2014-08-012024-05-13https://scholars.lib.ntu.edu.tw/handle/123456789/645944目前全世界大約有超過十萬名病患使用腹膜透析治療以維持其身體的健康，這些病患 大約佔所有接受透析的人口的百分之十至十五左右。影響這些腹膜透析病患成功治療的主 要障礙是透析藥水引起的腹膜功能的改變，而這些腹膜功能的改變已被證實影響到病患的 健康與死亡率。超過一半的病患會因透析藥水而改變其腹膜功能，這些變化牽涉到漸行性 的腹膜纖維化、血管新生、血管退化，並增加溶質通透性與降低超過濾的效果。當中發現 極少數的的病患會發生極度嚴重的腹膜纖維化，即包囊性腹膜硬化症(encapsulating peritoneal sclerosis)，往往造成死亡。雖然近年來在接管系統與新腹膜透析藥水的研發已大 為改善所謂的生物相容性並降低感染性腹膜炎發生率，但伴隨腹膜透析治療而來的腹膜纖 維化與功能喪失卻仍然無法有效克服。因此更深入了解腹膜功能退化的分子機轉與研究促 進受傷腹膜再生就越顯重要。結構上腹膜由腹膜間皮細胞、腹膜間皮下基質母細胞、及細 胞外基質所構成。造成腹膜纖維化的肌肉纖維母細胞究竟從何種細胞分化而來，以及是否 有幹細胞能夠再生以修補受傷害的腹膜間皮細胞仍然具有相當多的爭論。腹膜間皮細胞受 傷害之後轉分化為肌肉纖維母細胞是目前最廣為接受的假說之一，不過這些假說的立論基 礎主要來自免疫化學染色的研究，真正在生體上觀察到腹膜間皮細胞受傷害之後轉分化為 肌肉纖維母細胞的證據則是闕如。最近我們利用基因改造小鼠進行的研究證實急性腎臟傷 害後修補受傷害腎小管上皮細胞的細胞來源是存活下來的腎小管上皮細胞，而造成纖維化 的間質纖維母細胞的真正來源是血管周邊細胞。這些都與上皮細胞-間葉細胞的轉分化無 關。因此我們將在本研究中先利用以第一型膠元蛋白促進子所控制的綠色螢光蛋白質小鼠 來研究正常與受傷害的腹膜中製造膠元蛋白質的細胞真相。然後我們將利用可以專一地在 腹膜間皮細胞或腹膜間皮下基質細胞表現螢光蛋白質的小鼠，在特定的時間表現其螢光蛋 白以利於追蹤這些腹膜間皮細胞或腹膜間皮下基質細胞的命運。 我們也將利用骨髓移植的 技術來探討自骨髓而來的細胞是否會分化成腹膜肌肉纖維母細胞或修補受傷的腹膜間皮細 胞。我們將接著研究腹膜間皮細胞或腹膜間皮下基質細胞在受到腹膜纖維化的刺激後其轉 錄組的變化，以供後續尋找預防腹膜纖維化及促進腹膜間皮細胞修復的治療標的。由於過 去已知Wilms’ tumor 1在胚胎時期器官生成時的間葉細胞轉分化為上皮細胞時扮演非常重 要的角色，而我們的初步結果顯示腹膜間皮下基質細胞會在腹膜傷害後活化及表現Wilms’ tumor 1。因此我們將專一性地在引發腹膜纖維化時將腹膜間皮下基質細胞的Wilms’ tumor 1 剔除，以觀察其對於腹膜纖維化及腹膜間皮細胞修復的影響。利用這些可靠的動物模式以 及先進的基因改造鼠將可以進一步了解腹膜纖維母細胞與腹膜間皮細胞修補的真正來源與 機轉。本研究的結果將有助於在臨床上思考如何維持健康的腹膜間皮細胞及阻斷腹膜纖維 化，讓腹膜透析的患者可以維持更健康的透析治療。對於手術後的腹膜沾黏也可提供進一 步的治療標的。Peritoneal dialysis (PD) is a life-sustaining therapy used by > 100,000 patients with end-stage renal disease worldwide, accounting for approximately 10 to 15% of the dialysis population. One of the major obstacles to successful long-term PD is deleterious functional alteration in the peritoneal membrane after exposure to dialysis solutions; this loss of dialysis capacity is responsible for increased morbidity and mortality. These alterations, involving approximately 50% of all PD patients, include progressive fibrosis, angiogenesis, and vascular degeneration associated with increased solute transport and loss of ultrafiltration. In a small percentage of cases, a poorly defined but catastrophic fibrogenic response occurs primarily in the visceral peritoneum, leading to the onset of encapsulating peritoneal sclerosis (EPS) with an associated high mortality rate. Although advances in new dialysis solutions and connection systems improve the biocompatibility and reduce the incidence of peritonitis, membrane failure due to progressive fibrosis remains a big problem. There is thus a growing need to understand the molecular basis of these membrane-degenerative events and a need to improve the regeneration of injured peritoneum. Peritoneum is basically composed of mesothelial cells, submesothelial stromal cells and extracellular matrix. It is highly controversial about the origin of peritoneal myofibroblasts responsible for the progressive fibrosis and whether there are progenitor cells to regenerate injured mesothelium. The popular hypothesis that mesothelial cells contribute to the most peritoneal myofibroblasts through epithelial-mesenchymal transition is proposed based on the immunohistochemical staining. This hypothesis lacks direct evidence in vivo. We have recently used the genetically engineered mice to study the origin of regenerated tubular epithelial cells in acute kidney injury and interstitial myofibroblasts in chronic progressive kidney fibrosis. We have clearly shown that the regeneration by surviving tubular epithelial cells is the predominant mechanism of repair after ischemic tubular injury and the origin of interstitial myofibroblasts are pericytes, not from epithelial-mesenchymal transition of injured tubular epithelial cells. In this study we will first use the green reporter mice whose GFP is driven by collagen I (α1) promoter to study the collagen producing cells in normal and injured peritoneum. We will then use cell-specific and time-specific genetically fluorescence-labelled mesothelial cells and submesothelial stromal cells to trace the origin of the collagen-producing cells and regenerated mesothelial cells in injured peritoneum. Bone marrow chimeric mice with bone marrow from different reporter mice will help us clarify the role of bone marrow derived cells in the peritoneal myofibroblasts and regenerated mesothelial cells. This state-of-the-art and most reliable fate-tracing technique will lead us to understand the origin of both peritoneal myofibroblasts and progenitor cells to repair injured mesothelium. We will next study the evolution of transcriptomes of mesothelial and submesothelial stromal cells during peritoneal injury by microarray analyses that will unravel novel targets for prevention of peritoneal fibrosis and promotion of mesothelial regeneration. We will further study the role of Wilms’ tumor 1 in the peritoneal fibrosis by conditional knock out in submesothelial stromal cells. Through this study we may advance our knowledge how to maintain the healthy mesothelial cell mass and specifically prevent the formation of pathological myofibroblasts. The results may also help the development of prevention and therapy for adhesion after abdominal surgery.腹膜透析腹膜間皮細胞腹膜間皮下基質母細胞腹膜纖維化祖細胞peritoneal dialysismesothelial cellssubmesothelial stromal cellsperitoneal fibrosisprogenitor cell