脱细胞真皮基质的改建及其作为软骨细胞移植载体在兔软骨缺损修复中的应用

Using Acellular Dermal Matrix for Chondrocyte Implantation in Repairing Cartilage of Rabbit

目的:探讨脱细胞真皮(acellular dermal matrix,ADM)经改建后作为软骨细胞移植载体的可行性。方法:取新生小牛背部真皮组织,分别用自制脱细胞剂和表面活性剂对其进行脱细胞处理后,观察得出自制脱细胞剂脱细胞的效果良好,但脱细胞后的胶原纤维结构需要改建;将自制脱细胞剂脱细胞后的真皮组织分别用按一定浓度配比的消化液和胃蛋白酶消化,发现经配比消化液改建后的胶原纤维表面光滑、孔隙均匀,符合软骨细胞移植载体的要求;比较戊二醛和水溶性交联剂对改建后脱细胞真皮基质的交联效果,可见水溶性交联剂交联后的ADM载体植入大鼠皮下后,其生物相容性良好;用生长因子对改建和交联后的ADM进行纤维表面修饰,然后接种乳兔软骨细胞,观察细胞粘附及生长情况;最后将接种有兔自体软骨细胞的ADM植入兔胫骨软骨缺损部位,术后定期取材观察修复效果。结果:(1)经戊二醛交联后的ADM植入大鼠皮下后有强烈的炎症反应,并有组织出血坏死;而经水溶性交联剂交联的ADM的组织相容性较好。(2)在植入接种有自体软骨细胞的ADM24周后,大白兔胫骨的软骨缺损修复完好,附和的细胞能够存活且增殖,ADM本身基本降解。结论:经脱细胞、纤维改建、交联及生长因子修饰的ADM孔隙均匀,组织相容性好,适于细胞粘附及长期生长。ADM胶原支架在兔体内可基本降解,未见排异反应,移植24周后骨缺损修复。

Objective To study the feasibility of acellular dermal matrix （ADM） as a scaffold for chondrocyte implantation. Methods Calf full-thickness back skin was incubated with cell free buffer or ABS/AES to determine their efficacy for decellularization. After harvesting a cell free matrix, ADM was then digested with digestive buffer or pepsin for comparing surface modification. Both digested ADMs formed type I/Ⅲ collagen scaffold. Inflammatory reaction after implantation with both ADMs was compared and the histology of repaired cartilage surface was determined after 4, 8, 12 and 24 weeks of implantation. Results The ADM cross-linked by glutaralde- hyde demonstrated an obvious inflammatory reaction. Conversely, ADM treated with cross-link solution caused no inflammation after 12 weeks in the rat; if further treated with growth factor, significant improvement in cell attachment and growth appeared as compared with untreated controls. It was found that the cartilage defects were repaired completely and the graft exhibited significant biocompatibility after 24-week of surgery. Conclusions The ADM decellularized with cell free buffer, digested with digestive buffer, cross-linked by cross-link solution, and further decorated with growth factor was suitable for cell attachment and growth, exhibiting significant biocompatibility.