自体BMSCs复合改建脱细胞真皮基质体内构建组织工程软骨的实验研究Bone marrow stromal cells seeded in acellular dermal matrix scaffolds for cartilage regeneration
时长江;王鹏飞;辛大森;姜文学;
摘要(Abstract):
[目的]目的:探讨BMSCs复合脱细胞真皮基质(ADM)三维多孔支架体内构建组织工程软骨的可行性。[方法]首先对支架材料进行大体及扫描电镜观察,分别测定支架的孔隙直径、孔隙率、降解率;其次,分离培养兔骨髓基质细胞(BMSCs),采用扫描电镜及MTT法检测BMSCs在支架材料上的生长、增殖情况;最后取3个月龄清洁级新西兰大白兔27只,雌性,体质量(2.4±0.2)kg,随机分为3组,即A组:空白对照组(仅髁部造缺损),B组:单纯ADM覆盖,C组:ADM+BMSCs,细胞支架复合物体外培养1周后植入兔股骨髁软骨缺损处,分别在4、8、12周后对标本进行组织学、免疫化学检测及ICRS评分。[结果]测量示支架孔隙直径为(101.2±9.3)um,孔隙率为88.6%±2.7%,降解率2周为(13.83±7.12)%,4周为(25.66±8.19)%。MTT法显示细胞在支架上生长良好,与对照组相比,吸光度值差异无统计学意义(P>0.05),提示支架相容性良好,无细胞毒性。经过最长12周培养后取材进行组织学、免疫化学检测,结果显示ADM+BMSCs组软骨缺损获得了更为满意的修复效果:关节表面平坦光滑,与周围组织未见明显界限;组织学染色证实软骨基质分泌旺盛并有典型软骨细胞形成,组织结构类似正常关节软骨,ADM+BMSCs组术后第4、8、12周ICRS评分均高于其他两组,差异具有统计学意义(P<0.05)。[结论]ADM孔隙均匀,组织相容性好,适于细胞粘附及长期生长,是一种适用于软骨组织工程的良好载体。BMSCs-ADM支架复合体在体内成功构建组织工程软骨,可进一步用于软骨组织工程研究。
关键词(KeyWords): 脱细胞真皮基质;BMSCs;软骨修复;组织工程
基金项目(Foundation): 天津市卫生局科技基金重点攻关课题(编号:11KG109)
作者(Author): 时长江;王鹏飞;辛大森;姜文学;
Email:
DOI:
参考文献(References):
- [1]Hunziker EB.Articular cartilage repair:basic science and clinical progress.A review of the current status and prospects[J].Osteoarthritis Cartilage,2002,6:432-463.
- [2]于院龙,胡蕴玉,毕龙,等.脱细胞软骨基质三维支架的制备及特性研究[J].中国矫形外科杂志,2010,9:743-747.
- [3]王玉,彭江,张莉,等.软骨细胞外基质/壳聚糖复合多孔支架和骨髓间充质干细胞构建组织工程软骨[J].中国矫形外科杂志,2010,20:1715-1718.
- [4]Wu X,Liu Y,Li X,et al.Preparation of aligned porous gelatin scaf olds by unidirectional freeze-drying method[J].Acta Biomater,2010,3:1167-1177.
- [5]Yan LP,Wang YJ,Ren L,et al.Genipin-cross-linked collagen/chitosan biomimetic scaffolds for articular cartilage tissue engineering applications[J].J Biomed Mater Res A,2010,2:465-475.
- [6]Cao L,Yang F,Liu G,et al.The promotion of cartilage defect repair using adenovirus mediated Sox9 gene transfer of rabbit bone marrow mesenchymal stem cells[J].Biomaterials,2011,16:3910-3920.
- [7]van den Borne MP,Raijmakers NJ,Vanlauwe J,et al.International cartilage repair society(ICRS)and Oswestry macroscopic cartilage evaluation scores validated for use in autologous chondrocyte implantation and microfracture[J].Osteoarthritis Cartilage,2007,15:1397-1402.
- [8]Mainil-Varlet P,Van Damme B,Nesic D,et al.A new histology scoring system for the assessment of the quality of human cartilage repair:ICRSⅡ[J].Am J Sports Med,2010,38:880-890.
- [9]Urita Y,Komuro H,Chen G,et al.Regeneration of the esophagus using gastric acellular matrix:an experimental study in a rate model[J].Pediatr Surg Int,2007,1:21-26.
- [10]Kim YJ,Kim HJ,Im GI.PTHrP promotes chondrogenesis and suppresses hypertrophy from both bone marrow-derived and adipose tissue-derived MSCs[J].Biochem Biophys Res Commun,2008,1:104-108.
- [11]Martin I,Shastri VP,Padera RF,et al.Selective differentiation ofmammalian bone marrow stromal cells cultured on three-dimensional polymer foams[J].J Biomed Mater Res,2001,2:229-235.
- [12]Martinez-Alvernia EA,Rudnick JA,Mankarious LA.Transforming growth factor beta3 increases chondrocyte proliferation and decreases apoptosis in murine cricoid cartilage in vitro[J].Otolaryngol Head Neck Surg,2008,4:435-440.
- [13]Wang W,Li B,Li Y,et al.In vivo restoration of full-thickness cartilage defects by poly(lactide-co-glycolide)sponges filled with fibrin gel,bone marrow mesenchymal stem cells and DNA complexes[J].Biomaterials,2010,23:5953-5965.
- [14]Tay LX,Ahmad RE,Dashtdar H,et al.Treatment outcomes of alginate-embedded allogenic mesenchymal stem cells versus autologous chondrocytes for the repair of focal articular cartilage defects in a rabbit model[J].Am J Sports Med,2011,1:83-90.
- [15]Muraglia A,Martin I,Cancedda R,et al.A nude mouse model for human bone formation in unloaded conditions[J].Bone,1998,5:131-134.
- [16]Dai W,Kawazoe N,Lin X,et al.The influence of structural design of PLGA/collagen hybrid scaffolds in cartilage tissue engineering[J].Biomaterials,2010,8:2141-2152.
- [17]Nettles DL,Elder SH,Gilbert JA.Potential use of chitosan as a cell scaffold material for cartilage tissue engineering[J].Tissue Eng,2002,6:1009-1016.