Abstract
Purpose: To investigate the effect of recombinant adenovirus-mediated human β-nerve growth factor (Ad-EGFP-hβ-NGF) on the differentiation of endothelial progenitor cells (EPCs) in rats.Methods: The successfully constructed Ad-EGFP-hβ-NGF and its negative control Ad-EGFP were infected into the isolated and purified rat EPCs to observe their morphological changes. Enzyme-linked immunosorbent assay (ELISA) was conducted to detect the levels of vascular endothelial growth factor (VEGF), von Willebrand factor (vWF) and basic fibroblast growth factor (bFGF) in different rat EPC culture solutions. Western blot was performed to determine the expression of tyrosine kinase receptor A (TrKA) protein in different groups of EPCs.Results: Primary fibrous EPCs were converted into epithelium-like cells. After infection with Ad-EGFPhβ- NGF for 1 week, some EPCs became round and exhibited neural stem cell-like changes. The expression levels of VEGF, vWF and bFGF in the Ad-EGFP-hβ-NGF infection group were significantly higher than those in the control group (p < 0.01). TrKA protein in Ad-EGFP-hβ-NGF infection was also significantly up-regulated compared with that in the negative control and blank control groups (p <0.01).Conclusion: β-NGF up-regulates the expression of TrKA receptor protein and secretion of angiogenic growth factors (i.e., VEGF, vWF and bFGF), thereby promoting the differentiation of rat EPCs, which may contribute to angiopoiesis or vascular repair.Keywords: β-Nerve growth factor, Endothelial progenitor cells, Angiogenic growth factors, Tyrosine kinase receptor A, Cell differentiation
Highlights
Endothelial progenitor cells (EPCs), known as angioblasts, are bone marrow-derived stem cells characterised by migration; these cells can further proliferate and differentiate into vascular endothelial cells[1]
The EPCs cultured for 28 days were infected with adenovirus
After the EPCs were infected for 6 days, the cell culture media were collected for the detection of vascular endothelial growth factor (VEGF), von Willebrand factor (vWF) and basic fibroblast growth factor (bFGF) through Enzyme-linked immunosorbent assay (ELISA)
Summary
Endothelial progenitor cells (EPCs), known as angioblasts, are bone marrow-derived stem cells characterised by migration; these cells can further proliferate and differentiate into vascular endothelial cells[1]. Recombinant adenovirus-mediated β-NGF was infected into rat EPCs to investigate its effects on EPC differentiation by determining whether NGF promotes endothelial regeneration through regulating EPC differentiation. The EPCs of rats cultured until the 28th day were divided into three groups and infected with Ad-EGFP-hβ-NGF and Ad-EGFP with 100 multiplicity of infection. After 48 h infection, the cell morphological changes were continuously observed. After the cells were infected for 6 days, the culture supernatant was collected and centrifuged at room temperature (2000 rpm, 3 min). The EPCs cultured for 28 days were infected with adenovirus (the negative control and blank control groups were set at the same time). After 48 h, the cell morphological observation under a fluorescence-inverted phase-contrast microscope revealed the expression of green fluorescent protein in the AdEGFP-hβ-NGF and Ad-EGFP groups.
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