高效去除异体组织中脱细胞试剂十二烷基硫酸钠残留的清洗方法  

Efficient removal of a decellularized reagent sodium dodecyl sulfate in allogeneic tissue

作　　者：胡凯 李淼 邵怡然 汪晶晶 陈金发 Hu Kai;Li Miao;Shao Yiran;Wang Jingjing;Chen Jinfa(Shanghai Gencong Biomedical Materials Research Center,Shanghai 201201,China;Shanghai Yapeng Biotechnology Co.,Ltd.,Shanghai 201201,China)

机构地区：[1]上海亘从生物医用材料研究中心,上海市201201 [2]上海亚朋生物技术有限公司,上海市201201

出　　处：《中国组织工程研究》2023年第15期2311-2317,共7页Chinese Journal of Tissue Engineering Research

摘　　要：背景:十二烷基硫酸钠是一种常用于同种异体组织脱细胞处理的脱细胞试剂,然而其难以从组织内去除,少量残留即可产生细胞毒性。目的:提出一种高效去除异体组织中脱细胞试剂十二烷基硫酸钠的清洗方法。方法:选择纯水、钾盐溶液和乙醇作为清洗介质,并通过不同介质的排列组合进行清洗,对十二烷基硫酸钠残留量、异体真皮和异体肌腱的组成、结构、力学性能和细胞毒性等方面进行检测,评价清洗效果。结果与结论:总体上,乙醇+氯化钾、体积分数75%乙醇+氯化钾对样品的成分和结构无大影响。首先,十二烷基硫酸钠浸泡处理将异体真皮和异体肌腱的DNA残留控制在50 ng/mg以下之后再进行清洗,异体真皮、异体肌腱中的十二烷基硫酸钠含量分别降低了96.7%和97%;其次,异体真皮、异体肌腱的糖胺聚糖和胶原蛋白含量均无较大变化;此外,异体肌腱在清洗后,其最大拉力、强度和模量高于脱细胞后,与原材料相比无显著性差异;最后,细胞毒性结果显示异体真皮和异体肌腱清洗后的细胞存活率高于80%,细胞毒性分级不大于1级。BACKGROUND:Sodium dodecyl sulfate is commonly used for decellularization of allogeneic matrix,whereas it is hard to remove from matrix.Only a small amount of sodium dodecyl sulfate in matrix can cause cytotoxicity.OBJECTIVE:To provide a high effective method to remove sodium dodecyl sulfate,which residuals in allogeneic matrix.METHODS:The pure water,potassium salt solution and ethanol were chosen to remove sodium dodecyl sulfate.The decellularization degree,residual amount of sodium dodecyl sulfate,composition,structure and tensile properties and cytotoxicity of allogeneic dermis and allogeneic tendon were detected to evaluate the cleaning effect.RESULTS AND CONCLUSION:In total,ethanol+potassium chloride and 75%ethanol+potassium chloride produced little significant impacts on the component and structure of allogeneic matrix.Firstly,on condition that sodium dodecyl sulfate was infiltrated into allogeneic matrix since DNA amount was controlled under 50 ng/mg(dry weight),the sodium dodecyl sulfate residual results showed a tremendous reduction of 96.7%and 97%in allogeneic dermis and allogeneic tendon,respectively.Secondly,no significant changes occurred in allogeneic dermis and allogeneic tendon for glycosaminoglycan and collagen.Moreover,after carefully washing,the allogeneic tendon exhibited higher the maximum tensile force,strength,and modulus than those after decellularization.Meanwhile,there was no significant difference compared with the raw material.Finally,the cytotoxicity test showed that cell viability was higher than 80%after washing of allogeneic dermis and allogeneic tendon;the cytotoxicity grade was not greater than lever 1.

关 键 词：脱细胞 十二烷基硫酸钠 同种异体组织 异体真皮 异体肌腱 

分 类 号：R459.9[医药卫生—治疗学] R318.08[医药卫生—临床医学] R34