【摘要】：凝血酶、胰蛋白酶、類胰蛋白酶和彈性蛋白酶等絲氨酸蛋白酶能調(diào)節(jié)凝血和纖維蛋白溶解平衡、降解神經(jīng)肽以參與神經(jīng)性水種，調(diào)控補(bǔ)體系統(tǒng)從而在炎癥反應(yīng)時(shí)發(fā)揮免疫調(diào)節(jié)作用。近年的研究顯示，它們還能通過蛋白酶激活受體(proteinase activated receptor，PARs)調(diào)節(jié)細(xì)胞的功能而在炎癥和免疫反應(yīng)中起重要作用。PARs是新的G蛋白偶聯(lián)受體，目前已發(fā)現(xiàn)有四種PARs，分別為PAR-1、PAR-2、PAR-3和PAR-4，凝血酶可以激活PAR-1、PAR-3和PAR-4，胰蛋白酶可以激活PAR-1、PAR-2和PAR-4，類胰蛋白酶和彈性蛋白酶可以激活PAR-2。單核細(xì)胞和T淋巴細(xì)胞是炎征和免疫反應(yīng)重要的效應(yīng)細(xì)胞，但是PARs在人外周血單核細(xì)胞和T細(xì)胞上表達(dá)及它們分泌的細(xì)胞因子上的影響則知之甚少。 采用磁激活的細(xì)胞分選法(magnetic activated cell sorting，MACS)純化人外周血單核細(xì)胞和T細(xì)胞，分別用RT-PCR和流式細(xì)胞術(shù)檢測外單核細(xì)胞和T細(xì)胞上的PARs在基因和蛋白質(zhì)水平的表達(dá)；用凝血酶、胰蛋白酶、類胰蛋白酶和彈性蛋白酶及PARs的激動肽分別激發(fā)MACS純化后單核細(xì)胞和淋巴細(xì)胞16小時(shí)后，用ELISA檢測單核細(xì)胞培養(yǎng)上清液中的IL-6、IL-1β和IL-12，，T細(xì)胞培養(yǎng)上清液中的IL-6和IL-13。 實(shí)驗(yàn)顯示，人外周血單核細(xì)胞表達(dá)PAR-1、PAR-3和PAR-4但不表達(dá)PAR-2蛋白質(zhì)，單核細(xì)胞表達(dá)PAR-1和PAR-3，但是不表達(dá)PAR-2和PAR-4 mRNAs。凝血酶、胰蛋白酶、類胰蛋白酶和彈性蛋白酶呈濃度依賴性地誘導(dǎo)單核細(xì)胞產(chǎn)生IL-6，當(dāng)這些蛋白酶酶活性被它們的抑制劑抑制后，它們誘導(dǎo)單核細(xì)胞產(chǎn)生IL-6的能力被抑制。PAR-1激動肽SFLLR-NH_2和PAR-4激動肽GYPGQV-NH_2可濃度依賴性地誘導(dǎo)單核細(xì)胞產(chǎn)生IL-6，PAR-1反激動肽RLLFS-NH_2和PAR-4反激動肽VQGPYG-NH_2不能誘導(dǎo)單細(xì)胞產(chǎn)生IL-6，而PAR-3激動肽TFRGAP-NH_2和反激動肽PAGRFT-NH_2對于單核細(xì)胞產(chǎn)生IL-6無明顯作用。蛋白酶及PAR激動肽在各自工作濃度對于單核細(xì)胞產(chǎn)生IL-1β和IL-12無明顯作用。 實(shí)驗(yàn)還顯示，人外周血T細(xì)胞表達(dá)PAR-1、PAR-和PAR-3但不表達(dá)PAR-4蛋白質(zhì)，與流式細(xì)胞結(jié)果一致，T細(xì)胞表達(dá)PAR-1、PAR-2和PAR-3，但是不表達(dá)PAR-4 mRNAs。凝血酶、胰蛋白酶、類胰蛋白酶呈濃度依賴性地誘導(dǎo)T細(xì)胞產(chǎn)生IL-6，當(dāng)這些蛋白酶酶活
[Abstract]:Serine proteases such as thrombin, trypsin, trypsin and elastase regulate coagulation and fibrinolysis balance, and degrade neuropeptides to participate in neurotic water. Regulate the complement system and thus play an immunomodulatory role in the inflammatory response. Recent studies have shown that they also play an important role in inflammation and immune response through protease activated receptor (proteinase activated receptor,PARs) regulation of cell function. PARs is a new G-protein-coupled receptor, and four kinds of PARs, have been found. PAR-1,PAR-2,PAR-3 and PAR-4, thrombin can activate PAR-1,PAR-3 and PAR-4, trypsin can activate PAR-1,PAR-2 and PAR-4, trypsin and elastase can activate PAR-2.. Monocytes and T lymphocytes are important effectors of inflammation and immune response, but the effect of PARs on the expression of PARs on human peripheral blood monocytes and T cells and the cytokines secreted by them is little known. Human peripheral blood monocytes and T cells were purified by magnetic activated cell sorting (magnetic activated cell sorting,MACS), and the expression of PARs on outer monocytes and T cells was detected by RT-PCR and flow cytometry, respectively. Monocytes and lymphocytes were stimulated by thrombin, trypsin, elastase and PARs activation peptide after MACS purification for 16 hours respectively. ELISA was used to detect IL-6,IL-1 尾 and IL-12, in the supernatant of monocyte culture. IL-6 and IL-13. in supernatant of T cell culture Human peripheral blood monocytes expressed PAR-1,PAR-3 and PAR-4 but did not express PAR-2 protein. Monocytes expressed PAR-1 and PAR-3, but did not express PAR-2 and PAR-4 mRNAs.. Thrombin, trypsin, trypsin and elastase induce monocytes to produce IL-6, in a concentration-dependent manner when the activity of these proteases is inhibited by their inhibitors. Their ability to induce IL-6 production in monocytes was inhibited. PAR-1 activator SFLLR-NH_2 and PAR-4 GYPGQV-NH_2 could induce IL-6, production in a concentration-dependent manner. PAR-1 RLLFS-NH_2 and PAR-4 VQGPYG-NH_2 could not induce the production of IL-6, in monocytes, but PAR-3 TFRGAP-NH_2 and PAGRFT-NH_2 had no effect on IL-6 production in monocytes. Protease and PAR activator peptide had no significant effect on the production of IL-1 尾 and IL-12 in monocytes at their respective working concentrations. The results also showed that human peripheral blood T cells expressed PAR-1,PAR- and PAR-3 but did not express PAR-4 protein. In accordance with the flow cytometry results, T cells expressed PAR-1,PAR-2 and PAR-3, but did not express PAR-4 mRNAs.. Thrombin, trypsin, trypsin induce T cells to produce IL-6, in a concentration-dependent manner when these proteases are active.
【學(xué)位授予單位】：汕頭大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2006
【分類號】：R392

【引證文獻(xiàn)】

相關(guān)碩士學(xué)位論文 前1條

1 楊雪;傷寒沙門氏菌致病仔豬胃腸道組織中蛋白酶激活受體2的定位研究[D];華中農(nóng)業(yè)大學(xué);2007年

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