MAPK信号通路在瘦素诱导小鼠巨噬细胞IL-1α表达中的作用

Leptin-induced IL-1α expression by peritoneal macrophages of murine:involvement of MAPK pathways

通过体外培养小鼠腹腔巨噬细胞(Peritoneal macrophages,PM),按瘦素(leptin)不同质量浓度和/或丝裂原活化蛋白激酶(Mitogen-activated protein kinases,MAPK)特异性抑制剂PD98059、U0126、SB203580、SP600125进行分组,分别收集培养细胞和上清液,用酶联免疫吸附试验(Enzyme-linked immunosorbent assay,ELISA)测定了上清液中的白介素1α(Interleukin-1α,IL-1α)水平,用免疫印迹(Western blot)方法测定细胞内磷酸化细胞外调节蛋白激酶(phosphorylated extracellular signal-regulated kinase,p-ERK)1/2、p-p38以及磷酸化Jun氨基末端激酶(phosphorylated Jun N-terminal kinases,p-JNK)的表达水平,用逆转录-聚合酶链式反应(Reverse transcription polymerase chain reaction,RT-PCR)测定IL-1α mRNA的表达.结果发现,瘦素能够剂量依赖性地诱导小鼠PM产生IL-1α,在瘦素质量浓度为75 ng/mL时,IL-1α表达至峰值,且为对照组的12.2倍.瘦素可以活化ERK1/2和p38 MAPK信号转导通路,瘦素处理组的p-ERK1、p-ERK2、p-p38表达水平分别是对照组的2.3、2.4和2.6倍.ERK1/2的特异性抑制剂PD98059、U0126和p38的特异抑制剂SB203580能够分别抑制瘦素引起的ERK1/2、p38蛋白磷酸化以及IL-1α mRNA增加,两者的联合作用可以强烈抑制IL-1α mRNA的表达.瘦素在小鼠PM中通过同时活化ERK1/2、p38 MAPK信号转导通路而诱导细胞产生IL-1α.

The effect of leptin on inducing interleukin 1α（IL-1α） secretion in murine peritoneal macrophages（PM） and the involvement of mitogen activated protein kinases（MAPK） pathways in this process were investigated.The murine PM cultured in vitro were divided into groups according to the concentrations of leptin with or without the addition of specific inhibitors of MAPK.The level of IL-1α in the culture supernatant was measured by Enzyme-linked immunosorbent assay（ELISA） and Western blotting was used to determine the phosphorylated extracellular signal-regulated kinase（p-ERK）1/2,p38 and phosphorylated Jun N-terminal kinases（p-JNK）.Reverse transcription-polymerase chain reaction（RT-PCR） was performed to examine the expression of IL-1α mRNA in PM.The results indicated that leptin induced IL-1α expression in murine PM and that the secretion of IL-1α reached the maximum at 75 ng/mL of leptin.p-ERK1/2 and p38 played major roles in leptin induced IL-1α production.Expression of p-ERK1,p-ERK2 and p-p38 in leptin-treated group increased to 2.3,2.4 and 2.6-fold of the control group,respectively.The specific inhibitors of ERK1/2 and p38 could significantly suppress the activation of ERK1/2 and p38 induced by leptin in murine PM and partly inhibited the IL-1α mRNA expression.Combination of the inhibitors totally abolished the increasing of IL-1α mRNA.Leptin induced IL-1α production by simultaneously activating ERK1/2 and p38 signal pathway in murine PM.