I型胶原-PRP支架促进肌腱干细胞功能性分化作用的研究

The purpose of this study is to evaluate the effects of collagen type I-PRP composite on promoting functional differentiation of tendon-derived stem cells (TDSCs). In this study, ①TDSCs will be isolated form rabbit patellar or Achilles tendon; The isolated TDSCs will be characterized by immunostaining for stem cell markers and characterized for its multi-differentiation potential. ②TDSCs at passage 2 will be cultured on collagen I-PRP scaffold, qRT-PCR, immunofluorescence, western-Blot will be used to characterize the mRNA expression of Collagen type I, Tenascin C, Oct-4, Nanog and immunohistochemical analyses will be used to characterize the protein expression of cell markers; ③TDSCs will be mixed with collagen I-PRP composite and implanted subcutaneously into the abdominal areas of nude rats. HE staining will be used to test the organization of the tissue and immunohistochemical analyses will be used to test the protein expression of collagen types I and CD31. This study will provide us with a new scaffold for tendon repairing with tissue engineering method.

肌腱损伤后难以自身修复。富血小板血浆（PRP）中富含生长因子，激活形成凝胶后可以作为支架材料修复损伤肌腱，但PRP凝胶在组织内降解较快；研究表明I型胶原能激活血小板，且I型胶原能为种子细胞提供合适的生长环境。我们提出“使用肌腱来源的I型胶原激活PRP，形成I型胶原-PRP支架，适合肌腱干细胞（TDSCs）的生长，能促进TDSCs功能性分化，有促进TDSCs成肌腱的作用”。本项目：①分离、培养、鉴定兔TDSCs；②将TDSCs种植于I型胶原-PRP支架上，用qRT-PCR、免疫荧光、Western-Blot鉴定干细胞标记物和肌腱相关标记物的表达；③将TDSCs与I型胶原-PRP支架混合培养，种植于裸鼠皮下，检测TDSCs功能蛋白的表达量，并移植于兔肌腱缺损部位，通过组织学及生物力学方法，评估其对损伤肌腱的修复作用。此研究将为组织工程方法修复肌腱损伤提供新的支架材料。