電場及膠原蛋白基質的厚薄與排列對韌帶細胞移動的影響

Abstract

本論文旨在研究電刺激在不同排列和厚度的膠原蛋白纖維裡對於韌帶細胞移動的影響。在生物體內，細胞原來就生長在有不同軟硬度的環境，而我們所研究的前十字韌帶細胞原生在具有高度排列的膠原蛋白纖維裡，因此本研究有有排列�無排列及不同厚薄的膠原蛋白基質。我們用不同厚薄的膠原蛋白基質是因為根據文獻顯示，對細胞而言，不同厚薄的膠原蛋白基質會有類似不同軟硬度基質的效果。我們的實驗發現細胞在這些不同排列和厚薄的膠原蛋白上會有不一樣的形態，而膠原蛋白纖維的排列會影響細胞移動的方向：在無排列的膠原蛋白裡的細胞無特別的移動方向；然而在有排列的膠原蛋白裡的細胞會沿著纖維方向移動。 當施加電場於無排列的膠原蛋白纖維時，不管在薄的還是厚的膠原蛋白基質裡的細胞都會往電場負極移動。而當施加的電場方向平行於有排列的纖維時，不管在薄的還是厚的膠原蛋白裡細胞都會沿著纖維方向移動，但不會特別往電場負極或正極移動，這代表纖維走向比電場更有影響力。當細胞的RhoA訊號被抑制住時，在薄的膠原蛋白裡的細胞會往電場負極方向移動，然而在厚的膠原蛋白裡的細胞只會沿著纖維方向移動並不會特別往負極移動。 我們的研究顯示，細胞在薄的膠原蛋白裡是運用RhoA來感受膠原蛋白纖維走向，但在厚的膠原蛋白裡則不是。 這也說明了細胞在不同厚薄的膠原蛋白裡是經由不同的訊號傳遞來感受膠原蛋白纖維走向。

This work investigates the effects of electric field (EF), collagen gel thickness and collagen fiber alignment on ligament fibroblast migration. ACL cells live in highly aligned collagen fibers in vivo and studies had shown that cell can feel different substrate stiffness, and different thickness collagen gel have different effective modulus. Therefore, we develop a system that can produce highly aligned collagen fiber and control the thickness of collagen gel. We had two kinds of collagen fiber orientation, random and aligned and two kinds of collagen gel thickness. Cells have different morphology when they were seeded on different collagen fiber orientation and collagen gel thickness. When EF was applied, contact guidance is more potent than EF both in thin and thick collagen groups. When RhoA signaling was modulated, EF became more dominant than contact guidance in the thin collagen groups, whereas contact guidance was still more potent in the thick collagen groups. Our data suggested that, contact guidance mediates cell RhoA activating in the thin collagen groups but not in the thick groups. This also implicates that cell use different signaling pathways to have contact guidance when they were on different collagen gel thickness.