阳离子脂质体DOTAP介导报道基因转染骨关节病滑膜细胞的实验研究

目的：探索阳离子脂质体ＤＯＴＡＰ介导目的基因转染骨关节病滑膜细胞的最佳条件。方法：分离培养骨关节病滑膜细胞，使用荧光质粒pＥＧＦＰ－Ｃ３作为报道基因，与阳离子脂质体ＤＯＰＡＰ一起转染骨关节病滑膜细胞；通过设立不同转染时间组、不同表达时间组以及不同ＤＯＴＡＰ剂量组，观察绿色荧光蛋白的表达。结果：使用阳离子脂质体ＤＯＴＡＰ介导荧光质粒转染骨关节病滑膜细胞，转染时间在 ６ｈ左右， ＤＯＴＡＰ（μｌ）／ＤＮＡ（μｇ）比值在 ７：１左右，可于转染后４８ｈ左右表达出的蛋白量最大。结论：阳离子脂质体ＤＯＴＡＰ介导荧光质粒转染骨关节病滑膜细胞转染条件的优化，为今后骨关节病滑膜细胞非病毒基因转移的研究提供实验依据。

Experimental Study of Cationic Liposome DOTAP-mediated Report Gene Transfection of Osteoarthritis Synovial Cells

To study the optimization of cationic liposome DOTAP-mediated reporter gene transfection of osteoarthritis(OA) synovial cells, we separated and cultured the knee joint synovial cells from OA patients. Cotransfection of synovial cells ~ conducted using fluorescent Plastal PEGal as a reporter gene with cationic liposome DOTAP. For establishing optimal transfection conditions, we designed the techniques of determining the expression level of a reporter protein by varying experimental parameters,such as different transfection time,different expression time and differ- ent ration of DOTAP/DNA,and observed the expression of enhenced green fluorescent protein (EGFP). The result showed that when transfection time was 6 hours and the ratio of DOTAP(ul)/DNA(ug) was 7: 1,the transfection and expression efficiency of synovial cells about 48h after transfection was the highest. We concluded that the optimal transfection conditions of cationic liposome DOTP-mediated reporter gene transfec- tion of OA synovial cells provide experimental evidence for investigating non-viral gene transfer of OA synovial cells.