可注射性乙二醇壳聚糖/双醛功能化聚乙二醇水凝胶的细胞相容性  

Cytocompatibility of injectable glycol chitosan/dibenzaldehyde-terminated poly-ethyleneglycol hydrogel

作　　者：荆晓光[1,2] 刘舒云[2] 郭维民[2] 李旭[2] 吕奥 刘士臣[1] 孟昊业[2] 陈明学[2] 张学亮[2] 张增增 刘雪剑[1,2] 高超[1,2] 王泽浩 张彬[2] 沈师[2] 陶磊 杨建华[1,4] 郭全义 Jing Xiaoguang;Liu Shuyun;Guo Weimin;Li Xu;Lü Ao;Liu Shichen;Meng Haoye;Chen Mingxue;Zhang Xueliang;Zhang Zengzeng;Liu Xuejian;Gao Chao;Wang Zehao;Zhang Bin;Shen Shi;Tao Lei;Yang Jianhua;Guo Quanyi(Jiamusi University,Jiamusi 154007,Heilongjiang Province,China;Institute of Orthopedics,Chinese PLA General Hospital,Beijing 100853,China;Department of Chemistry,Tsinghua University,Beijing 100084,China;Longguang People’s Hospital,Shenzhen 518172,Guangdong Province,China)

机构地区：[1]佳木斯大学,黑龙江省佳木斯市154007 [2]解放军总医院骨科研究所,北京市100853 [3]清华大学化学系,北京市100084 [4]深圳市龙岗区人民医院,广东省深圳市518172

出　　处：《中国组织工程研究》2019年第2期196-203,共8页Chinese Journal of Tissue Engineering Research

基　　金：国家重点研发计划课题(2017YFC1104102;2017YFC1103404);国家自然科学基金(21134004;81472092;项目负责人:郭全义;81772319;项目负责人:郭全义);北京市自然科学基金(7172203;项目负责人:郭全义);北京市科技专项(Z161100005016059)~~

摘　　要：背景:课题组前期研究自主研发了可注射性乙二醇壳聚糖/双醛功能化聚乙二醇水凝胶(glycolchitosan/dibenzaldehyde-terminated poly-ethyleneglycol,GCS/DF-PEG),研究显示其具有较好的可注射性和自愈性。目的:进一步检测可注射性GCS/DF-PEG的物理学性质及细胞相容性。方法:将质量分数1.5%的乙二醇壳聚糖溶液与质量分数分别为2%,4%,8%的双醛功能化聚乙二醇溶液等体积混合,制备3组可注射性GCS/DF-PEG水凝胶,检测其弹性模量。将3组可注射性水凝胶分别浸入PBS中4周,检测凝胶的体外降解。提取第2代SD乳鼠脂肪间充质干细胞,实验组加入可注射性GCS/DF-PEG水凝胶浸提液,对照组常规培养,MTT法检测细胞增殖。将质量分数3%的乙二醇壳聚糖溶液与第2代SD乳鼠脂肪间充质干细胞混合,再与质量分数为4%的双醛功能化聚乙二醇溶液混合,培养第1,5天,采用死活染色法检测细胞在水凝胶内死活状态。将质量分数3%的乙二醇壳聚糖溶液与第2代SD乳鼠脂肪间充质干细胞混合,再与质量分数分别为2%,4%,8%的双醛功能化聚乙二醇溶液混合,MTT法检测细胞增殖。结果与结论:(1)质量分数分别为2%,4%,8%双醛功能化聚乙二醇组水凝胶的弹性模量分别为13.48,22.21,33.19kPa;(2)随时间的延长,3组水凝胶的降解率均逐渐增加,质量分数2%双醛功能化聚乙二醇组水凝胶的降解速率明显快于其他两组;(3)培养7d内,实验组细胞增殖与对照组比较无差异;(4)死活染色显示,脂肪间充质干细胞在水凝胶内呈球形,存活率在90%以上,且随时间延长细胞数量明显增多;(5)随着时间的延长,脂肪间充质干细胞在含不同质量分数双醛功能化聚乙二醇GCS/DF-PEG水凝胶内的数量逐渐增多,且含质量分数2%双醛功能化聚乙二醇组水凝胶内的细胞增殖快于含质量分数4%,8%双醛功能化聚乙二醇组(P <0.05);(6)结果表明,可注射性GCS/DF-PEG水凝胶的细胞相容性良好,力学BACKGROUND:Our research group independently developed an injectable glycol chitosan/dibenzaldehyde-terminated poly-ethyleneglycol(GCS/DF-PEG)hydrogel,which has good injectability and self-healing properties.OBJECTIVE:To test the physical properties and cytocompatibility of the GCS/DF-PEG hydrogel.METHODS:The injectable GCS/DF-PEG hydrogel was prepared by mixing GCS solution at a mass fraction of 1.5%with an equal volume of DF-PEG solution at a mass fraction of 2%,4%,and 8%,respectively.Their moduli of elasticity were measured.Three groups of injectable hydrogels were immersed in PBS for 4 weeks to detect the in vitro degradation of the hydrogels.Passage 2 adipose-derived mesenchymal stem cells from Sprague-Dawley neonatal rats were cultured in injectable GCS/DF-PEG hydrogel leaching solution as experimental group or cultured routinely as control group.MTT assay was used to detect the cell proliferation.Passage 2 adipose-derived mesenchymal stem cells from Sprague-Dawley neonatal rats were mixed with 3%GCS solution,and then mixed with 4%DF-PEG solution.On the 1st and 5th days of culture,the cell survival and death in the hydrogel were tested by live/dead staining.Passage 2 adipose-derived mesenchymal stem cells from Sprague-Dawley neonatal rats were mixed with 3%GCS solution,and then mixed with 2%,4%and 8%DF-PEG solution,respectively.MTT method was used for testing the cell proliferation.RESULTS AND CONCLUSION:(1)The moduli of elasticity of GCS/DF-PEG hydrogel with 2%,4%,8%DF-PEG were 13.48,22.21 and 33.19 kPa,respectively.(2)In vitro degradation experiments showed that GCS/DF-PEG hydrogels gradually degraded in PBS over time.And the degradation rate of the 2%DF-PEG hydrogel was significantly faster than the other two groups.(3)Within 7 days of culture,there was no difference in the cell proliferation between the experimental and control groups.(4)Live/dead staining results showed that adipose-derived mesenchymal stem cells were spherical in the hydrogel,the survival rate was over 90%,and the number of cells incre

关 键 词：可注射水凝胶 脂肪间充质干细胞 扫描电镜 降解实验 细胞增殖 细胞死活染色 国家重点研发计划课题 生物材料 水凝胶 材料试验 细胞增殖 组织工程 

分 类 号：R459.9[医药卫生—治疗学] R318[医药卫生—临床医学]