differences （ all P 〈 0.05 ）. The MMP-2 contents in blank group at 12 hours were lower than those at 2 hours and 6 hours,there were statistical differences （all P 〈 0.05 ）. The MMP-2 contents in leech group at 2 hours were higher than those at 6 hours and 12 hours, there were statistical differences （all P 〈 O. 05 ）. The contents of type IV collagen at 2 hours in blank groups, leech group and mixing group were （20.67 ± 1.02）ng . mL-1 ,（15.58 ±3.40）ng. mL-1 and （15.75 ±6.23）ng . mL-1, respectively,the former were higher than the latter two, there were statistical differ ences （ all P 〈 0.05 ） ; Which at 6 hours in leech group were obviously higher than other three groups,there were statistical differences （ all P 〈 0.05 ） ; Which at 12 hours in blank group and mixing group were （ 21.33 ±2.75 ）ng . mL -1 and （ 16.49 ± 1.59 ）ng .mL-1, respectively,the former was obviously higher than the latter,there was statistical difference （P 〈 0.05 ）, and the leech group were higher than thrombin group and mixing group, there were statistical differences （ all P 〈 0.05 ）. The contents of type IV collagen in blank group at 2 hours and 12 hours were higher than that at 6 hours,there were sta tistical differences （all P〈0.05） ;The formation of type IV collagen in leech group was obviously increased, the contents at 6 hours and 12 hours were higher than that at 2 hours, there were statistical differences （ all P 〈 0.05 ）. conelusion The leech extract （64 g 0 L-1 ） can increase the synthesis of type IV collagen, decrease the secretion of MMP-2, and inhibit the retinal neovascularization.