利用導電高分子聚吡咯為基材以電刺激促進幹細胞分化;Electrical Stimulation to Promote Stem cell Differentiation using Conducting Polypyrrole Films

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Please use this identifier to cite or link to this item: http://ir.lib.ncu.edu.tw/handle/987654321/53751

Title:	利用導電高分子聚吡咯為基材以電刺激促進幹細胞分化;Electrical Stimulation to Promote Stem cell Differentiation using Conducting Polypyrrole Films
Authors:	徐藝庭;Hsu,Yi-ting
Contributors:	化學工程與材料工程研究所
Keywords:	成骨;間質幹細胞;電刺激;聚吡咯膜;化學氧化聚合;Osteogenesis;Mesenchymal stem cells;Electrical stimulation;Polypyrrole;Chemical oxidative polymerization
Date:	2012-08-23
Issue Date:	2012-09-11 18:13:06 (UTC+8)
Publisher:	國立中央大學
Abstract:	聚吡咯(Polypyrrole, PPy)是一種具導電性的生醫材料。在本研究中，我們將PPy膜作為培養間葉幹細胞(mescnchymal stem cells, MSCs)的基材，觀察PPy對於細胞分化成骨的影響。利用化學氧化的方式，以過硫酸銨(ammonium persulfate, APS)作為起始劑可以成功製備出PPy膜，藉由改變單體濃度及與起始劑的莫爾比，可控制所製備的PPy薄膜具有不同導電度。首先以傅立葉紅外光譜(Fourier transform infrared spectroscopy, FTIR)進行確認以PPy膜成功地沉積在基材上，在材料的物性分析中，我們使用四點探針來量測材料片電阻，並以掃描電子顯微鏡(scanning electron microscopy, SEM)和原子力顯微鏡(atomic force microscopy, AFM)觀察薄膜厚度及表面形貌。另外，藉由電子能譜儀(X-ray spectroscopy, XPS)探討薄膜表面元素組成，解析C1s的吸收峰後發現PPy薄膜的導電性質與高分子的分枝率成正比，推測合成高交聯度的聚吡咯可以提升高分子網絡結構的完整性。最後我們將PPy膜用於培養大鼠的骨髓間葉幹細胞(RM1)， MTT分析結果表明PPy膜是生物相容性材料，並且以骨分化培養液進行細胞培養，將RM1細胞誘導分化為成骨細胞。根據鹼性磷酸酶(alkaline phosphatase activity assay, ALP)的活性檢測顯示，PPy膜應具有提早分化成骨的效果。以茜素紅染色(Alizarin red stain, ARS) 及Calcium-o- cresolphthalein complexone的分析結果表明，聚吡咯薄膜能促進骨的礦化，以增加細胞外基質中的鈣沉積。最後，藉由PPy膜上施加恆定電場，探討電刺激成骨作用，結果顯示經過電刺激培養的細胞比起未經處理的PPy膜，其鈣沉積明顯增加，說明電刺激可能是能夠增進細胞骨化的效果。這些結果表明，聚吡咯具有優越的生物傳導性，適合作為骨組織修復用的生醫材料。Polypyrrole (PPy) is a conductive biomaterial. In this study we would like to apply PPy films as substrates to culture mesenchymal stem cells (MSCs) to evaluate if PPy may promote osteogenesis. Chemical oxidation polymerization using ammonium persulfate (APS) as initiator was utilized to polymerize PPy films. By changing monomer concentrations and initiator ratio, films with different conductivity were fabricated. Fourier transform infrared spectroscopy (FTIR) was performed to confirm that PPy films were successfully deposited. Physical properties of PPy films were characterized using four point probe, scanning electron microscopy (SEM) and atomic force microscopy (AFM) to demonstrate sheet resistance, thickness, and surface morphology of PPy films, respectively. In addition, X-ray spectroscopy (XPS) was applied to investigate film composition. By diagnosing C1s peaks, PPy films with higher conductivities always had higher branch ratio, which should be due to that complete network would be formed by highly-crosslinked PPy chains.These films were then applied for culturing rat bone marrow-derived mesenchymal stem cells (RM1). MTT assay suggested that PPy films were biocompatible materials. By using osteogenic medium, RM1 cells were differentiated into osteoblasts. The alkaline phosphatase (ALP) activity assay revealed that PPy films may accelerate osteogenesis. Alizarin red stain (ARS) and calcium-o-cresolphthalein complexone assays suggested that PPy film promoted mineralization to increase calcium deposition in extracellular matrix.Finally, PPy films were subjected in a constant electric field to elucidate the effect of electrical stimulation on osteogenesis. Cells cultured on PPy films with electrical stimulation deposited more calcium than that cultured on untreated PPy films, suggesting that electrical stimulation may be able to improve cell differentiation. These results indicated that PPy should be an appropriate biomaterial with superior bioconductivity, which should be capableIIIof facilitating bone regeneration in tissue engineering application.
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