- 學位論文
臍帶間質組織衍生之內皮前驅細胞具有抑制內皮受損後內膜增生之作用: 藉由色素性上皮細胞衍生因子參與抑制作用
Late outgrowth endothelial cells derived from Wharton’s jelly in human umbilical cord reduce neointimal formation after vascular injury: involvement of pigment epithelium-derived factor
摘要
在以細胞植入治療心血管疾病方面，欲由骨髓及周邊血液中分離出具有潛能的內皮前驅細胞，結果發現骨髓及周邊血液中的內皮前驅細胞不論在數量或品質上都受到相當大的限制。本篇研究之主要目的為檢測可否從臍帶間質(Wharton’s jelly in human umbilical cord, WJC)中分離、培養並分化出內皮前驅細胞，及探討其是否可影響內皮受損後的內膜增生。首先由臍帶間質分離出的間質細胞(mesenchymal cells, MCs)，培養於適合內皮細胞生長的培養液(EGM-2)中可以分化成內皮前驅細胞(late outgrowth endothelial cells) (WJC-OECs)，其具有吞噬乙醯化的低密度脂蛋白(acetylated-LDL)之能力，並且表現出內皮細胞特有的蛋白。利用彈性線圈將老鼠股動脈進行去內皮手術後，再將這些已分化的人類內皮前驅細胞經由尾靜脈注射植入老鼠體內，結果發現，這些植入的細胞能夠快速到達內皮受損區，進行血管內皮修補(re-endothelialization)的作用。在去內皮手術後及細胞植入4周後，相較植入saline、MCs及臍帶血衍生之內皮前驅細胞(cord blood-OEC, CB-OECs)之組別，發現植入WJC-OECs之組別中，血管內膜層與中膜層之比例有顯著的降低(2周組，saline: 1.48±0.14, MC: 1.32±0.18, WJC-OEC: 0.95±0.05, CB-OEC: 1.32±0.05; 4周組，分別為saline: 1.81±0.25, MC: 1.68±0.09, WJC-OEC: 01.01±0.07, CB-OEC: 1.39±0.14)，而且在內皮受損區，經由組織免疫染色法觀察有大量的色素性上皮細胞衍生因子(pigment epithelium-derived factor, PEDF)表現，相同的結果也可在細胞培養時觀察到；進一步發現WJC-OECs的培養液會抑制人類主動脈平滑肌細胞移行及增生；而藉由加入PEDF抗體或轉染PEDF siRNA來減弱WJC-OECs之PEDF的表現後，顯著抑制其影響。本篇報告是第一篇陳述臍帶間質細胞具有分化成內皮前驅細胞的潛能，而且對於血管內皮細胞受損後的再修補扮演著相當重要的角色，可維持血管內皮的完整，達到抑制內膜增厚的作用。綜合本研究的結果顯示，在血管受損相關疾病的臨床治療，WJC-OEC可視為細胞治療法中的另一個新契機。
並列摘要
The number of endothelial progenitor cells (EPCs) that can be obtained from adult bone marrow and peripheral blood to treat cardiovascular diseases is limited. The goal was to examine the endothelial potential of Wharton’s jelly in human umbilical cord (WJC)-derived stem cells and evaluate their potential to affect neointimal formation after vascular injury. Mesenchymal cells (MCs) were isolated from WJC and cultured in endothelial growth medium. Differentiation into late outgrowth endothelial cells (WJC-OECs) was demonstrated by incorporation of acetylated low-density lipoprotein and expression of the endothelial-specific markers. Transplantation of these cells into wire-injured femoral arteries in mice led to rapid re-endothelialization. At 4 weeks after injury, the neointima/media area ratio was reduced and strong expression of pigment epithelium-derived factor (PEDF) compared to saline- or MC- or cord blood-OEC- treated mice. WJC-OECs-conditioned medium has an extremely strong capacity to inhibit the migration and proliferation of smooth muscle cells. The effects were inhibited by neutralizing antibody for PEDF and by siRNA silencing of PEDF. We firstly demonstrated the presence of a cell population within WJC that has the potential to differentiate into OECs. Transplantation of WJC-OECs may play a crucial role in reestablishing endothelial integrity in injured vessels, thereby inhibiting neointimal hyperplasia. These findings have implications for a novel and practical cell-based therapy for vascular diseases.
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