Basic Fibroblast Growth Factor Enhances the Growth and Expression of the Osteogenic Phenotype of Dexamethasone-Treated Human Bone Marrow-Derived Bone-like Cells in Culture S. PRI-CHEN, 1 S. PITARU, 2 F. LOKIEC, 1 and N. SAVION 1 1 Maurice and Gabriela Goldschleger Eye Research Institute and 2 Department of Oral Biology, Maurice and Gabriela Goldschleger School of Dental Medicine, Sackler Faculty of Medicine, Tel-Aviv University, Tel-Aviv, Israel Basic fibroblast growth factor (bFGF) was shown to enhance rat stromal bone marrow cells in culture to produce mineralized bone-like tissue in response to dexamethasone (Dex) treatment (Pitaru et al., J Bone Miner Res 8:919; 1993). The purpose of this study was to explore the effect of bFGF on Dex-treated human stromal bone marrow cells (hSBMC) in culture. Human SBMC from 6 patients were cultured for 14 days (P 0 ) and then subcultured and grown for 28 days in the presence of Dex (10 8 mol/L). The effect of bFGF on cell proliferation at P 0 and protein content, DNA content, alkaline phosphatase activity (ALP), osteocalcin secretion, and formation of mineralized bone-like tissue (MBT) at P 1 was analyzed. bFGF treatment resulted in a 2.4-fold increase in cell number at P 0 and a concentration-dependent increase in [ 3 H]-thymidine incorpora- tion at P 1 , reaching a maximum increase of 3.7-fold at a con- centration of 0.3 ng/mL. Furthermore, bFGF significantly in- creased both DNA content (two- to threefold), protein content (five- to sixfold), and the amount of MBT (up to 20-fold) at P 1 cultures. Morphological evaluation of the MBT at the electron microscope level revealed a mineralization process along colla- gen fibrils similar to the natural process. The osteogenic nature of the bFGF-treated cultures was further shown by their ALP activity, as well as osteocalcin secretion in response to 1,25- dihydroxyvitamin D 3 . In conclusion, bFGF demonstrated a stimulatory effect on the proliferation of Dex-treated hSBMC- derived osteoprogenitors while maintaining their capacity to fully differentiate and form bone-like tissue in culture. (Bone 23: 111–117; 1998) © 1998 by Elsevier Science Inc. All rights reserved. Key Words: Human bone cell; Basic fibroblast growth factor; Osteoblast; Mineralization; Bone marrow; Dexamethasone. Introduction Bone formation and remodeling involves the recruitment of osteo- progenitor cells from the bone marrow cells, their proliferation, and terminal differentiation into active osteoblasts. Failure or reduced activity of one of these processes results in impaired skeletal growth or reduction of bone mass. The majority of information regarding the development of the osteogenic lineage in the bone marrow has been obtained from in vitro or combined in vitro and in vivo studies that utilized animal-derived stromal bone marrow cells. 11,19 These studies point to the existence of stromal pluripotent stem cells that give rise to the fibroblastic, adipocytic, reticular, and osteoblastic cell lineages. The factors that promote and control a specific path- way of differentiation are still unknown. However, the aforemen- tioned studies indicate that, at least in vitro, serum concentration and source, steroid hormones, growth factors, and bone morphogenetic proteins might be important in this respect. 8,15 Dexamethasone (Dex) was found to be a prerequisite for the recruitment of osteoprogenitor cells and their terminal differen- tiation in cultures of rat stromal bone marrow cells. 16 Basic fibroblast growth factor (bFGF) was shown to enhance the proliferation of Dex-committed rat osteoprogenitors and to stim- ulate their differentiation in these cultures. 18,20 Data regarding the factors that regulate the osteogenic path- way of human stromal bone marrow cells (hSBMC) are consid- erably more sparse compared with that obtained from animal and particularly rodent models. It has been recently shown that Dex induces the differentiation of hSBMC along the osteoblastic lineage and enables the formation of mineralized-like tissue in culture. 13 Human bone cells were shown to express bFGF and to respond to this growth factor. 25 Basic fibroblast growth factor was also shown to activate the transcription of the human osteocalcin gene in ROS 17/2.8-transfected cells suggesting that bFGF may stimulate the differentiation of human bone cells. 23 However, in rat osteoblast-like cells, bFGF decreased the steady- state mRNA levels for osteocalcin, type I collagen, and alkaline phosphatase, indicating an inhibitory effect of bFGF on osteo- blastic differentiation. 22 Because of this conflicting evidence, and since we have shown that bFGF considerably stimulates the growth and differentiation of rat-derived stromal bone marrow cells, 20 we investigated the effect of bFGF on the growth and osteoblastic differentiation of hSBMC. Materials and Methods Materials Alpha-modified minimal essential medium (MEM), fetal calf serum, penicillin, streptomycin, amphotericin B, glutamine, and Address for correspondence and reprints: Prof. Naphtali Savion, Gold- schleger Eye Research Institute, Sheba Medical Center, Tel-Hashomer 52621, Israel. E-mail: eyeres@ccsg.tau.ac.il Bone Vol. 23, No. 2 August 1998:111–117 111 © 1998 by Elsevier Science Inc. 8756-3282/98/$19.00 All rights reserved. PII S8756-3282(98)00087-8