Lentiviral-mediated gene transfer into human adipose-derived stem cells: role of NELL1 versus BMP2 in osteogenesis and adipogenesis <italic>in vitro</italic>

Abstract

NEL-like molecule 1 (NELL1) is a potent osteogenic factor associated with craniosynostosis. Adenoviruses, the most commonly used viral vectors for gene therapy, have several disadvantages that may restrict osteogenesis. Previous studies have shown that lentiviruses can serve as ideal vectors for gene therapy for bone regeneration. In this study, two lentiviral vectors (LvNELL1 and LvBMP2) that encode human NELL1 and bone morphogenetic protein-2 (BMP2), respectively, were constructed. The effect of LvNELL1 infection on the proliferation, osteogenesis, and adipogenesis of human adipose-derived stem cells (hADSCs) in vitro was assessed and compared with that of LvBMP2. The results showed that hADSCs infected with LvNELL1 could efficiently and stably overexpress the target genes. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay results demonstrated that LvBMP2, but not LvNELL1, enhanced the proliferation of hADSCs. Assessment of alkaline phosphatase activity and cellular mineralization indicated that LvNELL1 infection promoted the osteogenic differentiation of hADSCs, and the effect was comparable with that of LvBMP2. Real-time polymerase chain reaction (PCR) revealed that LvNELL1 infection upregulated OSX expression but not RUNX2 expression in hADSCs. In addition, adipogenic markers (lipid droplets, peroxisome proliferator-activating receptor g, and lipoprotein lipase) analysis showed that LvNELL1 could dramatically inhibit the adipogenic differentiation of hADSCs, but LvBMP2 had no such effect. Taken together, these findings suggested that lentiviral-mediated NELL1 gene transfer in hADSCs may be a novel and promising approach to achieve effective and precise bone regeneration.

Effect of NELL1 gene overexpression in iPSC-MSCs seeded on calcium phosphate cement

“…In the present study, ALP activity was increased by 130% at 14 d and bone matrix mineralization was increased by 81% at 21 d due to NELL1 overexpression. These results suggest that NELL1 yielded osteogenic differentiation and mineral production of iPSC-MSCs comparable to those of BMP2, consistent with the previous findings [11,57]. …”

“…There was no significant difference in cell proliferation between RFP-iPSC-MSCs and NELL1-iPSC-MSCs. Therefore, NELL1 appeared to affect the osteogenic differentiation rather than the proliferation of iPSC-MSCs, consistent with the previous results on ADSCs [57]. RGD was immobilized on chitosan and incorporated into CPC in order to improve cell attachment, growth, and osteogenic differentiation.…”

“…In another study, however, NELL1 had no inhibition on the proliferation of human ADSCs [57]. In the present study, iPSC-MSCs were seeded on RGD-grafted CPC and the cell growth was determined by nuclear counting.…”

“…NELL1 cannot induce ectopic bone formation in muscle, which is different form BMP2 [15]. Previous studies compared the effects of BMP2 and NELL1 gene overexpression on osteogenic differentiation using goat BMSCs or human adipose-derived stem cells (ADSCs) [11,57]. It was demonstrated that NELL1 and BMP2 had comparable effects on osteogenic differentiation.…”

Potential therapeutic Strategies and Its Application in Correcting Birth Defects, Craniosynostosis, Neurological Disorders and Other Diseases

“…Basically there are two lanti viral vectors (Lv NELL 1 and Lv BMP2). Researcher observed that these genes transfer in hADSCs may be a novel and potential approach for bone regeneration [3]. NELL 1 is able to promote the osteogenic differentiation.…”