大鼠盆底重建组织工程补片的初步构建及其在体研究

Primary fabrication of a tissue-engineered mesh for pelvic floor reconstruction and research in vivo

目的 初步探讨自体来源脂肪间充质干细胞（ADSC）复合丝素蛋白支架构建盆底重建组织工程补片的可行性.方法 分离培养大鼠ADSC,扩增后种植于丝素蛋白溶液与丝素纤维网片冻干而成的支架上,制备组织工程补片,并观察形态.将组织工程补片植入相应大鼠皮下,对照组植入丝素蛋白支架.观察其在体组织化、力学性能及胶原形成情况.结果 ADSC体外增殖良好,丝素蛋白支架由编织部分与海绵部分组成,ADSC可黏附于支架并形成细胞层.组织工程补片在体生物相容性好,最大失效力（3.081 N±0.121 N比2.167 N±0.148 N）、杨氏模量（3.191 MPa±0.146 MPa比2.263 MPa±0.213 MPa）及胶原含量（18.648 μg/mg±0.867 μg/mg比14.123 μg/mg±0.989μg/mg）均优于对照组（P＜0.05）.结论 组织工程补片可满足盆底重建要求,具有良好应用前景.

Objective To explore the fabrication of a tissue-engineered mesh fabricated with autologous adipose-derived mesenchymal stem cells （ADSC） and silk fibroin scaffold for female pelvic reconstruction.Methods Sprague-Dawley rat adipose tissue was obtained from inguinal region.Mesenchymal stem cells were isolated and proliferated in vitro.The sericin of silk was removed after knitting in to web.Then a compound scaffold freeze-dried of silk fibroin web and silk fibroin solution was prepared for ADSC seeding.after 1-week culturing,the cellular morphology and extracellular matrix secretion on scaffold were observed by scanning electronic microscopy.The tissue-engineered meshes were implanted subcutaneously into the homologous rats while silk fibroin scaffolds served as control.Then the biomechanics,collagen matrix and local tissue response to tissue-engineered meshes in vivo with that to silk fibroin scaffolds were estimated.Results ADSC were fibroblast-like and proliferated well in vitro.SEM showed that the scaffold had an interwoven structure with a smooth surface and the silk sponges in scaffold had a uniform and porous structure.ADSC adhered firmly to the scaffold,secreted extracellular matrix and formed cell sheets on scaffold.In vivo studies showed that the tissue matrix of tissue-engineered meshes was better organized than silk fibroin scaffolds.They also had higher failure force （3.081 ±0.121 vs 2.167 ±0.148 N） and Young＆#39;s modulus （3.191 ± 0.146 vs 2.263 ± 0.213 M Pa） and more collagen content （18.648 ± 0.867 vs 14.123 ± 0.989 μg/mg） than silk fibroin scaffolds （P 〈 0.05）.Conclusion As a suitable cell type for tissue engineering,ADSC may be successfully isolated and stably proliferated in vitro.The tissue-engineered meshes have an excellent biocompatibility and appropriate properties for pelvic floor reconstruction.