摘要
目的:构建外源过氧化氢酶基因表达载体并使其在晶体上皮细胞中稳定表达,从而提高晶体上皮细胞的抗氧化能力减少白内障的发生。方法:克隆过氧化氢酶基因,测序确定。构建过氧化氢酶基因pIRESneo3表达载体。利用转染的方法将过氧化氢酶基因导入晶体上皮细胞SRA01/04中,筛选稳定表达克隆。以SRA01/04细胞和稳定转染过氧化氢酶基因的SRA01/04细胞为研究对象,使用过氧化氢为处理因素,MTT方法测定过氧化氢对晶体上皮细胞的半数致死量。结果:经上述方法处理后,获得过氧化氢酶高表达晶体上皮细胞SRA01/04-CAT,其过氧化氢酶的表达水平是SRA01/04细胞的3.5倍。过氧化氢对SRA01/04细胞的IC50是32.24μmol/L,对SRA01/04-CAT的IC50是85.32μmol/L,转染过氧化氢酶基因后SRA01/04细胞对过氧化氢的耐受能力提高2.65倍。结论:外源过氧化氢酶基因能够在晶体上皮细胞中稳定表达并使晶体上皮细胞的抗氧化能力提高。
AIM:To construct exterior catalase gene expression vector and make it stable in lens epithelial cells so as to improve the resistance of lens epithelial cells to oxidant damage and reduce the occurance of cataract. METHODS:The coding sequence of catalase gene was cloned and verified by DNA sequencing.pIRESneo3 vector of catalase gene was constructed and introduced in SRA 01/04 lens epithelial cells by transfection, and stable expression clones were screened.The expression of catalase gene in the transfected SRA01/04 cells and untransfected SRA01/04 cells was investigated and treated with hydrogen peroxide.MTT method was used to determine the half lethal dose of hydrogen peroxide to lens epithelial cells. RESULTS:The expression level of catalase in SRA01/04 CAT cells was 3.5 times higher than that in SRA01/04 cells.The IC50 of H2O2 to SRA01/04 CAT was 85.32 μmol/L and the IC50 of H2O2 to SRA01/04 was 32.24 μmol/L.After catalase transfection, tolerance capacity to hydrogen peroxide of SRA01/04 was strengthened by 2.65 folds. CONCLUSION:The catalase gene can stabilize the expression in lens epithelial cells and improve the resistance of lens epithelial cells to oxidant damage.
出处
《中国临床康复》
CSCD
2004年第17期3319-3321,i005,共4页
Chinese Journal of Clinical Rehabilitation
