2011-08-012024-05-13https://scholars.lib.ntu.edu.tw/handle/123456789/646580摘要：Statins 除廣泛作為降血脂的藥物外，也具有多重治療效果，而這些多重性的治療潛力的作用機轉主要是由於statins 會造成異戊二烯 (isoprenoid)生合成路徑的受損，抑制小分子G 蛋白質的訊息傳遞路徑，進而調節細胞功能達到多元的治療效果。近年研究發現在單核白血球或是人類週邊血液單核球細胞，給予statins 會造成capsase-1 的活化而誘導IL-1β的釋出，然而詳細的分子作用機轉仍待釐清。目前我們初步的實驗結果顯示fluvastatin 及lovastatin 的確可與脂多醣體 (LPS)可協同誘發NLRP3 活化，且此作用與geranylgeranyl pyrophosphate (GGPP)的減少有關。因此，本研究將更深入探討statin 刺激caspase-1 活性的詳細作用機制。目前報導已知NLRP3 inflammasome 的活化至少可受控於三大主要路徑，分別是 (1)外生性ATP 鍵結P2X7 受體促使細胞內鉀離子流失，(2)細胞內活性氧化物(ROS)堆積及 (3)溶酶體破裂(lysosomal rupture)釋出蛋白酶(如cathepsinB)。在此計畫我們將於人類單核球細胞株(THP-1)，老鼠骨髓巨噬細胞(BMDM) 及RAW264.7 巨噬細胞，針對此三條路徑進行statins 誘導caspase-1 活化的機制探討。由於NLRP3 inflammasome 是一個由NLRP3、 ASC 和 pro-caspase-1 蛋白所組合而成的一個聚合體，我們將於293T 和HeLa 重新組合NLRP3 inflammasome 的元件，以探討statins對NLRP3 inflammasome 聚合及活化的作用。我們也將探討異戊二烯化修飾蛋白作用和inflammasome 活化其兩者之間的連結關係。綜合以上，我們的研究將為異戊二烯化修飾蛋白作用如何調控NLRP3 inflammasom 提供一個新的觀點，並釐清statins 活化inflammasome 的分子作用機制。<br> Abstract: The HMG-CoA reductase inhibitors, namely statins, are widely used for treatment oflowering cholesterol in clinical. However, statins are not only therapeutically administered inhypercholesterolemia but also have multiple therapeutic potentials, such as inanti-inflammation, anti-tumor, and immunomodulation. Most of these pleiotropic and atypicalactions result from the impairment of isoprenoid biosynthesis and interference with signalcascades mediated by small G proteins (e.g. Ras, RhoA, Rac, Rab, Cdc42). Recent studiesfurther unexpectedly observe the stimulating effects of statins on caspase-1 activation andIL-1β secretion in monocytes and peripheral blood mononuclear cells, and suggest the crucialroles played by isoprenoids in the inhibition of caspase-1 activity. Nevertheless the molecularmechanism in detail has not been clearly elucidated. Currently our preliminary data confirmthat fluvastatin or lovastatin treatment can synergize with LPS to trigger inflammasomeNLRP3 activation. Moreover, statins-stimulated caspase-1 activation and IL-1β production inLPS-primed THP-1 cells are related to GGPP deficiency. Therefore, in this study we attemptto dissect the molecular mechanisms underlying the stimulating effects of statins on caspase-1.To this end, three commonly recognized mechanistic models for NLRP3 inflammasomeactivation (i.e. ATP/P2X7/K+ efflux, ROS and lysosomal rupture) will be investigated instatin-induced caspase-1 activation in human THP-1 monocytes, mouse bone marrow-derivedmacrophages and RAW264.7 macrophages. Since NLRP3 inflammasome is an assemblycomplex composed by NLRP3, ASC and pro-caspase-1, we will reconstitute inflammasomecomponents with pro-IL-1β in 293T and HeLa cells, the cell lines lacking inflammasomeproteins, to analyze effects of statins on NLRP3 inflammasome formation and activation. Inaddition, the molecular link between isoprenylation and inflammasome activation will also beelucidated. Taken together, this study will provide signaling basis underlyinggeranylgeranylation-dependent regulation of NLRP3 inflammasome in monocytes andmacrophages, and will lead to a clearer understanding on statin-elicited inflammasomeactivation.trophoblastinvasionhanging drop culturespheroid3D culturetransforming growth factor