Stromal activation associated with development of prostate cancer in prostate-targeted fibroblast growth factor 8b transgenic mice.

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.

Androgen and fibroblast growth factor 8 (FGF8) downregulation of thrombospondin 1 (TSP1) in mouse breast cancer cells.

In the search for androgen target genes responsible for malignant growth in S115 mouse mammary tumor cells we found that thrombospondin 1 (TSP1) expression was strongly downregulated by testosterone (Te). Experiments with cycloheximide suggested that Te repression of TSP1 was dependent on de novo protein synthesis. TSP1 repression by Te was preceded by the induction of fibroblast growth factor 8 (FGF8) expression. FGF8 has previously been shown to mediate androgen effects on proliferation of S115 cells by autocrine/paracrine mechanisms. It has also been shown to increase breast cancer cell growth as tumors in nude mice and to stimulate tumor angiogenesis. We studied here the possibility that FGF8 belonged to the Te-induced de novo synthesized proteins that mediate the effect of Te on TSP1 expression in these cells. We found that addition of FGF8b to in vitro cultures or ectopic expression of FGF8b in S115 cells repressed TSP1 expression at mRNA and protein levels even in the absence of Te. FGF2, another angiogenic member of FGF family, also downregulated TSP1 mRNA level in the in vitro cultures of S115 cells. The antisense oligonucleotides for FGF8 did not, however, prevent Te-repression of TSP1 mRNA expression and a neutralizing anti-FGF8b antibody only partially opposed Te induced downregulation of TSP1. These results suggest that both androgen and FGF8 inhibit TSP1 expression independently. They also suggest that opposite to many other androgen-induced responses in S115 cells, the effect of Te on the expression TSP1 is not mediated by FGF8.

Regulation of osteoblast differentiation: a novel function for fibroblast growth factor 8.

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.