Abstract: BACKGROUND: The tissue engineering module provides a new idea for the treatment of femoral head necrosis. A vascularized scaffold was constructed and transplanted into the necrotic area, and the bone marrow mesenchymal stem cells were induced to differentiate into vascular endothelial cells under the action of growth factors and scaffolds, which promoted the generation of microvessels and improved local blood vascularity, thereby promoting the repair of the necrotic area. 
OBJECTIVE: To investigate the feasibility of constructing a dendritic amphiphilic peptide self-assembly gel module with the ability to induce the differentiation of bone marrow mesenchymal stem cells into vascular endothelial cells in vitro. 
METHODS: Bone marrow mesenchymal stem cells were isolated and extracted from SD rats aged 4-6 weeks and identified by flow cytometry. In vitro directed adipogenic and osteogenic induced differentiation was used to detect the multidirectional differentiation potential of cells. Peptides            KGGGGAAAA(K)-C16H31O were synthesized by solid-phase method. The concentration was 10 mg/mL peptide solution. After the addition of normal salt solution, the peptide self-assembly was triggered, forming a translucent hydrogel. Transmission electron microscopy was utilized to observe the gel structure. In the experimental group, rat bone marrow mesenchymal stem cells were transfected with adenovirus carrying vascular endothelial growth factor 165 gene. The 10 mg/mL peptide solution was mixed with the cell suspension, and the cells were distributed inside the gel to form a three-dimensional culture system. In the control group, bone marrow mesenchymal stem cells transfected with empty virus were mixed with 10 mg/mL peptide solution to form a three-dimensional culture system. Immunofluorescence and RT-qPCR were used to detect the differentiation of vascular endothelial cells after 7 days of in vitro culture.
RESULTS AND CONCLUSION: (1) Flow cytometry results showed that the expression of CD29 and CD44 on the surface of mesenchymal stem cells was ≥95%, and the expression of CD34 and CD45 on the surface of hematopoietic stem cells was ≤2%. Bone marrow mesenchymal stem cells were successfully differentiated by osteogenic induction and adipogenic induction in vitro. (2) The dendritic peptide solution was triggered by ions to form a transparent gel, and the small-sized nanomicelles with uniform morphology were observed under electron microscopy. (3) The results of immunofluorescence and RT-qPCR showed that the expression level of vascular endothelial growth factor 165 in the experimental group was higher than that in the control group, and the expression levels of CD31 and CD34 were higher than those in the control group. These results suggest that vascular endothelial growth factor 165 gene transfected bone marrow mesenchymal stem cells implanted in a tissue engineering module constructed by peptide gel can induce the differentiation of bone marrow mesenchymal stem cells into vascular endothelial cells. 

Key words: dendritic amphiphilic peptide">, bone marrow mesenchymal stem cell">, gene transfection">, vascular endothelial growth factor 165">, three-dimensional culture">, vascularization">, femoral head necrosis