阿托伐他汀对瘦素诱导血管内皮细胞及平滑肌细胞增殖的作用

Effect of atorvastatin on leptin-induced proliferations of vascular endothelial cells and vascular smooth muscle cells

目的研究阿托伐他汀对瘦素刺激的血管内皮细胞（vascular endothelial cells,VECs）和血管平滑肌细胞（vascular smooth muscle cells,VSMCs）增殖的影响。方法 VECs、VSMCs的密度调整为1×105/m L接种于96孔培养板,每孔均为100μL,将细胞分为两组：（1）不同浓度瘦素（0、12.5、25、50、100 ng/m L）-VSMCs与（0、10、20、50、100 ng/m L）-VECs分别作用24、48、72 h,设150 ng/m L瘦素干预72 h为150 ng/ml瘦素组;（2）先用100 ng/m L瘦素干预细胞72 h,再加入各种浓度的阿托伐他汀（0、1、10、100μmol/L）共培养24 h,设单纯的100 ng/m L瘦素干预细胞72 h为100 ng/m L瘦素组;（3）同时,两组均设正常对照组。刺激细胞至时间点后,于每个孔各滴入噻唑蓝（methyl thiazolyl tetrazolium,MTT）（10 mg/m L）10μL,持续培育4 h。弃培养液后加入200μL二甲基亚砜（DMSO）,小心吹打均匀,完全熔化后于全自动免疫酶标仪中测定的波长为490 nm的吸光度,每组6孔,重复实验3次。结果瘦素刺激组VECs与VSMCs的增殖高于正常对照组,且随着瘦素作用时间延长,VECs与VSMCs表达也随之增加,在72 h末促增殖效应达峰值,并且瘦素浓度在0~100 ng/m L之间呈剂量依赖性关系。100 ng/m L与150 ng/m L瘦素的促增殖效应无明显差异。阿托伐他汀组VECs与VSMCs的促增殖效应低于正常对照组及100 ng/m L瘦素组,且随着阿托伐他汀作用时间的延长,VECs与VSMCs的增殖反而会降低,并且最佳浓度为100μmol/L。结论瘦素能促进VECs与VSMCs的增殖,在一定范围内呈剂量及时间依赖性关系。阿托伐他汀能抑制瘦素诱导VECs与VSMCs的促增殖效应。

Objectives To study the stimulation of atorvastatin on leptin-induced proliferations of vascular endothelial cells（VECs） and vascular smooth muscle cells（VSMCs）. Methods VECs and VSMCs with adjusted densities of 1 ×105/ m L were seeded in 96-well culture plates（each well =100 μL） and divided into two groups ：（1）Different concentrations of leptin（0, 12.5, 25, 50, 100 ng / m L）-VSMCs and（0,10,20, 50,100 ng / m L）-VECs were treated for 24,48,72 h. The 150 ng / m L leptin for 72 h was located as 150 ng / m L leptin group.（2）100 ng / m L leptin intervented for 72 h before adding different concentrations of atorvastatin（0,1,10,100 μmol / L） to co-culture for 24 h.Pure 100 ng / m L leptin for 72 h was located as 100 ng / m L leptin group.（3） At the same time, normal control groups of the two groups were set. After stimulating the cells to the time point, 10 μL of methyl thiazolyl tetrazolium（MTT）（10 mg / m L）was added to each well. Cultivation was continued for 4 h. After aspirating the culture medium,200 μL of dimethylsulfoxide（DMSO） was added. The cells were carefully blown to be uniformed. When they were fully melted,autoimmune microplate reader was used to measure the absorbance spectra（wavelength=490 nm）. There were 6 holes in each group and the experiment was repeated for 3 times. Results Proliferations of VSMCs and VECs stimulated by leptin were higher than those in normal control group, and with the role of leptin prolonged, expressions of VECs and VSMCs also increased. Proliferation effects peaked at the end of 72 h. Leptin concentrations at 0-100 ng / m L had a dose-dependent relationship with the proliferations. Proliferation effects between leptin concentrations at 100 ng / m L and150 ng / m L had no significant difference. Proliferation effects of VECs and VSMCs in atorvastatin group were lower than those in normal control group and 100 ng / m L leptin group. With the effect duration of atorvastatin prolonged,proliferations of VECs and VSMCs decreased, and the best concentration of atorvastatin was at 100 μmol / L.Conclusions Leptin can promote the proliferations of VECs and VSMCs, which shows a dose- and time-dependent manner within a certain range. Atorvastatin inhibits leptin-induced proliferation effects of VECs and VSMCs.