Rabbit bone marrow mesenchymal stem cells transfected with recombinant lentivirus and decalcified bone matrix to construct transgenic tissue engineering materials

doi:10.12307/2025.501

Abstract

Abstract: BACKGROUND: The use of autologous or allogeneic transplantation materials for tissue repair and reconstruction is limited and carries limitations in clinical applications. Transgenic stem cells and tissue engineering materials offer new therapeutic possibilities.
OBJECTIVE: To investigate the in vitro biological characteristics of enhanced green fluorescent protein recombinant lentivirus transfected into rabbit bone marrow mesenchymal stem cells and the biommineralization characteristics of transgenic tissue engineering materials constructed using decalcified bone matrix.
METHODS: Rabbit bone marrow mesenchymal stem cells were obtained through adherent culture and density gradient centrifugation. The fifth-generation bone marrow mesenchymal stem cells were transfected with enhanced green fluorescent protein recombinant lentivirus at the optimal multiplicity of infection (MOI=100). The differences in cell proliferation capacity, cell phenotype, cell cycle, alkaline phosphatase expression, osteogenic transcription factor (Runx2) expression, and osteocalcin gene expression after osteogenesis were observed between the transfected cells and non-transfected cells in vitro. The micromorphology and elemental energy spectrum analysis of the transgenic tissue engineering materials constructed from cells and decalcified bone matrix scaffold were also observed.
RESULTS AND CONCLUSION: Bone marrow mesenchymal stem cells transfected with enhanced green fluorescent protein recombinant lentivirus showed that cell proliferation at 24 and 48 hours after transfection was slower than that of untransfected cells (P < 0.05). After 72 hours, the cell phenotype remained unchanged, and the expressions of alkaline phosphatase, Runx2, and osteocalcin after osteogenic induction were not significantly different from those of untransfected bone marrow mesenchymal stem cells (P > 0.05). The transgenic tissue engineering materials constructed in vitro using enhanced green fluorescent protein recombinant lentivirus transfected bone marrow mesenchymal stem cells and decalcified bone matrix showed good biocompatibility on the decalcified bone matrix scaffold. Based on fluorescence expression intensity, it was estimated that the target gene would exert its maximum biological function in about 2 weeks, and calcium phosphate deposits would continue to appear, demonstrating superior biomineralization characteristics.

Key words: recombinant lentiviral vector, enhanced green fluorescent protein, bone marrow mesenchymal stem cell, gene therapy, biological characteristics

CLC Number:

Ning Yinkuan, Liu Linzhi, Zhou Cila, Long Yubin. Rabbit bone marrow mesenchymal stem cells transfected with recombinant lentivirus and decalcified bone matrix to construct transgenic tissue engineering materials[J]. Chinese Journal of Tissue Engineering Research, 2025, 29(23): 4851-4858.

Figures/Tables 8

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