高糖对人脐静脉内皮细胞CD26的表达及MBL补体激活途径的影响及作用机制

发布时间：2018-05-14 00:15

本文选题：CD26 + 血管内皮细胞　；参考：《遵义医学院》2015年硕士论文

【摘要】：目的:观察高糖培养下人脐静脉内皮细胞(HUVEC)的CD26表达变化、甘露聚糖结合凝集素(MBL)补体途径的激活及炎症因子(IL-6和NF-k B)的表达情况,探讨CD26在糖尿病肾病(DN)慢性炎症发病机制中的作用,为CD26抑制剂治疗DN寻求基础研究依据。方法:体外培养人脐静脉内皮细胞,分别设置正常对照组(含葡萄糖5.5mmol/L),甘露醇组(含葡萄糖5.5mmol/L+甘露醇24.5mmol/L),高糖组(含葡萄糖30mmol/L),采用免疫组织化学法检测细胞表面CD26、MBL、C3和膜攻击复合物(MAC)的表达,采用Western Blot检测细胞的CD26、MBL、C3及MAC的蛋白表达,采用实时荧光定量PCR(real-time PCR)检测细胞中CD26、IL-6和NF-k B m RNA的表达情况。结果:⑴免疫组化提示:在HUVEC中CD26、MBL、C3、膜攻击复合物(MAC)主要表达于胞膜和胞浆。高糖组CD26、MBL、C3及MAC的蛋白表达均明显高于正常组与甘露醇组(P0.05);在正常组与甘露醇组之间,各指标蛋白表达量均无统计学意义(P0.05)。CD26与MBL、C3、MAC蛋白表达量均呈明显正相关(r=0.874,r=0.952,r=0.947,P0.01)。⑵Western Blot提示:高糖组CD26、MBL、C3及MAC的蛋白表达亦明显高于正常组及甘露醇组(P0.05);在正常组与甘露醇组之间,各指标蛋白表达量均无统计学意义(P0.05)。CD26与MBL、C3、MAC蛋白表达量呈明显正相关(r=0.825,r=0.869,r=0.828,P0.01)。⑶RT-PCR提示:高糖组CD26、IL-6、NF-k B的m RNA表达均高于正常组及甘露醇组(P0.05);在正常组与甘露醇组之间,各指标m RNA表达量均无统计学意义(P0.05)。CD26与IL-6、NF-k B的m RNA表达量均呈正相关(r=0.858,r=0.794,P0.05)。结论:体外培养的人脐静脉内皮细胞可少量表达CD26。高糖条件下,HUVEC的CD26表达增高,后者可能通过激活MBL介导的甘露糖结合凝集素补体途径,引起下游细胞因子IL-6、NF-k B表达增加,加重高糖作用下HUVEC的炎症反应。
[Abstract]:Aim: to observe the changes of CD26 expression in human umbilical vein endothelial cells (HUVECs), the activation of mannan-binding lectin (MBL) complement pathway and the expression of inflammatory cytokines (IL-6 and NF-k B) in cultured human umbilical vein endothelial cells (HUVECs). To explore the role of CD26 in the pathogenesis of chronic inflammation in diabetic nephropathy (DN), and to seek the basis for the treatment of DN with CD26 inhibitor. Methods: human umbilical vein endothelial cells were cultured in vitro. The expression of CD26MMBL3 and membrane attack complex (MAC-1) was detected by immunohistochemistry in normal control group (including glucose 5.5 mmol 路L ~ (-1), mannitol group (containing glucose 5.5mmol/L _ (2.5) mmol 路L ~ (-1) and high glucose group (containing glucose 30 mmol / L ~ (-1). The expression of CD26ML-C _ 3 and MAC was detected by Western Blot, and the expression of IL-6 and NF-k B m RNA was detected by real-time fluorescence quantitative PCR(real-time. Results Immunohistochemistry showed that CD26ML-C3, a membrane attack complex, was mainly expressed in the membrane and cytoplasm of HUVEC. The protein expressions of CD26ML-C _ 3 and MAC in high glucose group were significantly higher than those in normal group and mannitol group (P 0.05), and between normal group and mannitol group. There was no significant difference in the protein expression of P0.05G, CD26 and MBLL-C3C3MAC protein expression. The results showed that there was a significant positive correlation between the protein expression of P0.05, CD26 and MBC, and the expression of CD26ML-C3 and MAC in high glucose group was significantly higher than that in normal group and mannitol group, and between normal group and mannitol group, the protein expression of CD26ML-C _ 3 and MAC in mannitol group was significantly higher than that in normal group, while in mannitol group, the expression of CD26ML-C _ 3 and MAC in high glucose group was significantly higher than that in normal group and mannitol group. There was no statistical significance between the expression of P0.05U. CD26 and the MAC protein expression of MBLML-C3C3. The results of RT-PCR indicated that the expression of m RNA in high glucose group was higher than that in normal group and mannitol group, and between normal group and mannitol group, the expression of m RNA of NF-kB in high glucose group was higher than that in normal group and mannitol group, and in mannitol group, the expression of m RNA in high glucose group was significantly higher than that in normal group and mannitol group. There was no statistical significance between the expression of m RNA and the expression of m RNA in IL-6 nF-kB. There was a positive correlation between the expression of m RNA and the expression of m RNA in IL-6 nF-kB. Conclusion: in vitro cultured human umbilical vein endothelial cells can express a small amount of CD 26. The expression of CD26 in HUVEC was increased under high glucose condition. The latter may increase the expression of IL-6NF-K B by activating the MBL mediated mannose binding lectin complement pathway and aggravate the inflammatory response of HUVEC under high glucose.
【学位授予单位】：遵义医学院
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R587.2;R692.9

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