壳聚糖-胶原基可注射水凝胶制备及生物相容性检测  

Investigation of preparation and biocompatibility of injectable chitosan-collagen-based hydrogels

作　　者：宋克东[1] 刘天庆[1] 姜波[1] 李瑞鹏[1] 张文[1] 姜丽丽[1] 马学虎[1] 崔占峰[2] 

机构地区：[1]大连理工大学大连市干细胞与组织工程研发中心,辽宁大连116024 [2]牛津大学工程科学系,英国牛津OX1 3PJ

出　　处：《大连理工大学学报》2011年第5期644-652,共9页Journal of Dalian University of Technology

基　　金：国家自然科学基金资助项目(30670525,30700181);教育部高等学校博士学科点新教师基金资助项目(20070141055);大连理工大学青年教师培养基金资助项目(893228)

摘　　要：制备了可注射的新型温敏性壳聚糖-甘油磷酸钠-胶原（chitosan-glycerophosphate-collagen，C-GP-Co）水凝胶并检测了其与脂肪间充质干细胞（adipose tissue-derived stem cells，ADSCs）的生物相容性．首先将体积分数2．2％的壳聚糖溶液与质量分数50％的B．甘油磷酸钠溶液按一定比例混合后，将2mg／mL胶原溶液加入上述溶液中混合得到壳聚糖-甘油磷酸钠-胶原水凝胶；随后用扫描电镜观察其微观结构，测定了37℃时的成胶时间并测量水凝胶中提取培养基的渗透压；最后将分离、培养和免疫表型鉴定后的ADSCs接种在C-GP—Co水凝胶中培养，倒置显微镜下观察细胞的生长形态，并用Live／Dead Viability-Cytotoxieity试剂盒染色观察细胞凋亡状况，CCK-8试剂盒测定细胞增殖情况．结果表明C—GP—Co水凝胶37℃的成胶时间为12min，水凝胶内部呈海绵状多孔结构，水凝胶中提取培养基的渗透压为310-330mmol／kg．培养7d后，C-GP-Co水凝胶表面的ADSCs贴壁生长，并显示出良好的细胞活性．上述结果表明制备的C-GP-Co水凝胶具有良好的细胞相容性，可作为ADSCs生长增殖的良好支架．An injectable new-type temperature-sensitive hydrogel consisted of chitosanglycerophosphate-collagen （C-GP-Co） was prepared and the biocompability for adipose tissue-derived stem cells （ADSCs） was detected. The C-GP-Co hydrogel was prepared as follows.chitosan solution （2. 2%, volume fraction） and β-glycerophosphate solution （50%, mass fraction） were mixed at a series of proportion, and then, the collagen solution （2 mg/mL）was added into the previous mixture to get the C-GP-Co hydrogel. The gelation time was measured at 37 ℃, and the osmotic pressure of the media within the hydrogels was measured, and then, the micro structure analysis was carried out by SEM. The isolated, cultured and identified ADSCs were seeded into C-GP-Co hydrogels and cell growth status was observed under inverted microscope. The celt apoptosis was detected using Live/ Dead Viability-Cytotoxicity Kit, while the proliferation of ADSCs was determined by CCK-8 kits. The experimental results show that the gelation time is about 12 min at 37℃, the osmotic pressure of media in the hydrogel is in the normal range of 310-330 mmol/kg and the inner structure of hydrogel is spongy porous. And later, the biocompability of the C-GP-Co hydrogel was further examined. After 7 days＇ culture, cells within C-GP-Co hydrogels display adherent cell morphology and high cell viability. It is demonstrated that the C-GP-Co hydrogel has excellent cell compatibility and is a proper scaffold for ADSCs proliferation.

关 键 词：水凝胶 壳聚糖 胶原 脂肪间充质干细胞 组织工程 

分 类 号：Q813.1[生物学—生物工程] R318[医药卫生—生物医学工程]