【摘要】 目的 观察嘌呤能受体P2X配体门控离子通道7的配体(P2X7R)过表达白血病细胞诱导分化的巨噬细胞单钠尿酸盐(MSU)晶体诱导痛风炎症反应过程中NOD样受体家族3(NLRP3)蛋白、IL-1β、TNF-α表达情况。方法 取人单核细胞白血病细胞系THP-1，并随机分为过表达组、空白组、模型组、对照组；过表达组和空白组分别转染P2X7R过表达质粒、空白载体质粒，转染5 d，将过表达组、空白组、模型组THP-1细胞用100 ng/mL的PMA刺激3 h后分化为巨噬细胞，另将MSU晶体用氢氧化钠溶解配制成浓度为100μg/mL的MSU乳糜状悬液加入培养液中孵育6 h；对照组正常培养。分别采用RT-PCR法和Western blot法测算巨噬细胞P2X7R mRNA、蛋白，ELISA法检测巨噬细胞上清液IL-1β、TNF-α,Western blot法测算巨噬细胞NOD样受体家族3(NLRP3)蛋白。结果 与对照组比较，过表达组、空白组、模型组P2X7R mRNA和蛋白相对表达量升高，细胞上清液IL-1β、TNF-α水平升高，细胞NLRP3蛋白相对表达量升高(P均<0.05)；与模型组、空白组比较，过表达组P2X7R mRNA、蛋白相对表达量升高，细胞上清液IL-1β、TNF-α水平升高，细胞NLRP3蛋白相对表达量升高(P均<0.05)。结论 P2X7R过表达白血病细胞诱导分化的巨噬细胞MSU晶体诱导痛风炎症反应过程中IL-1β、TNF-α、NLRP3表达增加，IL-1β、TNF-α水平升高可能通过激活NLRP3蛋白来实现。更多还原

【Abstract】 Objective To observe the expression of NOD-like receptor family 3(NLRP3) protein,interleukin-1β(IL-1β) and tumor necrosis factor-α(TNF-α) in the process of gout inflammation induced by macrophage monosodium urate(MSU) crystals in macrophages differentiated from leukemia cells overexpressing purinergic receptor P2X ligand-gated ion channel 7(P2X7R).Methods THP-1 cell line were taken and randomly divided into the overexpression group,empty vector group,model group and control group,respectively.Cells in the overexpression group and empty vector group were transfected with P2X7R overexpression plasmid and empty vector plasmid for five days,respectively.THP-1cells in the vector group,empty vector group and model group were stimulated with 100 ng/mL PMA for three hours and then were differentiated into macrophages.In addition,MSU crystals were dissolved with sodium hydroxide to prepare MSU chyle suspension at a concentration of 100 μg/mL and were incubated in the culture medium for six hours.Cells in the control group were cultured normally.The mRNA and protein of P2X7R in macrophages were measured by RT-PCR and Western blotting,respectively.IL-1β and TNF-α in the supernatant of macrophages were detected by ELISA,and the NLRP3 protein in macrophages was measured by Western blotting.Results Compared with the control group,the relative expression levels of P2X7R mRNA and protein,IL-1β and TNF-α in cell supernatant and NLRP3 protein all increased in the overexpression group,empty vector group and model group(all P<0.05).Compared with the model group and empty vector group,the relative expression levels of P2X7R mRNA and protein,IL-1β and TNF-α in cell supernatant and NLRP3 protein all increased in the overexpression group(all P<0.05).Conclusion The expression of IL-1β,TNF-α and NLRP3 increases and the levels of IL-1β and TNF-α levels increase in the process of gout inflammation induced by MSU crystals of macrophages differentiated from leukemia cells with P2X7R overexpression,which may be achieved by activating NLRP3 protein.更多还原