过敏性紫癜患儿血清对人脐静脉内皮细胞的影响及甲基泼尼松龙的保护作用

Impact of sera from children with active HenochSchnlein purpura on human umbilical venous endothelial cells (HUVECs) and protective effects of methylprednisolone against HUVECs injury

摘要:

目的：探讨过敏性紫癜（HSP）患儿血清对人脐静脉内皮细胞（HUVEC）炎症损伤的作用及甲基泼尼松龙（甲泼尼龙）在其中的调节作用。方法：人脐静脉内皮细胞培养分为4组：空白对照组、正常血清组、HSP血清组及甲泼尼龙干预组。分别检测各组上清液白细胞介素-8（IL-8）、肿瘤坏死因子-α（TNF-α）和一氧化氮（NO）水平；半定量RT-PCR和Western blot法检测HUVEC中核因子-κB（NF-κB）和不规则趋化因子(Fractalkine) mRNA及蛋白表达。 结果：HSP血清组上清液IL-8、TNF-α和NO水平均高于空白对照组和正常血清组（P<0.05）， 甲泼尼龙干预组上清液IL-8、TNF-α和NO水平较HSP血清组明显降低（P<0.05）；HSP血清组HUVEC中NF-κB和Fractalkine mRNA表达量明显高于空白对照组（P<0.05），两者的蛋白表达量明显高于空白对照组和正常血清组（P<0.05）；甲泼尼龙干预组HUVEC中NF-κB和Fractalkine mRNA及蛋白表达量较HSP血清组明显降低（P<0.05）。结论：HSP急性期患儿血清可以诱导体外培养的HUVEC活化并分泌炎性细胞因子，甲泼尼龙可能通过抑制NF-κB表达、减少炎性因子产生而减轻血管炎症。

Abstract:

OBJECTIVE: To observe the changes of human umbilical venous endothelial cells (HUVECs) induced by the sera from children with active Henoch-Sch-nlein purpura (HSP) and the protective effects of methylprednisolone against HUVECs injury. METHODS: HUVECs were divided into four groups based on the culture conditions: blank control group, normal serum group, HSP serum group, and HSP serum plus methylprednisolone group. The levels of tumor necrosis factor (TNF)-α and interleukin (IL)-8 in the supernatants of each group were detected using ELISA and the nitric oxide (NO) level by nitrate reductase determination. Moreover, the expressions of nuclear factor-kappa B (NF-κB) and Fractalkine in HUVECs were examined by semiquantitative reverse transcription polymerase chain reaction (RT-PCR) and Western blot, respectively. RESULTS: The levels of IL-8, TNF-α, and NO in the HSP serum group were significantly higher than those in the blank control and normal serum groups (P<0.05). Compared with the HSP serum group, the levels of IL-8, TNF-α, and NO in the HSP serum plus methylprednisolone group decreased significantly (P<0.05). The mRNA expression levels of NF-κB and Fractalkine in the HSP serum group were significantly higher than those in the blank control group (P<0.05). The protein expression levels of NF-κB and Fractalkine in the HSP serum group were significantly higher than those in the blank control and normal control group (P<0.05). Compared with the HSP serum group, the mRNA and protein expression levels of NF-κB and Fractalkine in the HSP serum plus methylprednisolone group decreased significantly (P<0.05). CONCLUSIONS: The sera from children with active HSP can induce the in vitro cultured HUVECs to become activated and excrete cytokines. Methylprednisolone may inhibit NF-κB expression, reduce the production of inflammatory factors, and thus alleviate vascular inflamation.