Effect of overexpressing fibroblast growth factor 2 protein isoforms in osteoblastic ROS 17/2.8 cells

Abstract

Fibroblast growth factor-2 (FGF-2) is made by osteoblasts and modulates their function. There are high molecular weight (HMW) protein isoforms of FGF-2 that have nuclear localization sequences and a low molecular weight (LMW) 18 kDa FGF-2 protein that is exported from cells. Since FGF-2 is a trophic factor and potent mitogen for osteoblasts, the goal of this study was to utilize targeted overexpression of FGF-2 as a novel means of assessing different FGF-2 isoforms on osteoblastic cell viability and proliferation. Either LMW or HMW human Fgf2 cDNAs were cloned downstream of 3.6 kb alpha1(I)-collagen 5' regulatory elements (Col 3.6). A set of expression vectors, called Col3.6-Fgf2 isoforms-IRES-GFPsaph, capable of concurrently overexpressing either LMW or HMW FGF-2 isoforms concomitant with GFPsaph from a single bicistronic mRNA were built. Viable cell number in ROS 17/2.8 cells stably transfected with Vector (Col3.6-IRES-GFPsaph) versus each of the Col3.6-Fgf2-IRES-GFPsaph constructs were compared. In the presence of 1 or 10% serum, DNA synthesis was increased in cells expressing any isoform of FGF-2 compared with vector. However, cells transfected with HMW isoform had augmented DNA synthesis in 1 or 10% serum compared with cells expressing either ALL or LMW FGF-2 isoforms. A neutralizing FGF-2 antibody significantly reduced the mitogenic response in cells harboring ALL or the LMW FGF-2 isoforms but did not block the mitogenic effect of cells harboring the HMW isoforms. In summary, overexpression of any isoform of FGF-2 protein increased viable cell number and OB proliferation in the presence of low or high concentrations of serum. However, the HMW/nuclear isoforms preferentially mediate augmented OB proliferation. We conclude that differential expression of FGF-2 proteins isoforms is important in modulating OB function.

The cardioprotective effect of the low molecular weight isoform of fibroblast growth factor-2: The role of JNK signaling

“…Transfection of FGF2 isoforms in an osteoblastic cell line showed that neutralizing FGF2 antibodies blocked cell growth in FGF2 LMW overexpressing cells but not in FGF2 HMW isoforms overexpressing cells, suggesting that the FGF2 HMW isoforms modulate cell growth and DNA synthesis by a mechanism independent of its cell surface receptors [29]. In cardiomyocytes, both LMW and HMW isoforms increase proliferation while FGF2 HMW overexpression significantly increased binucleation [30].…”

In Vitro Functional Assessment of Human Skeletal Myoblasts After Transduction With Adenoviral Bicistronic Vector Carrying Human VEGF165 and Angiopoietin-1

“…14, 24 Xiao and colleagues recently reported a bicistronic vector encoding for basic fibroblast growth factor (bFGF) along with the green fluorescent protein (GFP) reporter gene. 25 There is however, no information about 1 vector simultaneously encoding for 2 angiogenic growth factors to achieve therapeutic angiogenesis. Moreover, adenoviral vector administration triggers host inflammatory reactions and immune response.…”

High molecular weight FGF2: the biology of a nuclear growth factor

“…The proliferative activity of HMW FGF2 has been well documented in NIH3T3 and A31 fibroblasts, adult bovine aortic endothelial (ABAE) cells, cardiac myocytes and ROS17/2.8 osteosarcoma cells [39, 48–53]. In NIH3T3 cells, this proliferation also takes place in low-serum conditions [29, 49, 50].…”