高糖和MBL对体外培养的人肾小球内皮细胞表达IL-18的影响

目的观察高糖和甘露聚糖结合凝集素（MBL）对体外培养的人肾小球内皮细胞（HRGEC）表达白细胞介素-18（IL-18）的影响。方法体外培养正常HRGEC,按干预条件随机分为对照组（5mmol/L D-葡萄糖）、高糖组（30mmol/L D-葡萄糖）和甘露醇组（5mmol/L D-葡萄糖＋25mmol/L甘露醇）,培养72小时后分别采用实时荧光定量RCR法（Real-time PCR）和ELISA法检测各组细胞IL-18的mRNA及细胞培养上清液中的蛋白表达。另取细胞随机分为单纯高糖组和高糖＋MBL组,均用高糖培养72h后,分别加入等量的30%MBL缺乏人血清,高糖＋MBL组另加10μg/ml MBL,继续培养4h。采用免疫荧光法检测各组细胞表面凝集素补体途径（LCP）激活终产物膜攻击复合物（MAC）的沉积,并检测各组细胞IL-18的mRNA和蛋白表达。结果与其他各组相比,高糖组细胞的IL-18mRNA以及蛋白表达均明显增加（P〈0.05）;高糖＋MBL组细胞表面有明显MAC沉积,IL-18mRNA及蛋白表达与单纯高糖组相比明显增加（P〈0.05）。结论高糖可以诱导人肾小球内皮细胞炎症因子过表达;高糖和MBL共存时可促成LCP激活,并对DN的炎症状态具有促进作用。

Influence of high glucose and mannose binding lectin on interleukin-18 expression in cultured human renal glomerular endothelial cells

Objective To investigate the effects of high glucose and mannose binding lectin(MBL) on expression of interleukin-18 of cultured human renal glomerular endothelial cells(HRGEC).Methods Normal HRGEC cultured in vitro were divided randomly into three groups according to different intervention methods: normal glucose group(5 mmol/L D-glucose),manicol group(5mmol/L D-glucose+25mmol/L manicol)and high glucose group(30mmol/L D-glucose).Real-time PCR and Euzyme-linked Immunosorbent Assay(ELISA) were used to detect IL-18 mRNA expression of HRGEC and protein concentration in supernatant fluid after 72 hours’ culture.HRGEC was then randomly divided into two groups: single high glucose group and high glucose+MBL group.After 72 hours’ culture with 30mmol/L D-glucose,equivalent volume of 30% MBL deficiency human serum was added into two groups,10μg/mL MBL was only added into high glucose+MBL group,continue the culture for 4 hours.Immunofluorescence was used to detect Menbrane Attacks Complex(MAC)deposition on cell surface,which is the product of Mannose Binding Lectin Complement Pathway(LCP).Real-time PCR and ELISA were used to detect mRNA and protein expression of IL-18 in each group.Results Compared with other groups,the mRNA and protein expression of IL-18 in high glucose group was apparently increased(P<0.05).MAC was confirmed obviously deposition on cell surface in high glucose+MBL group.Both mRNA and protein expression of IL-18 in high glucose + MBL group were significantly higher(P<0.05) compared with single high glucose group.Conclusion High glucose can bring inflammatory factors’ overexpression of HRGEC;high glucose together with MBL can bring LCP activation and promote inflammatory status of diabetic nephropathy.