血中可溶性白细胞分化抗原40配体对内皮细胞损伤的影响

Effect of Soluble Cluster of Differentiation 40 Ligand on the Injury in Human Umbilical Vein Endothelial Cells

目的研究血中可溶性白细胞分化抗原40配体(s CD40L)对人脐静脉内皮细胞(HUVEC)的损伤并探讨其发生机制。方法应用不同浓度人重组CD40L(r-CD40L)刺激HUVEC 24 h,建立单核细胞(THP-1)渗出模型,评价内皮细胞通透性。应用不同浓度r-CD40L刺激HUVEC 24 h,逆转录PCR技术检测HUVEC中TNF-αm RNA水平,观察不同浓度r-CD40L对内皮细胞功能的影响(以TNF-αm RNA水平表示)。采用ELISA法测定不同浓度r-CD40L刺激HUVEC后上清液中TNF-α蛋白水平。应用25μg/L r-CD40L刺激HUVEC不同时间,流式细胞技术检测HUVEC凋亡情况。结果随着r-CD40L刺激浓度的增加,单核细胞透膜数逐渐升高,HUVEC中TNF-αm RNA表达水平逐渐升高,HUVEC上清液中TNF-α的浓度逐渐增加。应用25μg/L r-CD40L刺激HUVEC不同时间,未诱导出细胞凋亡。结论 s CD40L对脐静脉内皮细胞通透性的影响呈浓度依赖性,其机制可能是通过上调内皮细胞核内TNF-αm RNA表达并促进其分泌TNF-α实现的。s CD40L对HUVEC功能的影响明显早于细胞的凋亡。

Objective To investigate the effect of soluble cluster of differentiation 40 ligand （sCD40L） on human umbilical vein endothelial cell （HUVEC） and explore its underlying mechanisms.Methods The THP-1 leakage model was established by stimulating HUVEC with different concentration of r-CD40L for 24 hours.The permeability of endothelial cell was evaluated.The level of tumor necrosis factor-α （TNF-α） mRNA in HUVEC cell was examined 24 hours after stimulation by different concentration of r-CD40L using RT-PCR technology.The concentration of TNF-α in supematant was measured by ELISA.The apoptosis of HUVEC after r-CD40L stimulation （25 μg/L） was detected by flow.Results The number of THP-1 has been shown to gradually increase with the concentration of r-CD40L.Along with the increase of r-CD40L concentration,TNF-α mRNA in HUVEC expression level increased gradually.TNF-α concentration of HUVEC in supernatant gradually increased with the increase of rCD40L concentration.No apoptosis was observed in HUVEC with 25 μg/L r-CD40L stimulation.Conclusion The permeability of endothelial cells was depended on the concentration of sCD40L,the mechanism may rely on the induced expression of TNF-α mRNA and increased the secretion of TNF-α.The effect of sCD40L on the function of HUVEC was observed much earlier than cell apoptosis.