透明质酸-MMP敏感肽仿生杂化水凝胶的制备及其对骨髓间充质干细胞分化调控的影响

摘要

设计合成了3种不同序列的基质金属蛋白酶(MMP)敏感肽,与活性基团马来酰亚胺修饰后的透明质酸进行共价交联,制备一类对MMP具有响应性的透明质酸仿生杂化水凝胶,并研究其对诱导兔骨髓间充质干细胞(BMSCs)向软骨方向分化的影响.以 N-羟基硫代琥珀酰亚胺(Sulfo-NHS)与1-(3-二甲氨基丙基)-3-乙基碳二亚胺盐酸盐(EDC?HCl)为催化剂,活化透明质酸羧基后,与 N-(2-氨基乙基)马来酰亚胺三氟乙酸盐(AEM)常温搅拌反应,得到接枝马来酰亚胺基团的透明质酸(HA-MAL),马来酰亚胺基团与MMP敏感肽上的巯基在常温下即可发生加成反应.共价交联MMP敏感肽的水凝胶具有良好的机械性能,能够包埋细胞3 D培养达4周以上.对水凝胶3D及孔板2D培养的BMSCs进行实时荧光定量PCR测定软骨方向分化的特征性基因Marker表达,结果显示,3种水凝胶对 COL1A2、COL2A1、ACAN、SOX94种基因表达具有不同程度的上调作用,可促进BMSCs的软骨分化.%Three different sequences of matrix metalloproteinases (MMP)sensitive peptides were designed and covalently crosslinked with hyaluronic acid which was modified with active group of maleimide to prepare a class of hyaluronic acid bionic hybrid hydrogel and then its effect on the cartilage differentiation of rabbit bone mar-row mesenchymal stem cells (BMSCs)was studied.The hyaluronic acid(HA)was activated with (Sulfo-NHS) and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC·HCl)as catalyst,then N-(2-amin-oethyl)maleimide trifluoroacetate (AEM)was grafted with activated HA at room temperature to obtain HA-MAL.The addition reaction of the maleimide group with the mercapto group on the MMP-sensitive peptide could be carried out at room temperature.Covalently cross-linked MMP-sensitive peptide hydrogel has good me-chanical properties,cells can be embedded in which for more than 4 weeks.The results show that compared with 2D orifice plate cells culture,hydrogels have different effects on the up regulation of COL1A2,COL2A1, ACAN and SOX9 gene expression,which can promote BMSCs to cartilage differentiation.