2010-08-012024-05-13https://scholars.lib.ntu.edu.tw/handle/123456789/645313摘要：蛋白尿是腎絲球疾病的生物標記。它可被視為腎絲球屏障受傷的後果，但同時也可以刺激腎小管細胞，誘導腎小管間質發炎和纖維化。因此，無論是何種病因，減輕或阻斷蛋白尿嚴重度已成為治療蛋白尿型腎臟病的最主要目標之一。目前用於控制蛋白尿的治療藥物包括類固醇和免疫抑制劑 (用於腎絲球腎炎)，腎素-血管張力素-皮質醛酮素系統阻斷劑 (ACEi, ARB, aldosterone blocker, 和直接腎素抑制劑)，和其它輔助性藥物如statins, PPARgamma agonists, 和 pentoxifylline。在這些藥物中， pentoxifylline 是一種非特定性的磷酸雙脂酶抑制劑，它單獨或與血管張力素阻斷劑合併，已被廣汎地用於治療蛋白尿型腎臟病。雖然此藥的降蛋白尿效果只算中度，但相對於血管張力素阻斷劑，它不致產生低血壓、腎衰竭、或高血鉀等副作用，安全性較高。不過，即便pentoxifylline 的降蛋白尿效果已被認可，但其作用機轉卻尚未釐清。目前只知道它降尿蛋白同時，也降低尿中發炎激素 (TNF-alpha) 和趨化因子(MCP-1)。但除此之外，我們對於其降蛋白尿的分子機轉，完全不清楚。過去十年間，有關蛋白尿的分子致病機轉研究進展快速，迄今所累積的資料顯示腎絲球過濾屏障扮演極重要的角色，其中又以slit-diaphragm 和足細胞蛋白最為重要。同時，亦有研究發現發炎激素 (TNF-alpha, IL-1beta)、趨化因子 (CCL2/MCP-1)、和TGF-beta1具有抑制slit-diaphragm 和足細胞蛋白基因轉錄的作用。最近足細胞被報告會表現另一種趨化因子，CX3CL1/fractalkine，的受器，暗示此趨化因子亦可能如同MCP-1，影響足細胞的移動。因此，本計畫假設pentoxifylline 抑制腎絲球病變蛋白尿的主要機轉是透過抑制足細胞內特定磷酸雙脂酶 (如 4B 亞型)，再經由細胞內cAMP (或 cGMP) 升高，活化其標的蛋白酶 (如蛋白酶A)，進而恢復受到發炎媒介物 (TGF-beta1, TNF-alpha) 或趨化因子 (CX3CL) 所抑制的足細胞slit-diaphragm 蛋白表現，並修補足細胞的屏障功能。為證實此一假說，我們預計在未來兩年利用足細胞和動物模式研究pentoxifylline 降低蛋白尿的機轉，具體目標包括：(1) 探討pentoxifylline 對 (a) TGF-beta1 所改變的足細胞slit-diaphragm 蛋白表現的調控及機轉；(b) CX3CL1 所刺激之足細胞移動的調控及機轉；(c) 大鼠 anti-Thy1 腎炎模式 (無直接足細胞傷害仍併發蛋白尿)，蛋白尿變動相對於slit-diaphragm 和足細胞蛋白表現的調控作用及機轉。(2) 探討pentoxifylline 對 (a) TNF-alpha 所改變的足細胞slit-diaphragm 蛋白表現的調控及機轉；(b) 小鼠 adriamycin nephrosis 模式 (有直接足細胞傷害且合併蛋白尿)，蛋白尿變動相對於slit-diaphragm 和足細胞蛋白表現的調控作用及機轉。吾人預期將來本計畫所獲得的成果，除了瞭解pentoxifylline 這類磷酸雙脂酶抑制劑降蛋白尿的分子機轉外，更可以引領出治療蛋白尿型腎臟病的新式策略。<br> Abstract: Proteinuria (albuminuria) is both a marker of injured glomerular barrier, and a predictorfor end-stage renal disease in glomerular disorders. As such, measures to reduce proteinuriahas become a standard therapeutic goal in the management of proteinuric kidney diseases.Current antiproteinuric therapies include corticosteroids and immunosuppressive drugs (in theevent of glomerulonephritis), blockers for the renin-angiotensin-aldosterone system, as wellas other adjunctive agents such as statins, PPARgamma agonists, and pentoxifylline (anonselective phosphodiesterase (PDE) inhibitor). We and others have shown thatpentoxifylline can attenuate proteinuria in patients with diabetic and non-diabetic proteinurickidney diseases. However, despite the well-recognized antiproteinuric efficacy of this drug,the mechanism underlying its action remains largely unexplored. So far, we only know thatpentoxifylline lowers proteinuria in association with reduction of urinary TNF-alpha andMCP-1.Over the past decade, accumulating data indicates that glomerular barrier, in particularthe podocyte slit-diaphragm apparatus, plays the most critical part in the pathogenesis ofproteinuria. There is also evidence showing that inflammatory cytokines (TNF-alpha),chemokines (CCL2/MCP-1), or TGF-beta1 downregulate the expression of the slit-diaphragmpodocyte proteins. More recently, podocytes are reported to display the receptor forCX3CL1/fractalkine, which presumably might affect podocyte mobility as does MCP-1.Given these facts, we hypothesize that pentoxifylline may exert its antiproteinuric effectthrough suppression of podocyte PDE isoenzymes (such as PDE4B); the resultant increase inintracellular cyclic nucleotides (presumably cAMP) then activates target protein kinases (eg.,protein kinase A), which in turn orchestrate the restoration of dysregulated slit-diaphragmpodocyte proteins induced by inflammatory mediators (TGF-beta1, TNF-alpha) orchemotactic cytokines (CX3CL1).To substantiate this hypothesis, the present study will be conducted both in culturedpodocytes, and in animal models with or without direct podocyte injury. In the next two years,we aim to demonstrate:(1) the modulatory effect of pentoxifylline on (a) TGF-beta1-altered slit diaphragm proteinexpression and CX3CL1-induced podocyte motility in vitro; (b) dysregulated expressionof podocyte-specific markers in relation to proteinuria in rats with anti-Thy1 nephritis (amodel without direct podocyte injury) and the mechanism act by this drug in vivo.(2) the modulatory effect of pentoxifylline on (a) TNF-alpha-altered slit diaphragm proteinexpression and the mechanism whereby it acts in vitro; (b) dysregulated expression ofpodocyte-specific markers in relation to proteinuria in mouse adriamycin nephrosis (amodel with direct podocyte injury) and the mechanism act by this drug in vivo.It is hoped that results obtained from this study may shed light on the molecularmechanism of the antiproteinuric effect of pentoxifylline in glomerular disorders and lead to anovel anti-proteinuric strategy in the management of proteinuric kidney diseases.