Several members of the fibroblast growth factor (FGF) family have an important role in the development of skeletal tissues. FGF-8 is widely expressed in the developing skeleton, but its function there has remained unknown. We asked in this study whether FGF-8 could have a role in the differentiation of mesenchymal stem cells to an osteoblastic lineage. Addition of FGF-8 to mouse bone marrow cultures effectively increased initial cell proliferation as well as subsequent osteoblast-specific alkaline phosphatase production, bone nodule formation, and calcium accumulation if it was added to the cultures at an early stage of osteoblastic differentiation. Exogenous FGF-8 also stimulated the proliferation of MG63 osteosarcoma cells, which was blocked by a neutralizing antibody to FGF-8b. In addition, the heparin-binding growth factor fraction of Shionogi 115 (S115) mouse breast cancer cells, which express and secrete FGF-8 at a very high level, had an effect in bone marrow cultures similar to that of exogenous FGF-8. Interestingly, experimental nude mouse tumors of S115 cells present ectopic bone and cartilage formation as demonstrated by typical histology and expression of markers specific for cartilage (type II and IX collagen) and bone (osteocalcin). These results demonstrate that FGF-8 effectively predetermines bone marrow cells to differentiate to osteoblasts and increases bone formation in vitro. It is possible that FGF-8 also stimulates bone formation in vivo. The results suggest that FGF-8, which is expressed by a great proportion of malignant breast and prostate tumors, may, among other factors, also be involved in the formation of osteosclerotic bone metastases.

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Fibroblast growth factor 8 (FGF-8) is a secreted heparinbinding protein, which has transforming potential. Alternative splicing of the mouse Fgf-8 gene potentially codes for eight protein isoforms (a ± h) which di er in their transforming capacity in transfected cells. S115 mouse mammary tumor cells express a transformed phenotype and secrete FGF-8 in an androgen-dependent manner. In order to study the role of FGF-8 isoforms in the induction of transformed phenotype of breast cancer cells, we over-expressed FGF-8 isoforms a, b and e in S115 cells. Over-expression of FGF-8b, but not FGF-8a or FGF-8e, induced androgen and anchorage independent growth of S115 cells. FGF-8b-transfected S115 cells formed rapidly growing tumors with increased vascularization when injected s.c. into nude mice. FGF-8a also slightly increased tumor growth and probably tumor vascularization but FGF-8e was not found to have any e ects. The angiogenic activity of FGF-8b and heparinbinding growth factor fraction (HBGF) of S115 cell conditioned media was tested in in vitro and in vivo models for angiogenesis using immortomouse brain capillary endothelial cells (IBEC) and chorion allantoic membrane (CAM) assays. Recombinant FGF-8b protein was able to stimulate proliferation, migration, and vessellike tube formation of IBECs. In addition, stimulatory e ect of S115-HBGF on IBE cell proliferation was evident. A positive angiogenic response to FGF-8b was also seen in CAM assay. The results demonstrate that the expression of Fgf-8b is able to promote vessel formation. Angiogenic capacity probably markedly contributes to the ability of FGF-8b to increase tumor growth of androgen-regulated S115 mouse breast cancer cells. Oncogene (2001) 20, 2791 ± 2804.

Expression of fibroblast growth factor 8 (FGF-8) is commonly increased in prostate cancer. Experimental studies have provided evidence that it plays a role in prostate tumorigenesis and tumor progression. To study how increased FGF-8 affects the prostate, we generated and analyzed transgenic (TG) mice expressing FGF-8b under the probasin promoter that targets expression to prostate epithelium. Prostates of the TG mice showed an increased size and changes in stromal and epithelial morphology progressing from atypia and prostatic intraepithelial neoplasia (mouse PIN, mPIN) lesions to tumors with highly variable phenotype bearing features of adenocarcinoma, carcinosarcoma, and sarcoma. The development of mPIN lesions was preceded by formation of activated stroma containing increased proportion of fibroblastic cells, rich vasculature, and inflammation. The association between advancing stromal and epithelial alterations was statistically significant. Microarray analysis and validation with quantitative polymerase chain reaction revealed that expression of osteopontin and connective tissue growth factor was markedly upregulated in TG mouse prostates compared with wild type prostates. Androgen receptor staining was decreased in transformed epithelium and in hypercellular stroma but strongly increased in the sarcoma-like lesions. In conclusion, our data demonstrate that disruption of FGF signaling pathways by increased epithelial production of FGF-8b leads to strongly activated and atypical stroma, which precedes development of mPIN lesions and prostate cancer with mixed features of adenocarcinoma and sarcoma in the prostates of TG mice. The results suggest that increased FGF-8 in human prostate may also contribute to prostate tumorigenesis by stromal activation.