脂肪血管基质成分复合骨软骨一体化支架的体外评价.

BACKGROUND: A large number of animal and clinical experiments have confirmed that local transplantation of adipose-derived stromal vascular fraction can promote articular cartilage repair. OBJECTIVE: To explore the feasibility of constructing tissue-engineered cartilage with acellular dermal matrix/biomineralized collagen integrated bone cartilage scaffold combined with autologous adipose-derived stromal vascular fraction. METHODS: The groin fat pads of both sides of the female rabbit were taken out under sterile conditions, and the stromal vascular fraction was obtained by enzyme digestion. Flow cytometry was used to detect the surface specific protein and observe the ability of adipogenic, osteogenic and chondrogenic differentiation. Adipose-derived stromal vascular fraction (experimental group), adipose-derived mesenchymal stem cells containing transforming growth factor-β3 (cartilage induction group) and adipose-derived mesenchymal stem cells containing 10% fetal bovine serum (control group) were co-cultured on acellular dermal matrix/biomineralized collagen scaffold. CCK8 assay was used to detect cell proliferation. The number of cells was observed under fluorescence microscope. Real time-quantitative PCR detection was used to detect cartilage related gene expression. The content of glycosaminoglycan in extracellular matrix was determined by dimethylmethylene blue colorimetry. RESULTS AND CONCLUSION: (1) The results of flow cytometry showed that the adipose-derived stromal vascular fraction had high expression of CD44, CD105 and CD90, and low expression of CD14, CD19 and CD45. Adipose-derived stromal vascular fraction had the ability to differentiate into adipocytes, osteoblasts and chondrocytes. (2) CCK8 assay showed that with the extension of culture time, the number of three kinds of cells on the acellular dermal matrix/ biomineralized collagen scaffold increased, and the number of cells in the experimental group was higher than that in the other two groups (P < 0.05). Under the fluorescence microscope, the number of cells in the experimental group was more than that in the other two groups at 7 days after culture. (3) Real timequantitative PCR results showed that at 21 days after culture, the mRNA expression levels of Sox9, proteoglycan and type II collagen in the experimental group were higher than those in the other two groups (P < 0.05). (4) Dimethylmethylene blue colorimetry results demonstrated that at 7, 14, and 21 days, the content of glycosaminoglycan in the experimental group was higher than that in the other two groups (P < 0.05). (5) The results showed that adipose-derived stromal vascular fraction combined with acellular dermal matrix / biomineralized collagen scaffold could effectively support cartilage formation in vitro. [ABSTRACT FROM AUTHOR]