免疫活化血小板对人脐静脉内皮细胞COX-2及PPAR-α表达与活性的影响及LDL的作用

Effects of immune-activated platelets and LDL on expression and activity of COX-2 and PPAR-α in HUVECs

目的:观察免疫活化血小板对人脐静脉内皮细胞(HUVECs)环氧合酶2(COX-2)及过氧化物酶体增殖物活化受体α(PPAR-α)表达及活性的影响,并探讨低密度脂蛋白(LDL)对其作用。方法:利用腺苷二磷酸(ADP)刺激血小板免疫活化,然后利用免疫活化血小板与HUVECs加或不加LDL共孵育,利用实时定量RT-PCR、Westernblotting分别检测HUVECsCOX-2及PPAR-αmRNA及其蛋白的表达,ELISA法检测PGE2浓度反映COX-2活性、核因子活性检测试剂盒检测PPAR-α活性。结果:血小板活化组COX-2及PPAR-αmRNA表达量较血小板对照组均有较明显升高(1.49±0.27vs0.68±0.21,1.45±0.21vs1.17±0.16,均P<0.01);血小板活化组COX-2及PPAR-α蛋白表达量较血小板对照组亦有较明显升高(1.600±0.145vs0.700±0.073,1.630±0.143vs0.960±0.073,均P<0.01),血小板活化组孵育液中PGE2浓度较血小板对照组明显升高[(42.46±5.57)ng/Lvs(23.73±2.53)ng/L,P<0.01],但血小板活化组PPAR-α活性较血小板对照组无明显变化;LDL本身无明显增加HUVECsCOX-2及PPAR-α的表达及活性,但其能增加免疫活化血小板的这一作用。结论:免疫活化血小板显著促进HUVECsCOX-2表达及活性增加,亦能显著促进HUVECsPPAR-α表达,但并不改变PPAR-α活性;一般浓度的LDL并不对HUVECsCOX-2、PPAR-α的表达及活性产生影响,但能促进免疫活化血小板对内皮细胞的这一作用。

AIM ： To observe the effect of immune - activated platelets and low - density lipoprotein cholesterol （LDL） on the expression and activity of cyclooxygenase -2 （COX -2） and peroxisome proliferator activated receptor α （PPAR -α） in human umbilical vein endothelial cells （HUVECs） treated with activated platelets and LDL. METHODS： The platelets were activated by ADP. The co - culture system of HUVECs with immune activated platelets and/or LDL were established. The activity of COX -2 and expression of PPAR -α at mRNA and protein levels in HUVECs were detected by RT - PCR and Western blotting. The concentration of PGE2 was measured by ELISA for representing the COX - 2 activity. The PPAR - α activity was determined by a nuclear factor assay kit. RESULTS ： The COX - 2 activity and mRNA expression of PPAR - α, the protein levels of COX - 2 and PPAR - α and PGE2 concentration in activated platelets group were significant higher than those in un - activated platelets group （all P 〈 0.01 ）. No difference of PPAR - α binding activity was observed between two groups. LDL didn＇t affect the COX -2 activity and PPAR-α expression, but significantly promoted the stimulating effect of immune - activated platelets. CONCLUSION： Immune - activated platelets significantly promote COX -2 activity and PPAR -α expression in HUVECs, but don＇t change the PPAR -α binding activity. LDL at general concentration does not affect the expression and activity of COX - 2 and PPAR - α, but promote the effect of activated platelets on HUVECs.