自体脂肪源性间充质干细胞修复兔软骨缺损的实验研究

目的 评价腺病毒介导的人转化生长因子β2(Ad-hTGF-β2)基因转染自体兔脂肪间充质干细胞(ADSCs)复合聚乳酸(PLGA)/藻酸钙对关节软骨缺损的修复效果.方法 将20只新西兰大白兔制备双侧膝关节软骨缺损模型.体外培养扩增自体ADSCs,转染Ad-hTGF-β2后,种植于PLGA上,然后将细胞_支架复合物植入兔关节软骨缺损模型;缺损处植入单纯支架、缺损处不做任何处理者,作为对照组.分别于术后4、12及24周处死动物进行大体及组织学观察,并行Ⅱ型胶原免疫组织化学染色.结果 实验组修复区可见新生的陷窝状细胞形成,与周围结合紧密.对照组见大量成纤维细胞,未见软骨样陷窝细胞形成.Ⅱ型胶原染色实验组呈阳性,对照组无明显阳性可见.结论 Ad-hTGF-β2基因转染的白体ADSCs能够修复关节软骨缺损.

Repair of articular cartilage defects by autologous adipose derived mesenchymal stem cell experiment with rabbits

Objective To evaluate the effectiveness of autologous adipose derived mesenchymal stem cells (ADSCs) tranduced by adenovirus-mediated human transforming growth factor 2 (Ad-hTGF-β2) gene in the repair of articular cartilage defects. Methods Rabbit ADSCs were obtained, cultured, and transfected with Ad-hTGF-β2 containing human transforming growth factor (hTGF) - β2. Three days later RT-PCR was used to detect the mRNA expression of hTGF- β2 in the ADSCs, and ELISA was used to detect the protein expression of hTGF-β2 in the supernatant, phosphorylation of Smad was examined by Western blotting. Articular cartilage defects at the femoral trochlea were made on 20 rabbits (40 sides) so as to establish animal models. The culture-expanded rabbit ADSCs transfected with Ad-hTGF- 2 were seeded on poly (L-lactic-co-glycolic acid) (PLGA) scaffolds. The cell-adhered PLGA scaffolds were implanted into the articular cartilage defects. Plain PLGA was implanted into the left-side defects of 10 rabbits as control group and the defects of 10 sides remained untreated as blank control group. The rabbits were sacrificed 4, 12, and 24 weeks after the operation respectively. The specimens of defects were examined histologically and stained immunohistochemically for type Ⅱ collagen. Results After transfection the ADSCs expressed mRNA and protein expression of hTGF- β2 and Western blotting showed bands of phosphorylated Smad. The cartilage specimens harvested from the experimental group rabbits demonstrated hyaline cartilage formation mingled closely with the nearby tissues and expression of type Ⅱ collagen. However, only fibroblasts, not cartilage-like cells, were seen in the control groups that lacked the expression of type Ⅱ collagen too. Conclusion Culture-expanded autologous ADSCs adhered with PLGA composites facilitate the formation of hyaline-cartilage.