利用H202誘使培養於PVA細胞產生不可逆的老化

Abstract

前言：由過去組織工程的研究可知，不同生醫材料表面的化學和物理性質與細胞的生存、生長等細胞行為表現息息相關，但是對於細胞成球與老化的相關研究也較少。因此，本研究的主要目標為利用PVA親水性的表面使細胞成球，進而探討在PVA材料上培養的成球細胞生長和生存的情況，以及細胞是否會直接走向死亡或者是會走老化路徑及前趨性老化等可能發生的路徑。 材料與方法：實驗選用兩種不同種類的PVA（一個是鹼化程度較高的BP-20H、一個是黏度較高的BC-24）以及商業用組織培養盤(TCPS)，在細胞方面使用肺癌細胞株H1299及轉殖FAK的H1299細胞（H1299/ FAK），觀察比較其在PVA生長和生存的情況，再利用纖維母細胞(HFF)培養於不同的基材上，以不同條件之雙氧水處理，建立細胞長期之生長曲線(PD)與 SA-β-galactosidase酵素活性等老化相關指標，最後進行老化相關的基因分析。 結果： 細胞生長在PVA材料上，長時間的觀察方面H1299/ FAK 生長的情況比H1299較好WST-1讀值也較高。使用長時間低濃度雙氧水處理對HFF細胞的傷害似乎較短時間高濃度的大，HFF細胞經培養在PVA材料再移置TCPS的細胞較易變扁平老化的比例也較明顯。 結論：H1299 / FAK在PVA生長和生存的狀況較H1299良好。經由雙氧水處理可以誘使HFF細胞產生不可逆的老化，若處理後再培養在PVA材料上，會明顯影響細胞生長的型態進而產生類似於老化之現象。

Introduction: It has been known that different chemical and physical surface properties of biomembranes controlled the cell behavior, such as cell survival and growth. However, there is no report which is focused on the effects of biomembranes to cellular senescence. Thus, the objective of this study was to investigate the effects of cell aggregates cultivated on the surface of PVA biomembranes. The influences of biomembranes to the proliferation and senescence of cells were also studied. Materials and methods: We chose three kinds of biomembranes, PVA (polyvinyl Alcohol) BP-20H has high alkalization degree, PVA (polyvinyl Alcohol) BC-24 has high viscosity, and tissue culture polystyrene (TCPS). The survival and proliferation of H1299 and H1299/FAK when cultured on PVA membranes were investigated. Human foreskin fibroblast (HFF) were cultured on these membranes and received different conditions of H2O2 treatment to establish senescence correlation index, such as growth curves (P.D) and SA-β-galactosidase activity. Senescence associated gene expression was also analyzed. Results: Our results show that H1299/FAK has better survival and WST-1 value than the H1299. Prolonged low-dose H2O2 treatment not only induced irreversible cell growth arrest but also increased senescent cells, especially cultured on the PVA membranes. Conclusions: H1299/FAK has better survival on the PVA membranes than the H1299 cells. H2O2 treatment significantly induced irreversible cell growth arrest and the percentage of senescent cells to the HFF cells. This phenomenon could be further enhanced by transferring the H2O2 treated cells to the PVA membranes for subsequent cell culture.