Objective: To explore the effects of Notch signaling pathway and the vascular endothelial growth factor [VEGF(165)] gene on the functions of endothelial cells derived from rat bone marrow mesenchymal stem cells (MSCs).
Methods: Isolated and cultivated rat bone marrow MSCs in vitro, then the cells were treated by VEGF165 and basic fibroblast growth factor (bFGF) for 2 weeks to induce them to differentiate into endothelial cells. The gene of VEGF165 was transfected into differentiated endothelial cells to promote the functions of the cells. The receptor Notch1 and the ligand Jagged1 of the Notch signaling were detected by reverse transcription-polymerase chain reaction (RT-PCR) before and after the transfection. γ-secretase inhibitor L-685458 was used to block Notch pathway. Migration ability of cells was detected by scarification test. Cells were inoculated on semisolid gel to study their ability of forming capillary-like structure.
Results: After transfection, VEGF165 mRNA could be detected on the differentiated endothelial cells. The expression of Jagged1 mRNA was up regulated(1.08 ± 0.01 vs. 1.01 ± 0.02,P < 0.01) and there was no change in Notch1 mRNA(0.60 ± 0.02 vs. 0.59 ± 0.01,P > 0.05). The ability of migration was enhanced (number of cells on the scratched area:46.45 ± 4.46 vs. 41.61 ± 1.42,P < 0.05), and the ability of forming capillary-like structure on semisolid gel showed no change (cells classification: 3.00 ± 0.89 vs. 2.00 ± 0.89,P > 0.05). After the transfection, using the γ-secretase inhibitor L-685458 to block the Notch signaling transduction, the ability of migration of the differentiated endothelial cells (number of cells on the scratched area: 51.72 ± 3.47 vs. 46.45 ± 4.46,P < 0.05), and that of forming capillary--like structure (cells classification: 4.17 ± 0.75 vs. 3.00 ± 0.89, P < 0.05), was also further enhanced.
Conclusion: Transfection of the gene of VEGF165 into the differentiated endothelial cells can reinforce the function of these cells, and when Notch signaling was blocked, this effect can be further amplified.