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目的:检测大鼠视网膜色素上皮细胞系(RPE-J)是否表达载脂蛋白的基因及其是否具有合成和分泌中性脂质的功能。方法:将RPE-J细胞培养于组织培养板及Transwell上,用RT-PCR检测载脂蛋白mRNA的表达。将荧光标记颗粒加入细胞中培养以检测细胞吞噬功能。细胞中加入光感受器细胞外节,培养不同时间点后提取脂质,用薄层层析法和酶学荧光法分析细胞中脂质含量及变化。用放射性标记的OleicAcid([3H]-oleate)检测细胞新合成和分泌中性脂质的功能。结果:RPE-J细胞有载脂蛋白A-I,A-II,B,E,C-I,C-II,C-III,以及微粒体甘油三酯酸转移蛋白基因的表达。RPE-J细胞具有吞噬功能。吞噬光感受器细胞外节后,RPE-J细胞中甘油三酯含量增加5.8倍,磷脂含量增加2.6倍,非酯化胆固醇增加1.5倍,酯化胆固醇增加0.3倍。RPE-J细胞和培养基内可以检测到新合成的放射标记的中性脂质。细胞内脂质含量随时间延长而增加;培养基中只有磷脂随时间延长而增多。结论:RPE-J细胞有多种载脂蛋白及微粒体甘油三酯酸转移蛋白的基因表达。吞噬光感受器细胞外节后,RPE-J细胞中脂质增加。培养基中加入长链脂肪酸后,RPE-J细胞可以新合成和分泌中性脂质。
OBJECTIVE: To detect whether the retinal pigment epithelial cell line (RPE-J) expresses the gene of apolipoprotein and whether it has the function of synthesizing and secreting neutral lipids. Methods: RPE-J cells were cultured on tissue culture plates and Transwell, and the expression of apolipoprotein mRNA was detected by RT-PCR. Fluorescently labeled particles were added to the cells for culture to detect phagocytosis. Cells were added photoreceptor extracellular sections, cultured at different time points after the extraction of lipids, by thin-layer chromatography and enzymatic fluorescence analysis of cell lipid content and changes. Radioactively labeled OleicAcid ([3H] -oleate) was used to examine the function of the cells in the new synthesis and secretion of neutral lipids. Results: The expression of apolipoprotein A-I, A-II, B, E, C-I, C-II, C-III and microsomal triglyceride transfer protein genes were detected in RPE- RPE-J cells have phagocytosis. After phagocytosis of extracellular sections of photoreceptors, triglyceride content in RPE-J cells increased 5.8-fold, phospholipid content increased 2.6-fold, non-esterified cholesterol increased 1.5-fold and esterified cholesterol increased 0.3-fold. Newly synthesized radiolabeled neutral lipids can be detected in RPE-J cells and media. The intracellular lipid content increased with time; only the phospholipid in the medium increased with time. CONCLUSIONS: RPE-J cells have a variety of apolipoprotein and microsomal triglyceride transfer protein gene expressions. Lipophilicity increased in RPE-J cells after phagocytosis of photoreceptor extracellular sections. RPE-J cells can newly synthesize and secrete neutral lipids after adding long-chain fatty acids to the medium.