探討微米尺寸的基底結構對於細胞貼附型態與生長的影響

Abstract

為了瞭解並調控細胞與移植材料介面間的作用行為，我們製造了一維的密集線與二維的密集柱狀陣列，兩種不同地貌的表面，試圖去了解材料的表面圖案是如何影響細胞貼附後的種種行為。利用微影技術，不僅可以容易的定義出欲探討的表面圖案，更可以精準的製作成我們想要研究的的地貌尺寸。在實驗中，以微影曝光並加以蝕刻製作成之矽基材，做為母模，利用翻模的方式，製作了具有1微米大小的線寬與柱狀直徑表面的PDMS基材，其圖案深度皆為350奈米，希望藉此探討微米尺度下的地貌表面，對於子宮頸癌細胞貼附及生長的調控。 在我們的實驗結果發現在一維密集線表面生長貼附的細胞，傾向與溝槽平行且對其排列生長，與二維的密集柱狀陣列表面與平滑的材料表面相比較，生長於平行溝槽的細胞整合蛋白(integrin-□5)的表現增加，且細胞有更為良好貼附的現象。此外，在細胞的型態上，貼附於二維的密集柱狀陣列表面與平滑的材料表面細胞，多呈現圓球態表面多絲狀偽足伸出，貼附狀況差，伴隨著腫瘤抑制蛋白(p53)的表現，基質金屬蛋白酶9(MMP-9)的表達增高。 由結果我們得知，藉由外在地貌的差異，進而影響由細胞表面整還蛋白(integrin)所介導的訊號傳遞，造成了細胞生長、DNA合成、細胞移動以及凋亡的不同結果產生。如此，藉由改變細胞貼附的基底材質地貌，進而改變細胞中由整合蛋白所介導的訊號傳遞而影響細胞行為，將會是生醫材料及生醫工程上很好的應用及操作細胞的方式。

In order to find the ways for controlling the cell-material interface, we made two different topographies of one dimensional (1D) periodic lines/space pattern and two dimensional (2D) arrayed pillars pattern for cell behavior analysis. Here we used lithographic techniques can control not only the topographic pattern but also the scale of such topography within microscale ridge widths (~1□m) and submicron deep grooves (~350nm). We investigated the microscal topography regulated cell functions using human epithelial carcinoma (HeLa) cell culture on poly(dimethylsiloxane) (PDMS), the silicon substrate with microstructures on it were used as templates for micromolding a silicon elastomer, PDMS, into tissue scaffolds for cell patterning purpose. We observed that on 1D periodic lines surface cells tend to align with the direction of microscale ridges and grooves and have better attachment through an integrin □5 subunit expression compared with 2D periodic pillars pattern or flat PDMS surfaces. By contrast, cells cultured on the 2D periodic pillars and smooth PDMS substrates were mostly round and worse adherent with higher filopodia protruded, tumor suppressor protein 53 (p53) increased and matrix metalloproteinase-9 (MMP-9) expression. Considering the important role of integrin-mediated intracellular signaling in anchorage-dependent cell function, we found the external topography regulate cell function as cell growth, DNA synthesis, motility, and apoptosis. Modulation of cellular morphology related integrin-mediated signaling by altering substrate topography will have useful applications in biomaterials science and tissue engineering.