幾丁聚醣膜接上不同蛋白對細胞的影響

Effect of albumin and gelatin coating on chitosan scaffold on cells

李維(Wei Lee)

指導教授 ： 謝瑞香

中原大學/工學院/生物醫學工程學系/碩士(2008年)

https://doi.org/10.6840/cycu200900148

摘要

本實驗的目的為探討表面鍵結不同蛋白質的幾丁聚醣薄膜對細胞生長之影響，並且利用紅光刺激，觀察經過光刺激後細胞是否有進一步的增生現象。本實驗主要分成三個部分：（1）幾丁聚醣薄膜表面鍵結不同蛋白質對細胞生長之影響；（2）幾丁聚醣薄膜造孔與否對細胞生長之影響；（3）光刺激對細胞生長之影響。蛋白質的部分採用明膠以及牛血清白蛋白；細胞選用的是人類包皮纖維母細胞以及人類上皮角質細胞；造孔方式為添加1.8％的PEG 8000；化學鍵結的方式採用添加0.25％的戊二醛做為交聯劑；光刺激的參數為630 nm之紅光波段，照射能量為16.3 mW、照射時間為400秒；細胞計數方式採用MTT assay。
實驗結果顯示，纖維母細胞培養在表面鍵結不同蛋白質的幾丁聚醣薄膜上，的確會造成貼附性以及增生速率的改變，培養在鍵結明膠的幾丁聚醣薄膜上貼附性以及增生速率都有明顯的增加，培養在鍵結牛血清白蛋白的幾丁聚醣薄膜上生長速率則略為抑制；在造孔與否方面，孔徑大小約為20μm的幾丁聚醣支架似乎不會造成纖維母細胞的增生；而利用紅光刺激則可明顯加速纖維母細胞的增生速率，不論是培養在幾丁聚醣表面、鍵結明膠的幾丁聚醣表面，或是鍵結牛血清白蛋白的幾丁聚醣表面。
角質細胞的部分，實驗結果則發現培養在表面鍵結明膠的幾丁聚醣薄膜上不會對細胞的貼附性以及生長速率造成影響，而培養在鍵結牛血清白蛋白的幾丁聚醣薄膜上則發現貼附性以及生長速率都有略微增加的現象；而孔洞大小20μm的幾丁聚醣支架，則不會造成角質細胞的增生；紅光刺激的部分，不論是培養在幾丁聚醣表面、鍵結明膠的幾丁聚醣表面、或是鍵結牛血清白蛋白的幾丁聚醣表面，經過光刺激後，都無法促進角質細胞的進一步的增生。
由上述結果可得知，表面鍵結明膠的幾丁聚醣薄膜，可以促進纖維母細胞的貼附與生長；而表面鍵結牛血清白蛋白的幾丁聚醣薄膜，可以促進角質細胞的增生；利用630 nm的LED光刺激，則可進一步的加速纖維母細胞增生。各項結果都顯示，表面鍵結不同蛋白的幾丁聚醣薄膜，在臨床上可做為幫助表皮組織重建的良好基材。

關鍵字

表面改質 ； 光刺激 ； 幾丁聚醣

並列摘要

The objective of this study is to discuss the effect of proliferation of cells on chitosan membrane with different protein surface coating. More than that, we use red light irradiation to stimulate cells and observe whether cells will further proliferation or not. This study separate into three parts: First, the effect of coating gelatin and BSA on chitosan membrane on cells. Second, cell proliferation on chitosan membrane or scaffold. Third, the effect of light irradiation on cells on chitosan membrane. We choose gelatin and BSA, use 0.25% glutaraldehyde as crosslinker. The cell types are human foreskin fibroblast and human epidermal keratinocyte. Light irradiation detail is 630 nm, 16.3 mW, irradiate 400 seconds. We use MTT assay to count cells.
As data shows, coating protein on chitosan membrane will cause fibroblast proliferation rate change: surface coating gelatin will cause more proliferation, and surface coating BSA will cause restrain. On film/scaffold test, combine PEG as a porogen to form a scaffold will not excite fibroblast proliferation. Using red light irradiation will clearly make fibroblast proliferate, no matter culture cells on chitosan membrane, chitosan membrane with gelatin surface coating, or on chitosan membrane with BSA surface coating.
When coating gelatin on chitosan membrane, keratinocyte proliferation rate doesn’t change, but when cells culture on chitosan membrane with BSA surface coating, it will cause more proliferation. On film/scaffold test, we can find out that combine PEG as a porogen to form a scaffold will not excite kerationcyte proliferation. Using red light irradiation will not cause keratinocyte further proliferation.
So we can make a conclusion, surface coating gelatin on chitosan membrane can promote fibroblast proliferation, surface coating BSA on chitosan membrane can promote keratinocyte proliferation, using 630 nm LED irradiation can further promote fibroblast proliferation. All data show that surface coating gelatin or BSA on chitosan membrane will be a good model to assist epidermal reconstruction.

並列關鍵字

surface modification ； light irradiation ； chitosan

參考文獻

謝岳峰（民94）。光刺激對纖維母細胞及角質細胞的影響。私立中原大學生物醫學工程系碩士論文，中壢市。

葉國全（民95）。紅光刺激對支架中細胞生長之影響。私立中原大學生物醫學工程系碩士論文，中壢市。

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