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前列腺素E₂不同受体对足月妊娠子宫肌细胞分泌炎性细胞因子的不同调节作用

张友义¹, 刘伟娜^1,2, 游兴姬¹, 古航³, 徐晨^1,4, 倪鑫^1,5,*

¹海军军医大学(第二军医大学)生理学教研室，上海 200433；²中国人民解放军第413医院妇科，舟山 316000；³上海市长海医院妇产科，上海 200433；⁴复旦大学基础医学院生理学与病理生理学系，上海 200032；⁵中南大学湘雅医院分子代谢组学研究中心，长沙 410008

摘要

前列腺素E2 (prostaglandin E2, PGE2)在妊娠维持和分娩启动中起着关键作用。愈来愈多的研究表明，人类妊娠的维持和分娩启动是一个炎症过程。本研究用原代培养的足月妊娠人子宫平滑肌细胞(human uterine smooth muscle cells, HUSMCs)作为研究对象，观察PGE2四种受体亚型(EP1、EP2、EP3和EP4)对妊娠HUSMCs分泌炎性细胞因子的影响。用药物处理和/或转染各受体siRNA后，使用ELISA试剂盒检测HUSMCs培养液中炎性分泌因子的浓度。结果显示，PGE2可剂量依赖性促进白细胞介素6 (interleukin 6, IL-6)和肿瘤坏死因子α (tumor necrosis factor α, TNFα)分泌，而抑制趋化因子(c-x-c基序)配体8 (CXCL8)释放，但对IL-1β和趋化因子(c-c基序)配体2 (CCL-2)的分泌没有影响。EP1/EP3激动剂17-苯基-trinor-PGE2刺激IL-6和TNFα的分泌，抑制IL-1β和CXCL8的分泌。转染EP3 siRNA能逆转17-苯基-trinor-PGE2对IL-6和TNFα分泌的影响，但是17-苯基-trinor-PGE2对IL-1β和CXCL8的作用仍然存在。敲低EP1可阻断17-苯基-trinor-PGE2对IL-1β分泌的抑制效应，而17-苯基-trinor-PGE2促进IL-6和TNFα分泌的作用仍然存在。EP2和EP4激动剂均可刺激IL-1β和TNFα分泌，这种效应可分别被EP2 siRNA和EP4 siRNA所逆转。磷脂酶C (phospholipase C, PLC)和蛋白激酶C (protein kinase C, PKC)的抑制剂可阻断17-苯基-trinor-PGE2对TNFα和CXCL8分泌的影响。PI3K抑制剂LY294002和P38抑制剂SB202190可分别阻断17-苯基-trinor-PGE2诱导的IL-1β和IL-6分泌。腺苷酸环化酶和PKA的抑制剂均能逆转EP2和EP4激动剂促进IL-1β和TNFα分泌的作用，而PLC和PKC抑制剂则可阻断EP2和EP4诱导的TNFα分泌，但不能阻断IL-1β的分泌。以上结果表明，PGE2不同受体对子宫肌细胞分泌炎性细胞因子具有不同作用，提示PGE2可能通过不同受体的精细调节来调控妊娠期子宫肌层中的炎性微环境。

关键词： 子宫肌层; 前列腺素E2; 促炎细胞因子; 妊娠

分类号：Q25

Prostaglandin E₂ receptors differentially regulate the output of proinflammatory cytokines in myometrial cells from term pregnant women

ZHANG You-Yi¹, LIU Wei-Na^1,2, YOU Xing-Ji¹, GU Hang³, XU Chen^1,4, NI Xin^1,5,*

¹Department of Physiology, Navy Military Medical University (Second Military Medical University), Shanghai 200433, China；²Department of Gynecology, Chinese PLA 413th Hospital, Zhoushan 316000, China；³Department of Obstetrics and Gynecology, Changhai Hospital, Shanghai 200433, China；⁴Department of Physiology and Pathophysiology, School of Basic Medical Sciences, Fudan University, Shanghai 200032, China；⁵Research Center of Molecular Metabolomics, Xiangya Hospital, Central Southern University, Changsha 410008, China

Abstract

Prostaglandin (PG) E2 plays critical roles during pregnancy and parturition. Emerging evidence indicates that human labour is an inflammatory event. We sought to investigate the effect of PGE2 on the output of proinflammatory cytokines in cultured human uterine smooth muscle cells (HUSMCs) from term pregnant women and elucidate the role of subtypes of PGE2 receptors (EP1, EP2, EP3 and EP4). After drug treatment and/or transfection of each receptor siRNA, the concentrations of inflammatory secreting factors in HUSMCs culture medium were detected by the corresponding ELISA kits. The results showed that, PGE2 increased interleukin 6 (IL-6) and tumor necrosis factor alpha (TNFα) output, decreased chemokine (c-x-c motif) ligand 8 (CXCL8) output in a dose-dependent manner, but had no effect on IL-1β and chemokine (c-c motif) ligand 2 (CCL-2) secretion of HUSMCs. EP1/EP3 agonist 17-phenyl-trinor-PGE2 stimulated IL-6 and TNFα whilst suppressing IL-1β and CXCL8 output. The effects of 17-phenyl-trinor-PGE2 on IL-1β and CXCL8 secretion were remained whereas its effect on IL-6 and TNFα output did not occur in the cells with EP3 knockdown. The stimulatory effects of 17-phenyl-trinor-PGE2 on IL-6 and TNFα were remained whereas the inhibitory effects of 17-phenyl-trinor-PGE2 on IL-1β secretion was blocked in the cells with EP1 knockdown. Either of EP2 and EP4 agonists stimulated IL-1β and TNFα output, which was reversed by EP2 and EP4 siRNA, respectively. The inhibitors of phospholipase C (PLC) and protein kinase C (PKC) blocked EP1/EP3 modulation of TNFα and CXCL8 output. PI3K inhibitor LY294002 and P38 inhibitor SB202190 blocked 17-phenyl-trinor-PGE2-induced IL-1β and IL-6 output, respectively. The inhibitors of adenylyl cyclase and PKA prevented EP2 and EP4 stimulation of IL-1β and TNFα output, whereas PLC and PKC inhibitors blocked EP2- and EP4-induced TNFα output but not IL-1β output. Our data suggest that PGE2 receptors exhibit different effects on the output of various cytokines in myometrium, which can subtly modulate the inflammatory microenvironment in myometrium during pregnancy.  

Key words: myometrium; prostaglandin E2; proinflammatory cytokines; pregnancy

收稿日期：2018-11-22　　录用日期：2019-01-22

通讯作者：倪鑫　　E-mail: nixin@smmu.edu.cn

DOI: 10.13294/j.aps.2019.0028

引用本文：

张友义, 刘伟娜, 游兴姬, 古航, 徐晨, 倪鑫. 前列腺素E₂不同受体对足月妊娠子宫肌细胞分泌炎性细胞因子的不同调节作用[J]. 生理学报 2019; 71 (2): 248-260.

ZHANG You-Yi, LIU Wei-Na, YOU Xing-Ji, GU Hang, XU Chen, NI Xin. Prostaglandin E₂ receptors differentially regulate the output of proinflammatory cytokines in myometrial cells from term pregnant women. Acta Physiol Sin 2019; 71 (2): 248-260