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

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 α1(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. © 2003 Wiley‐Liss, Inc.