氧化型低密度脂蛋白对人肝窦内皮细胞舒张功能及窗孔的影响

Effects of oxidized low-density lipoprotein on relaxation function and fenestrae regulation in human liver sinusoidal endothelial cells

目的：探讨氧化型低密度脂蛋白（oxLDL）对人肝窦内皮细胞（HLSECs）舒张功能和窗孔调节的影响。方法用100 mg/L浓度的oxLDL刺激HLSECs（oxLDL组），培养24 h，另设正常对照组（不经处理）。使用实时定量聚合酶链反应（RT-PCR）和Western blotting法从基因和蛋白水平检测2组HLSECs中内皮型一氧化氮合酶（eNOS）、内皮素1（ET-1）和小凹蛋白（Cav-1）的表达水平，扫描电镜观察100 mg/L oxLDL干预下HLSECs窗孔分布、数量和直径的变化特点。采用t检验进行数据分析。结果与对照组相比，oxLDL组HLSECs ET-1和Cav-1 mRNA表达分别升高了2.62倍和2.49倍（2.62±0.32比1.00±0.00和2.49±0.27比1.00±0.00），而eNOS mRNA表达则下降了45%（0.55±0.12比1.00±0.00）；ET-1和Cav-1蛋白表达升高了2.31倍和2.96倍（1.34±0.10比0.58±0.03和0.71±0.03比0.24±0.02），而eNOS蛋白表达则降低了54%（0.34±0.02比0.74±0.04），两组间差异均有统计学意义（t=32.54、26.62、6.29、16.71、22.09、15.20，均P〈0.05）。与对照组比较，oxLDL组HLSECs窗孔分布稀疏，数量减少，窗孔直径变小，有的区域甚至消失。结论 oxLDL可上调HLSECs ET-1和Cav-1的表达、下调eNOS的表达，引起HLSECs舒张功能失调和发生去窗孔化。

Objective To investigate the effect of oxidized low-density lipoprotein（oxLDL） on the relaxation function and fenestrae regulation in human liver sinusoidal endothelial cells（HLSECs） . Methods Cells were treated with 100 mg/L oxLDL（oxLDL group）. Meanwhile, the normal group was established and untreated. After incubation of HLSECs for 24 h, real time quantitative reverse transcription-polymerase chain reaction （qRT-PCR） and Western blotting were used to detect the mRNA and protein expression of endothelial nitric oxide synthase（eNOS）, endothelin-1（ET-1） and caveolin-1（Cav-1） in HLSECs. The fenestra morphology, number and distribution in the cells were visualized by using scanning electron microscopy. Data were analyzed with t test. Results Compared with the those in control, the mRNA expression of ET-1 and Cav-1 overexpressed by 2.62-and 2.49-folds（2.62±0.32 vs 1.00±0.00 and 2.49±0.27 vs 1.00±0.00）, and the protein expression of ET-1 and Cav-1 overexpressed by 2.31-and 2.96-folds（1.34±0.10 vs 0.58 ± 0.03 and 0.71 ± 0.03 vs 0.24 ± 0.02） in oxLDL group, respectively;the mRNA and protein expression of eNOS significantly decreased by 45%and 54%（0.55 ± 0.12 vs 1.00 ± 0.00 and 0.34 ± 0.02 vs 0.74 ± 0.04） in response to oxLDL versus control（t=32.54, 26.62, 6.29, 16.71, 22.09, 15.20, all P〈0.05）. The fenestra morphology of HLSECs has changed obviously in oxLDL group, oxLDL caused a significant decrease in the porosity, diameter and number of fenestrae compared with those in the control group, even the fenestrae were disappeared in some regions. Conclusions oxLDL can upregulate the expression of ET-1 and Cav-1, downregulate the expression of eNOS. These effects promote the process of relaxation dysfunction and defenstration of HLSECs.