内源性硫化氢通过抑制炎症反应调控胎膜中前列腺素生成的机制及其与分娩启动的关系

Prostaglandin (PG) of intrauterine tissues has been identified as one of the final events leading to parturition. The local level of PG were determined by the balance between cyclooxygenase-2 (COX-2) and metabolic enzymes (15-hydroxyprostaglandin dehydrogenase,PGDH). The regulation of COX-2 and PGDH remain, however, largely unknown.Hydrogen sulfide (H2S) is endogenously produced from L-cysteine by cystathionineβ-synthase (CBS) and cystathionineγ-lyase (CSE).We found that CBS and CSE express in human chorion and amnion by IHC and western blotting. And the production of endogenous H2S in fetal membrane was measured by miniaturized H2S micro-resporation sensor. The results showed that the mRNA and protein expression of CBS and CSE and the production of H2S were significantly decreased in the fetal membrane during labour. Moreover, H2S promoted the expression of PGDH, while inhibited the expression of COX-2. Furhermore, COX-2 level correlated negatively with CBS while PGDH level correlated positively with CBS and CSE in term labour (TL) chorion but term non-labour (TNL) tissues. It suggested that H2S play a pivotal role in the regulation of PG production in chorion. As we known, cytokines (IL-8, IL-1β, TNF-α,et al) were involved in pregnancy and preterm labour. We found that kockdown of inflammatory cytokines in chorionic cells could block the above effect of H2S on PG production. So the results suggested that the effects of H2S on the regulation of COX-2 and PGDH expression in fetal membrane, possible through inhibiting cytokines production. Bioinformatics analysis showed that miR-26a,miR-26b,miR-101,miR-143, miR-199s and miR-101 may regualte the expression of COX-2 and PGDH. We found that miR-26a and miR-26b were significantly decreased, while miR-101 and miR-199a increased,in term labour fetal membrane compared to those of term non-labour samples. Meanwhile ours results showed that miRNA involved in the regulation of COX-2 and PGDH expression in human fetal membrane.On the basis of preliminary experiments, we will employ a set of molecular biological techniques to investigate the regulation of synthesis and metabolism of PG by H2S in human fetal membrane.The study will help to furtherly clarity the mechanism of human parturition and prevent and treatment of preterm birth.

前列腺素(prostaglandin,PG)是子宫肌收缩的主要内源性因子，其局部水平取决于合成酶COX-2和代谢酶PGDH的表达和活性，但调节这些关键酶的机制尚未阐明。本课题利用高表达COX-2和PGDH的胎膜，以硫化氢(H2S)抗炎作用为切入点，探讨H2S在PG生成中的作用。前期工作发现临产者胎膜H2S合成酶CBS和CSE表达、H2S生成明显减少，并与COX-2表达负相关、与PGDH表达正相关；敲低细胞炎性细胞因子的表达可阻断H2S促进PGDH、抑制COX-2表达的作用，提示H2S通过抑制胎膜炎症反应减少局部PG的生成。但关于H2S抑制胎膜炎症反应及细胞因子在H2S抑制胎膜PG生成中的机制尚不清楚；另发现miRNA参与COX-2和PGDH表达的调节，可能介导细胞因子调节胎膜PG生成的作用。本课题通过系统研究H2S抗炎功能在胎膜PG生成中的作用，为阐明人类分娩启动机制和防治早产提供新的思路

早产在人类发生率一直居高不下，是围产期胎儿发病率和死亡率的首要因素，究其原因是人类分娩机制尚未阐明。而硫化氢（Hydrogen Sulfide, H2S）在人组起器官广泛分布，前期研究证实宫内组织亦有H2S合成酶的表达，但作用尚不清楚。利用人绒毛膜组织检测发现H2S合成酶表达和H2S生成速率在临产后绒毛膜膜显著降低，且与COX-2表达呈负相关、与PGDH表达呈正相关；检测发现miR-26 a、miR-26b、miR-199a、miR-101可参与COX-2和PGDH表达的调节过程；H2S促进miR26a、miR-101和miR-199a的表达，而抑制miR26b的表达。. 检测羊膜组织PGDH、COX-2和H2S合成酶（CBS、CSE、3-MST）的表达及H2S的生成，发现临产羊膜组织COX-2蛋白表达量高于未临产羊膜组织，而CBS和CSE蛋白的表达及H2S生成与之相反，提示局部H2S合成酶CBS、CSE与羊膜COX-2的表达及前列腺素的生成密切相关，并且在体外培养的羊膜细胞上发现H2S下调人羊膜COX-2的表达和酶活性。. 检测子宫肌中H2S合成酶CBS、CSE，核转录因子κB（NF-κB），子宫收缩相关蛋白CAPs(CX43、OTR、PGFR)的表达，发现H2S合成酶与子宫收缩相关蛋白CX43、OTR、PGFR的表达密切相关；探究内源性H2S对人子宫平滑肌细胞子宫收缩相关蛋白CAPs(CX43、OTR、PGFR)表达，促炎性细胞因子（IL-1β、IL-6和TNFα）生成的影响，结果提示妊娠子宫肌所产生的内源性H2S可能通过激活KATP通道而抑制NF-κB的激活，进而抑制炎性细胞因子的生成，继而抑制子宫激活，维持子宫静息。