Abstract
摘要:由我們團隊先前的研究發現,若將邁入老化前的人類包皮纖維母細胞培養於幾丁聚醣基 材上時,細胞會懸浮聚集成球,三天後再將懸浮的細胞球移至一般的培養皿時,細胞自球中 逐漸爬出,且能有效地使老化細胞延長複製代數、提升增殖能力和減少老化表徵。因此,本 計晝希望藉由生醫材料改變細胞生長的微環境,促使老化細胞提升增殖能力和修復之功能 性。由目前臨床統計數據可知,老年者關節損傷機率高,且老化的生理環境也可能會減弱細 胞的自我修復能力。因此,結合先前的研究成果,我們將以幾丁聚醣基材處理關節腔内易損 耗之細胞:前十字韌帶細胞、滑膜細胞及軟骨細胞,期許未來本方法可應用於細胞治療的前 處理步驟,即先使老化細胞提升增殖和修復的能力,再行治療。且本計晝之方法屬非侵入性 治療,亦不涉及基因與病毒之轉殖,相信未來對病人而言,是一種安全性較高、風險較低且 效益較佳之治療方式。本計晝為三年之研究,各年特定目標如下:第一年計晝欲建立幾丁聚醣基材對前十字韌帶細胞提升增殖能力和延長複製代數的模式。研究之特定目標在探討透過材料參數和培養條件的調整,欲建立一最佳模式,其能有效 地影響前十字韌帶細胞體外培養的老化現象。第二年計晝將探討不同年齡前十字韌帶細胞、滑膜細胞、軟骨細胞於幾丁聚醣基材上的增殖表現。計晝將驗證關節炎患者其關節腔内的各類初級細胞增殖能力是否會受患者年齡影 響。也將探討不同年齡層病患不同細胞檢體,其直接分離後的增殖能力受幾丁聚醣調控之情 形。第三年計晝係探討前十字韌帶細胞、滑膜細胞、軟骨細胞於幾丁聚醣基材上的功能表現與模式之機制。傷口癒合的研究,除細胞增生外,細胞功能也相當重要,因此本計晝將探討 幾丁聚醣調控細胞分泌細胞間質和激素的量及遷移能力。最後,幾丁聚醣對細胞老化調節影 響的機制也將於此深入地探討。因此本計晝結束後,將可以分析不同年齡層病人的關節腔内細胞其增殖能力與患者年齡 之相關性;幾丁聚醣生醫材料是否能影響前十字韌帶、滑膜細胞與軟骨細胞的增殖特性和功能 性;同時建立幾丁聚醣影響細胞延長複製代數的模式機制。
Abstract: Fibroblasts have been extensively used as a model to study cell senescence. In our preliminary study, human foreskin fibroblasts on commercial tissue culture polystyrene (TCPS) progressively entered senescence after 55-60 population doublings (PDs). Before senescence was reached, fibroblasts could aggregate to form multicellular spheroids on chitosan. After 3 days of incubation, fibroblasts reseeded on TCPS could migrate out of spheroids to develop more proliferation ability and lower senescence-associated (3-galactosidase (SA (3-gal) activity to undergo additional doublings to extend the life span. Based on these findings, the purpose of this project is to apply the technique of the chitosan treatment-mediated life-span extension to different knee joint cells, including the anterior cruciate ligament cells, synovial cells and chondrocytes. It is expected that the technique might be useful to maintain senescent cells in a younger state for future cell therapy applications. In addition, the method is non-invasive therapy, and without genetic and viral transfect process, we believe it is more safety, lower risk, and better efficacy of treatment for patients in the future. The objectives in this project will be achieved through the following specific aims.Specific Aim 1: To establish the chitosan treatment-mediated life-span extension model of the anterior cruciate ligament cells. This aim will test the difference of in vitro replicate senescence between chitosan-treated and -untreated anterior cruciate ligament cells. The optimized model of the effect of chitosan biomaterials and culture conditions on ligament cells will be established.Specific Aim 2: To study the proliferation ability of the anterior cruciate ligament cells, synovial cells and chondrocytes before and after chitosan treatment. This aim will analyze the relationship of the proliferation of primary joint cavity cells and the ages of the arthritis patients. Further, the effect of the chitosan treatment-mediated life-span extension on cells from patients with different ages will be analyzed in this aim.Specific Aim 3: To study the functionality of the anterior cruciate ligament cells, synovial cells and chondrocytes after chitosan treatment and investigate the underlying mechanism of this model. In addition to cell proliferation, cell functionality such as the production of the extracellular matrix proteins and cytokines, and the ability of cell migration after chitosan treatment will be investigated, which are important in the wound healing process. Finally, the mechanism of chitosan treatment on cell behavior will be analyzed to improve cell therapy effectiveness in the future.At the end of this project, the outcome will be published in famous journals, and the entire people join this project will gain good training.
Keyword(s)
幾丁聚醣
增殖能力
前十字韌帶細胞
滑膜細胞
軟骨細胞
Chitosan
proliferation
the anterior cruciate ligament cells
synovial cells
chondrocytes