壳聚糖胶原海绵的抗菌止血性能  

Antibacterial and hemostatic properties of chitosan collagen sponge

作　　者：朱虹 林子恒 何柔烨 潘锦滨 刘小川 何晓玲 张静莹 Zhu Hong;Lin Ziheng;He Rouye;Pan Jinbin;Liu Xiaochuan;He Xiaoling;Zhang Jingying(Guangdong Medical University,First Affiliated Hospital of Guangdong Medical University,Key Laboratory of 3D Printing of Stomatology,Dongguan 523710,Guangdong Province,China)

机构地区：[1]广东医科大学,广东医科大学附属东莞第一医院,口腔医学3D打印技术重点实验室,广东省东莞市523710

出　　处：《中国组织工程研究》2023年第16期2525-2533,共9页Chinese Journal of Tissue Engineering Research

基　　金：2021年广东医科大学大学生创新实验项目立项(FCZF002),项目负责人:朱虹;2021年广东省大学生创新创业训练计划项目立项(GDMU2021106),项目负责人:何柔烨;广东省基础与应用基础研究基金联合基金(2020B1515120001),项目负责人:张静莹;广东省普通高校重点领域(2020ZDZX2013),项目负责人:张静莹;广东医科大学学科建设项目(4SG21015G,4SG21019G),项目负责人:张静莹。

摘　　要：背景:胶原因具有良好的生物相容性和降解性能,常用于促进伤口愈合,但由于其不具备抗菌性能,不能预防伤口感染。目的:通过冷冻干燥法制备壳聚糖胶原海绵,使其兼具良好的抗菌及止血效果。方法:壳聚糖溶液与碳酸氢钠溶液在37℃下成凝胶态,利用冷冻干燥法制备壳聚糖/碳酸氢钠/Ⅰ型胶原海绵作为实验组,以壳聚糖海绵及壳聚糖/碳酸氢钠海绵为对照组,扫描电镜观测海绵的微观形貌,测定其pH值、孔隙率、吸水倍数、降解率及对金黄色葡萄球菌和大肠杆菌的抗菌率;利用CCK-8法和Live/Dead染色检测海绵浸提液的细胞毒性;使用大鼠截尾出血模型检测海绵的止血效果;建立大鼠皮肤创伤模型,通过苏木精-伊红染色及Masson染色观察海绵植入7 d后的组织愈合情况。结果与结论:(1)相较于其他两种海绵,壳聚糖/碳酸氢钠/Ⅰ型胶原海绵的绵孔隙更大,外壁更粗糙,吸水性更强、孔隙率及吸水倍数更大;而且壳聚糖/碳酸氢钠/Ⅰ型胶原海绵孔隙率均大于另外两种海绵(P<0.01);平均吸水倍数复合海绵大于壳聚糖海绵(P<0.0001)和壳聚糖/碳酸氢钠海绵(P<0.05);(2)抗大肠杆菌率检测结果显示:壳聚糖海绵>壳聚糖/碳酸氢钠海绵(P<0.001)>壳聚糖/碳酸氢钠/Ⅰ型胶原蛋白海绵(P<0.0001);(3)抗金黄色葡萄球菌率检测结果显示:壳聚糖海绵>壳聚糖/碳酸氢钠海绵>壳聚糖/碳酸氢钠/Ⅰ型胶原蛋白海绵(P<0.01);(4)壳聚糖/碳酸氢钠/Ⅰ型胶原蛋白海绵无细胞毒性且可促小鼠成纤维细胞(L929)增殖,促细胞增殖效果优于空白组(P<0.05)、壳聚糖组(P<0.0001)与壳聚糖/碳酸氢钠组(P<0.05);(5)壳聚糖/碳酸氢钠/Ⅰ型胶原蛋白海绵止血效果良好,优于其他三组;(6)壳聚糖/碳酸氢钠/Ⅰ型胶原蛋白海绵植入大鼠背部创口1周,周围皮肤无明显炎症反应,新生毛细血管及胶原含量明显高于其他两种海绵,皮肤组织恢复最佳;(7)上述数据BACKGROUND:Collagen,which has good biocompatibility and degradability,is commonly used to promote wound healing,but it is not effective in preventing wound infection because of its non-antibacterial properties.OBJECTIVE:To prepare chitosan collagen sponge by freeze-drying method,which has good antibacte rial and hemostatic effects.METHODS:Chitosan solution and sodium bicarbonate solution became gel at 37℃.Chitosan/sodium bicarbonate/typeⅠcollagen composite sponge was prepared by freeze-drying method as the experimental group,and chitosan sponge and chitosan/sodium bicarbonate sponge were used as the control group.Scanning electron microscopy was used to observe the micro-morphology of the sponge and determine its pH valu e,porosity,water absorption,degradation rate and antimicrobial activity against Staphylococcus aureus and Escherichia coli.The cytotoxicity of the sponges was detected by CCK-8 assay and Live/Dead staining.The hemostatic effect of the sponges was detected by the rat model of amputation bleeding.The rat model of skin trauma was established.The tissue healing was observed by hematoxylin-eosin staining and Masson staining 7 days after sponge implantation.RESULTS AND CONCLUSION:(1)Compared with the other sponge groups,the chitosan/sodium bicarbonate/typeⅠcollagen composite sponge had larger porosity,rougher outer wall,stro nger water absorption,greater porosity and water absorption multiple,and the porosity was greater than the other sponge groups(P<0.01).The ave rage water absorption multiple was greater than chitosan sponge(P<0.0001)and chitosan/sodium bicarbonate sponge(P<0.05).(2)Anti-Escherichia coli test res ults showed that Escherichia coli resistance rate was chitosan sponge>chitosan/sodium bicarbonate sponge(P<0.001)>chitosan/sodium bicarbonate/typeⅠcollagen composite sponge(P<0.0001).(3)Anti-Stophylococcus oureus rate test results showed that anti-Stophylococcus aureus rate was chitosan sponge>chitosan/sodium bicarbonate sponge>chitosan/sodium bicarbonate/typeⅠcollagen composite s

关 键 词：壳聚糖 Ⅰ型胶原 碳酸氢钠 止血材料 抗菌 组织修复 止血海绵 伤口愈合 

分 类 号：R459.9[医药卫生—治疗学] R363[医药卫生—临床医学] R364