[Biological Characteristics of Human Umbilical Cord Mesenchymal Stem Cells Transduced with a BMP4-Carrying Adenovirus].

OBJECTIVE

To evaluate the effects of Bone Morphogenetic Protein 4(BMP4) overexpression mediated by an adenoviral vector on immunophenotype features, migration ability, colony-forming ability, and early osteogenic differentiation potential of human umbilical cord-derived mesenchymal stem cells (hUC-MSCs).

METHODS

hUC-MSCs were isolated and cultured. An adenoviral vector carrying the human BMP4 gene (Ad-BMP4) and a corresponding null vector (Ad-NC) were constructed. The optimal multiplicity of infection (MOI) was determined through preliminary experiments. Subsequently, the cells were divided into three groups: Control group, Ad-NC group, and BMP4 overexpressing group (Ad-BMP4). Transduction efficiency and expression levels of BMP4 were verified by real-time quantitative PCR and Western blot. Following transduction, the expression of MSC-related surface markers (CD73, CD90, CD105) was assessed by flow cytometry. A wound healing assay was performed to evaluate the effect of BMP4 overexpression on the migration ability of hUC-MSCs. A colony-forming unit-fibroblast (CFU-F) assay was used to compare the colony-forming ability among the groups. Early osteogenic differentiation potential was assessed by alkaline phosphatase (ALP) staining, and the mRNA expression levels of osteogenesis-related genes (ALP, SPP1) were measured by RT-qPCR.

RESULTS

hUC-MSC overexpressing BMP4 was successfully constructed. At the optimal MOI, the mRNA and protein levels of BMP4 in the Ad-BMP4 group were significantly higher than those in the Control and Ad-NC groups. Flow cytometry results indicated that the Ad-BMP4 group expressed high levels of CD73, CD90, and CD105, while showing low to no expression of CD34, CD45, and HLA-DR, confirming that the cells retained a typical MSC immunophenotype post-transduction. The wound healing assay revealed no significant effect of BMP4 overexpression on the in vitro migration of hUC-MSCs compared to the control groups. The CFU-F assay demonstrated that the number of cell colonies in the Ad-BMP4 group was significantly increased compared to the Control and Ad-NC groups (P<0.05). In osteogenic differentiation assays, the Ad-BMP4 group exhibited more intense ALP staining, and the expression levels of osteogenic genes ALP and SPP1 were significantly upregulated (P<0.05).

CONCLUSION

Adenovirus-mediated overexpression of BMP4 enhances the colony-forming ability and early osteogenic differentiation potential of hUC-MSCs, but has no obvious effect on their phenotype and migration ability, suggesting the potential application value of BMP4-modified hUC-MSCs in tissue regeneration.