Repair of cartilage defects with stem cells loaded with silk fibroin degradation products self-assembled nano-peptide hydrogel

Objective To evaluate the repair of cartilage defects in New Zealand rabbits by self-assembled nano-peptide hydrogel-loaded stem cells loaded with silk fibroin degradation products. Methods The self-assembled nano-peptide hydrogel was constructed. The structure of silk fibroin scaffolds was detected by scanning electron microscopy. The rabbit bone marrow mesenchymal stem cells （BMSCs） were cultured. The expression levels of CD34, CD45, CD90 and CD105 proteins on the surface of second-generation BMSCs were detected by flow cytometry. The biocompatibility between hydrogel and BMSCs was assessed by living/dead cell dual staining. The proliferation of BMSCs was determined by CCK-8 assay. The expression levels of aggrecan （AGN）, COL2a1 and Sox-9 mRNA were detected by real-time quantitative PCR. The effect of hydrogel on the cartilage differentiation of BMSCs was evaluated. The repairing effect of hydrogel on cartilage defects was assessed by toluidine blue staining. Results Atomic force microscopy observed that functionalized self-assembled peptides could form nanofiber network structure. The surface proteins of the second-generation BMSCs showed that the proportion of CD105-positive and CD45-negative BMSCs was 91.22%, whereas that of CD90-positive and CD34-negative BMSCs was 95.10%. Biocompatibility test revealed that rabbit BMSCs grew stably in the self-assembled hydrogel, mainly seen as the green-stained live cells. CCK-8 assay showed that the proliferation capability of BMSCs after 3-d culture in the compound hydrogel group was significantly higher than that in the BMSCs group （P < 0.001）. PCR indicated that the expression levels of AGN, COL2a1 and Sox-9 mRNA at 2-4 weeks significantly differed among the blank, hydrogel, CM and hydrogel combined with CM groups （all P < 0.001）. The environment containing hydrogen was beneficial to the cartilage differentiation of BMSCs. Toluidine blue staining revealed that hydrogel-loaded stem cells were more effective than the BMSCs and controls in repairing bone defects （both P < 0.05）. Conclusion The self-assembled nano-peptide hydrogel loaded stem cells can promote the repair of cartilage defects.