CRH促进大鼠腹腔巨噬细胞IL-6分泌及其受体途径  

Corticotrophin-releasing Hormone Promotes Interleukin-6 Release from Rat Peritoneal Macrophages and Its Possible Receptor Pathways

作　　者：王伍超[1] 陈金萍[1] 张岫竹[1] 王旭辉[1] 刘大维[1] 周继红[1] 

机构地区：[1]第三军医大学大坪医院野战外科研究所第四研究室,创伤、烧伤与复合伤国家重点实验室,全军/重庆市交通医学研究所,重庆400042

出　　处：《中国神经免疫学和神经病学杂志》2009年第4期276-280,共5页Chinese Journal of Neuroimmunology and Neurology

基　　金：国家重点基础研究发展规划项目(2005CB522602)

摘　　要：目的探讨促肾上腺皮质激素释放激素(CRH)对大鼠腹腔巨噬细胞白细胞介素-6(IL-6)分泌的影响及其受体途径。方法将原代培养的大鼠腹腔巨噬细胞分为对照组、CRH组、CRH受体(CRHR)1选择性阻断剂CP-154526+CRH组(简称CP+CRH组)、CRHR1/2非选择性阻断剂α-helical CRH9-41+CRH组(简称HL+CRH组)以及蛋白激酶A(PKA)抑制剂H89+CRH组(简称H89+CRH组)5组。对照组仅加入新鲜RPMI1640培养液2 mL培养;CRH处理组加入1×10^(-9) mol/L CRH培养液2 mL;后3组分别加入含CP-154526(5×10^(-4) mol/L)、α-helical CRH9-41(1×10^(-6) mol/L)以及H89(1×10^(-6) mol/L)培养基预孵育2 h,然后加入CRH使培养液CRH终浓度为1×10^(-9) mol/L。各组分别于CRH刺激后0.5、1、2、4、8 h收集培养皿中上清液,应用ELISA法检测IL-6水平。结果对照组巨噬细胞分泌IL-6维持在低水平。CRH组巨噬细胞分泌IL-6在2 h时增加,达(7.7±0.741)μg/g prot,高于同一时点的对照组(P<0.05),4 h时达高峰[(16.60±3.94)μg/gprot],接近同时相点对照组的3倍(P<0.01)。CP+CRH组巨噬细胞IL-6分泌量在2、4 h时下降非常显著,与同时点CRH组比较分别下降约88%、92%(P<0.01),低于同时点对照组(P<0.01)。H89+CRH组中巨噬细胞分泌IL-6在2、4 h显著下降(P<0.01),其抑制效应约66%～79%。结论 CRH可通过CRHR1激活巨噬细胞促进IL6分泌,与细胞内PKA信号密切相关。CRH在激活巨噬细胞的过程中可能起到允许和易化作用。Objective To investigate the production of interleukin-6 （IL-6） from macrophages in presence to corticotrophin-releasing hormone （CRH） and to further substantiate the implication of CRH receptor （CRHR） styles in the modulation of cytokines. Methods Rat peritoneal macrophages cultured in vitro were stimulated with CRH at a concentration of 10.9 mol/L or without any stimulation as healthy control group. When inhibitors were used, cells were pretreated with CRHR1 antagonist CP154526 at 5 × 10^-4 mol/L, CRHR1/2 nonselective antagonist a-helical CRH9-41 at 1× 10^-6 mol/L, protien kinase A （PKA） inhibitor H89 compound at 1 × 10^-6 mol/L for 2 h before stimulation with CRH. The concentrations of IL-6 for incubation 0.5, 1, 2, 4 and 8 h following stimulation were investigated by enzyme linked immunosorbent assay （ELISA） method. Results In the healthy control group, the average concentration of IL-6 kept at low level. The secretion of IL-6 from macrophages exposed to CRH for 2 h reached to （7.7±0. 741） μg/g prot, increased remarkably compared with that in CRH group at 1 h （P〈0.01）, it is two folds of that in the normal control group （P〈0.01）. Then the content of IL-6 reached its peak, about （16.60±3.94） μg/g prot, it is three times of that in healthy control group （P〈0.01）. The effects of CRH peptides on the production of IL-6 from macrophages especially during the crest time were prevented likewise by blocking CRHR1 by CP154526 or blocking CRHR1/2 by s-helical CRH9- 41. Pretreatment of macrophages with AC inhibitors H89 induced a significant inhibition of the expression for IL- 6 eytokines compared with CRH group at 2 or 4 h. Furthermore, when CRHR were blockaded or AC was inhibited in advance, the content of IL-6 secreted from macrophages exposed to CRH was lower than that in control group. Conclusions Our data suggest that CRH arguments proinflammatory cytokine IL-6 production from macrophages in vitro through CRHR1, involving in the intracellular PKA signal pathway.

关 键 词：促肾上腺皮质激素释放激素 促肾上腺皮质激素释放激素受体 蛋白激酶A 巨噬细胞 白细胞介素-6 

分 类 号：R392.6[医药卫生—免疫学]