蛋白酶切激活受体-2在输卵管运动及输卵管转运过程中的作用及机制研究

Protease activated receptor-2(PAR-2) belongs to a G-protein coupled receptor which can be activated by various of proteases, after activation it regulates the motility of smooth muscle, however, the role of PAR-2 in the motility and transport of fallopian tube remains unclear. According to our preliminary data and the literatures, we hypothesize that the PAR-2 is expressed in both human and rat fallopian tubes, the activation of PAR-2 stimulates the motility of tubal smooth muscle through two possible pathway: one is prostaglandin- prostaglandin receptor pathway, another is the PLC-IP3-PKC-L-type voltage gated Ca2+ channel pathway, PAR-2 takes part in the development of fallopian pregnancy by affecting the transport function of fallopian tubes. In this project, we will detect the expression of PAR-2 in human and rat fallopian tubes by the IHC,RT- PCR, and WB, investigate the role and mechanism of PAR-2 in regulating the motility of tubal smooth muscle by tension recording, patch clam and Ca2+ imaging, examine the effect of PAR-2 in the tubal transport by using animal in vivo experiment, detect the change of PAR-2 in expression and function in fallopian pregnancy oviducts, to identify the role of PAR-2 in the tubal movement and transport. This project will provide a new research target and experimental basis for the regulation of oviductal transport and the development of fallopian pregnancy .

蛋白酶切激活受体-2(PAR-2)是能被多种蛋白水解酶激活的G-蛋白偶联受体，调控多种平滑肌的运动，但PAR-2对输卵管平滑肌运动及转运的作用及机制还不清楚。根据预实验结果和文献分析，我们提出PAR-2表达于人和大鼠的输卵管，激活后通过前列腺素-前列腺素受体途径和PLC-IP3-PKC-L-型电压门控Ca2+通道途径兴奋输卵管平滑肌，并通过影响输卵管的胚胎转运参与输卵管妊娠的发生。本课题拟采用IHC、RT-PCR、WB检测PAR-2在人和大鼠输卵管组织的表达；采用张力记录、膜片钳、钙成像等技术研究PAR-2对输卵管平滑肌运动的影响及机制；采用动物在体实验，研究PAR-2对大鼠输卵管胚胎转运的影响；研究输卵管妊娠者输卵管中PAR-2的表达、功能改变及可能机制。拟通过上述研究明确PAR-2在输卵管运动及转运过程中的作用及机制，为输卵管转运功能的调控及输卵管妊娠的发生提供新的研究靶点和实验基础。

蛋白酶切激活受体-2(PAR-2)是能被多种蛋白水解酶激活的G-蛋白偶联受体，调控多种平滑肌的运动，但PAR-2对输卵管平滑肌运动的影响还不清楚。应激后PAR-2表达增加，肠粘膜屏障通透性增大，但应激激素NE是否调控PAR-2及紧密连接蛋白的表达还不清楚。瞬时受体电位通道A1(TRPA1)表达于多种非神经细胞上，参与炎症过程但其在子宫内膜上皮细胞炎症反应及氧化应激过程中的作用还不清楚。本研究采用IHC、WB、IF检测PAR-2在人和大鼠输卵管组织的表达，检测热应激后结肠及食管PAR-2及紧密连接蛋白Occludin,ZO-1的表达；采用张力记录技术研究PAR-2激活对输卵管平滑肌运动的影响；采用HE染色观察热应激后大鼠结肠及食管粘膜结构的改变；采用Elisa检测大鼠血浆NE的水平；分别培养Caco-2及人子宫内膜上皮细胞，并研究NE处理Caco-2后PAR-2及紧密连接蛋白的表达；研究人子宫内膜上皮细胞上TRPA1激活后，相关炎症及氧化应激因子的变化。结果显示：PAR-2表达于人和大鼠输卵管黏膜上皮及平滑肌，激活后促进输卵管平滑肌的运动；热暴露后大鼠食管、结肠PAR-2、TRPA1，紧密连接蛋白Occludin、ZO-1表达改变，NE处理Caco-2可调节PAR-2及Occludin, ZO-1的表达；TRPA1激活后影响iNOS,p-p65 536的表达，且效应随处理时间而不同，明显上调HO-1，上述效应不受L-NAME及indomethacin预处理的影响。结论：PAR-2激活后兴奋输卵管平滑肌；热应激后PAR-2、TRPA1及紧密连接蛋白的表达改变，NE可直接调节Caco-2 上PAR-2及紧密连接蛋白的表达；TRPA1参与人子宫内膜上皮细胞的炎性及氧化应激反应。