Experimental study of recombinant adenoviral-mediated PDGF gene delivery of human umbilical cord derived mesenchymal stem cells / 白血病·淋巴瘤

ABSTRACT

Objective To investigate the infect capability of recombinant adenovirus mediated PDGF to human umbilical cord derived mesenchymal stem cells (MSCs). Methods The PDGF cDNA sequence was amplified with RT-PCR and constructed recombinant adenovirus vector AdPDGF. The human umbilical cord derived MSCs were isolated and cultured. In vitro AdPDGF infected MSCs with various MOI,and determine the efficiency using FACS and fluorescence microscope. Trypan blue and MTT assay was used to detect cell viability and proliferation. ELISA was used to quantify PDGF secretion. Results Recombinant adenovirus AdPDGF was successfully and constructed,in which infection efficiency to MSCs was dose dependent. The highest efficiency was around 87.36 % when MOI =50,at which cell viability and proliferation wasn' t impaired. Cell viability of uninfected,AdPDGF infected and control virus infected MSCs was (97.8 ±2.3) %,(91.9±4.0) % and (92.8±4.0) %,separately. The cell proliferation was (100±16.8) %,(95.9±12.0) % and (87.5±9.7) %,separately. There was no statistic significance among AdPDGF infected,control virus infected and uninfected MSCs. 48 hrs post infection,PDGF secretion can be measured from infectious MSCs' supernatant. The secretion level was (1.53±0.37),(3.03±0.68) and (5.25±0.92) ng/ml,separately,when MOI= 10,30,50. No PDGF can be detected from MSC-GFP and control MSC group. Conclusion Recombinant adenovirus AdPDGF was constructed and can infect human umbilical cord derived MSC at high efficiency.