VEGF165 expressing bone marrow mesenchymal stem cells differentiate into hepatocytes under HGF and EGF induction in vitro

Abstract

A short half-life and low levels of growth factors in an injured microenvironment necessitates the sustainable delivery of growth factors and stem cells to augment the regeneration of injured tissues. Our aim was to investigate the ability of VEGF 165 expressing bone marrow mesenchymal stem cells (BMMSCs) to differentiate into hepatocytes when cultured with hepatocyte growth factor (HGF) and epidermal growth factor (EGF) in vitro. We isolated, cultured and identified rabbit BMMSCs, then electroporated the BMMSCs with VEGF 165 -pCMV6-AC-GFP plasmid. G418 was used to select transfected cells and the efficiency was up to 70%. The groups were then divided as follows: Group A was electroporated with pCMV6-AC-GFP plasmid ? HGF ? EGF and Group B was electroporated with VEGF 165 -pCMV6-AC-GFP plasmid ?HGF ? EGF. After 14 days, BMMSCs were induced into short spindle and polygonal cells. Alpha-fetoprotein (AFP) was positive and albumin (ALB) was negative in Group A, while both AFP and ALB were positive in group B on day 10. AFP and ALB in both groups were positive on day 20, but the quantity of AFP in group B decreased with prolonged time and was about 43.5% less than group A. The quantity of the ALB gene was increased with prolonged time in both groups. However, there was no significant difference between group A and B on day 10 and 20. Our results demonstrated that VEGF 165 -pCMV6-AC-GFP plasmid modified BMMSCs still had the ability to differentiate into hepatocytes. The VEGF 165 gene promoted BMMSCs to differentiate into hepatocyte-like cells under the induction of HGF and EGF, and reduced the differentiation time. These results have implications for cell therapies.

Assessment of the Developmental Toxicity of Epidermal Growth Factor using Embryonic Stem Cell Test

“…Compared to the above, the expression of the ectodermspecific gene AFP, which is widely used as a hepatic marker, showed a dose-dependent increase and decreased when the concentration of EGF exceeded 5 μg/mL, which suggests that a low concentration of EGF promotes embryonic stem cell differentiation into hepatic cells, while, toxicity at a high concentration of EGF leada to inhibition. It has been reported that 20-45 ng/mL EGF can be used as an inducer to promote hepatic cell directional differentiation from stem cells [21]. Our results showed the promotion of hepatic differentiation effects of EGF on increased expression of the AFP gene.…”

Long‐term maintenance of HepaRG cells in serum‐free conditions and application in a repeated dose study

“…The slight increase in viability after 18 days of cultivation could be because of the regeneration of the hepatic cells, as occurring in vivo (Vodovotz et al, 2012). As the increase in viability is limited to serum-free conditions, it might be related to the presence of HGF, which is known to be a mitogen as well as for playing a role in liver regeneration (Forte et al, 2006;Tan et al, 2012).…”

“…However, other medium components, such as growth factors, also play a crucial role in the upkeep of hepatic functions. For example, hepatocyte growth factor (HGF) and epidermal growth factor (EGF) are important for the differentiation of hepatocytes and they can lead to improved liver-specific functions in vitro (Kim et al, 2010;Tan et al, 2012).…”

Genetic Engineering of Mesenchymal Stem Cells for Regenerative Medicine

“…In the case of liver regeneration attempts, VEGF was likewise recognized to promote BM-MSC differentiation into hepatocyte-like cells under the specific condition of HGF and EGF stimulation [144]. However, most studies have been focused on attempts to induce hepatic differentiation of engineered MSCs with physiologically committed factors.…”

“…GDF-5 [102][103][104] TIMP-1 [106] Bcl-2 [101] BONE FORMATION BMP-2 [86][109-111] BMP-2/SDF-1β [113] BMP-2/HIF-1α [114] BMP-2/Ang-1 [47] BMP-2/VEGF [116][117][118] BMP-2/miR-148b [142] BMP-4/-6/-7 [125,126] VEGF [119,120] Epo [121] IGF-I [128,129] OGP [127] Leptin [130] PDGF [131] ephrinB2 [132] ITGA2 [133] NELL1 [134] β-catenin [135] HI Cb TA Fo Na LIVER REGENERATION VEGF [144] ATP7B [155] HN HN…”