波浪狀結構及機械拉伸調控間質幹細胞結構與分化

Abstract

韌帶由平行波浪狀結構的膠原蛋白纖維所組成，而此波浪狀結構提供韌帶良好的機械性質並給予韌帶細胞較好的行為表現。之前研究發現波浪狀電紡絲結構相較於直線電紡絲更能增進韌帶細胞的基因表現。基於探討不同曲率是如何影響細胞行為表現，我們製作了直線及不同彎曲度的PDMS基質並種入間質幹細胞。結果發現在直線及不同曲率中細胞型態有顯著性的差異；在0.5X wavy pattern中細胞擁有最大的細胞及細胞核，而細胞核的曲率隨著pattern曲率增加而增加。之前研究提到 ROCK 及 MLCK 是重要的兩個途徑影響細胞骨架及調控細胞行為。當我們給予ROCK與MLCK抑制後，發現細胞型態受到影響且mRNA表現也有所差異。此外，我們亦給予動態拉伸刺激發現拉伸過後在0.5X wavy pattern中細胞及細胞核尺寸都變小。我們猜測ROCK與 MLCK 在直線及不同曲率結構扮演不同的角色去調控細型態。之後我們會更進一步探討ROCK與 MLCK在拉伸中如何調控細胞的行為。

Native ligament tissues are composed by aligned-wavy collagen fibers and the wavy structures are believed to provide ligament with mechanical properties and to improve ligament fibroblast behaviors. Previous studies found that cells on electrospun fibers with wavy structures enhanced ligament phenotypes than on straight fibers. In order to investigate the relationship between the degree of waviness and cell behaviors, we fabricated PDMS microgroove substrates with straight and different wavy structures and then seeded human mesenchymal stem cells (MSCs) on the microgrooves. We observed distinct cell morphologies on straight and different wavy structures. On 0.5X wavy structures, cells displayed the largest cell and nucleus sizes and nucleus curvature increased in accordance with increasing curvature of wavy structures. Previous studies found that ROCK and MLCK are important mediators to affect cell cytoskeleton and regulate cell behaviors. Upon ROCK and MLCK inhibitor treatments, we found cell morphologies and mRNA expression were affected by ROCK and MLCK. Additionally, we provided tensile loading. In 0.5X wavy group, cell and nucleus sizes became smaller compared to the control group after tensile loading. We hypothesize that ROCK and MLCK have different role in straight and wavy groups resulting in cell phenotype. In the future, we will understand ROCK and MLCK how to regulate cell phenotypes on tensile loading even more.