喻日成,雷闽湘,陈卓雄
YU Ri-cheng, LEI Min-xiang, CHEN Zhuo-xiong (Department of Endoclinology

• 1:中南大学湘雅医院内分泌科
• 2:中南大学湘雅医院内分泌科
• 3:中南大学湘雅医院内分泌科 湖南长沙410008
• 4:湖南长沙410008

摘要(Abstract)：

目的:探讨溶血卵磷脂(LPC)对牛视网膜微血管内皮细胞(BRECs)凋亡及细胞内钙离子浓度的影响。方法:原代分离培养牛视网膜微血管内皮细胞,加入不同浓度的LPC,分别培养6h、12h、24h,通过吖啶橙染色在荧光显微镜下观察细胞凋亡,流式细胞检测细胞凋亡率,F2000荧光光度计测细胞内钙离子浓度,并计算游离钙浓度。结果:(1)60μmol/LLPC作用牛视网膜微血管内皮细胞24h后,细胞凋亡率明显高于LPC浓度为20μmol/L及40μmol/L时;(2)LPC浓度从20μmol/L增加到60μmol/L过程中,细胞内游离钙离子浓度明显增加,而且细胞内游离钙离子浓度的增加与LPC的作用时间有关,作用时间越长,钙离子浓度越高。结论:LPC能增加细胞内游离钙离子浓度,促使BRECs的凋亡,而且LPC的这种作用具有时间和剂量依赖性。因此,脂代谢紊乱可能是通过诱导BRECs的凋亡而发挥作用的。
Objective: To study whether lysophosphatidylcholine (LPC) can induce apoptosis of bovine retinal endothelial cells (BRECs) in vitro. Methods: BRECs were exposed to LPC at a range of concentrations and cultured in 12-well plates for 6, 12, 24 hours. Apoptosis was examined on acridine orange (AO) stained specimens under fluorescence microscope. Flow cytometry was used to detect apoptosis rate. The intracellular content of calcium was determined with F-2000 type spectrofluorometer, and the concentration of free calcium ion was calculated. Results: After 24 hours, the rates of apoptosis were significantly higher in BRECs received 60μmol/L of LPC than in those that received 20 or 40μmol/L of LPC. The concentration of free calcium ion was obviously increased as the concentrations of LPC rised from 20μmol/L to 60μmol/L, and the concentration of free calcium ion was influenced by the exposure duration of BRECs to LPC, that, the longer the affecting time, the higher the concentration of free calcium ion. Conclusions: LPC has effects on the intracellular concentration of free calcium ion to induce apoptosis of BRECs in a time- and dose-dependent fashion. Lipid metabolic disturbance might function through causing apoptosis of BRECs.

关键词(KeyWords)： 溶血卵磷脂;凋亡;BRECs;细胞内钙离子浓度
lysophosphatidylcholine; apoptosis; bovine retinal endothelial cells; the intracellular concentration of calcium ion

Abstract:

Keywords:

基金项目(Foundation):

作者(Author): 喻日成,雷闽湘,陈卓雄
YU Ri-cheng, LEI Min-xiang, CHEN Zhuo-xiong (Department of Endoclinology

DOI: 10.19367/j.cnki.1000-2707.2006.06.005

参考文献(References)：

• [1]Biljana M,Robert JG,David MN,JoAnn EManson,et al.A prospective study of serum lipids and risk of diabetic Mac-ular Edema in Type1Diabetes[J].Diabetes,2004,(53):2883-2892.
• [2]Larsson LI,Alm A,Lithner F,et al.The association of hy-perlipidemia with retinopathy in diabetic patients aged15-50years in the county of Umea[J].Acta Ophthalmol Scand,1999,77(5):585-591.
• [3]Cusick M,Chew EY.Histopathology and regression of reti-nal hard exudates in diabetic retinopathy after reduction of elevated serum lipid levels[J].Phthalmology,2003,110(11):2126-2133.
• [4]Mandarino U,Finlayson J,Hassell JR.High glucose down regulates glucose transport activity in retinal capillary peri-cytes but not endothelial cells[J].Invest Ophthalmol Vis Res,1994,35(8):964-972.
• [5]Song W,Barth JL,Lu K,et al.Effects of modified low-density lipoproteins on human retinal pericyte survival[J].Ann N Y Acad Sci,2005,(1043):390-395.
• [6]Mizutani M,Kern T,Lorenzi M.Accelerated death of reti-nal microvascular cells in human and experimental diabetic retinopathy[J].Clin Invest,1996,(97):2883-2890.
• [7]刘学政,萧鸿,曲维新,等.糖尿病大鼠视网膜毛细血管细胞凋亡及其凋亡相关基因的表达[J].中华[科杂志,2001,37(1):59-62.
• [8]Beltramo Z,Berrone E,Buttiglieris,at al.Thiamine and benfotiamine prevent increased apoptosis in endothelial cells and pericytes cultured in high glucose[J].Diabetes Metab Res Rev,2004,20(4):330-336.
• [9]Jump DB.Dietary polyunsaturated fatty acids and regula-tion of gene transcription[J].Curr Opin Lipidol.2002,(13):155-164.
• [10]Jump DB.The biochemistry of n-3polyunsaturated fatty acids[J].Biol Chem,2002,(277):8755-8758.
• [11]Christian AS,Christoph RWK,Christine G.Statins pre-vent oxidized low density lipoprotein and Lysophosphati-dy-choline induced proliferation of human endothelial cells[J].Vascular Pharmacology,2004,(41):67-73.
• [12]Dulak J,Jozkowicz A,Dichtl W,et al.Vascular endothe-lial growth factor synthesis in vascular smooth muscle cells is enhanced by7-ketocholesterol and lysophos-phatidylcholine independently of their effect on nitric ox-ide generation[J].Atherosclerosis,2001,(159):325-332.
• [13]Sata M,Walsh K.Endothelial cell apoptosis induced by oxidized LDLis associated with the down-regulation of the cellular caspase inhibitor FLIP[J].Biol Chem,1998,273(50):33103-33106.
• [14]Heiloth A,Heermeier K.Stimulation of NADPH oxidase by oxidized lowdensity lipoprotein induces proliferation of human vascular endothelial cells[J].Am Soc Nephrol,2000,11(10):1819-1825.
• [15]Rymaszewski Z,Szymanski PT.Human retinal Vascular cells differ from umbilical cells in synthetic functions and their response to glucose[J].PSEBM,1992,199(2):183-191.
• [16]Bucala R,Mitchell R,Arnold K,et a1.Identification of the major site of apolipo protein B modification by ad-vanced glycosylation end products blocking uptake by the lowdensity lipoprotein receptor[J].Biol Chem,l995,270(18):10828-10832.

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