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C. M. Cebulla, M.-E. Jockovich, Y. Pina, A. Alegret, H. Boutrid, A. S. Hackam, W. Feuer, T. G. Murray; Basic Fibroblast Growth Factor Expression in Retinoblastoma Progression and Survival. Invest. Ophthalmol. Vis. Sci. 2007;48(13):3987.
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Basic fibroblast growth factor (bFGF) is expressed by many tumors. It offers some a survival advantage by inhibiting apoptosis, while inducing apoptosis in others, such as PNET and Ewing’s sarcoma. Herein we test the hypothesis that bFGF is expressed in retinoblastoma (RB) tumors and determine whether exogenous bFGF is pro- or anti-apoptotic in RB.
The animal study protocol was approved by the University of Miami IACUC and human tissue studies approved by the IRB. Immunofluorescence analysis for bFGF was performed in the LH-ßTag transgenic mouse model of RB, human RB cell lines, and primary tumors. RT-PCR was performed to evaluate bFGF expression in human RB Y79 cells. A timecourse analysis for the expression of intraocular bFGF was performed in LH-ßTag mice and controls at 4 wk (preneoplastic stage, n=4), 8 wk (small tumor stage, n=4), and 16 wk (large tumor stage n=2). Proliferation assays to determine RB growth responses to exogenous bFGF (0, 0.01ng, 0.1ng, 1 ng, 10 ng) were performed with WERI and Y-79 human RB cells by counting cells after 4 days with a Coulter Particle Counter. TUNEL assays were performed to evaluate apoptosis in WERI and Y-79 cell lines in response to 24h carboplatin treatment, and test the hypothesis that exogenous bFGF enhanced carboplatin induced apoptosis.
During tumorigenesis, bFGF immunofluorescence is present in transgenic RB, primarily in a vascular pattern. The bFGF expression is lowest in 4 wk mice, highest in 8 wk mice and intermediate in 16 wk mice. The elevation in bFGF levels is statistically significant in the older animals (8 week, p=0.0059; 16 week, p=0.041). Immunofluorescence also demonstrates bFGF is expressed in primary human retinoblastoma. RT-PCR confirmed the presence of bFGF expression in Y-79 cells. Proliferation assays demonstrated no significant proliferation of Y-79 cells in response to bFGF. However, significant proliferation of WERI cells was induced in response to 0.1ng bFGF (22% increase, p=0.04). Interestingly, the effect of bFGF on carboplatin-induced apoptosis was very different in the Y-79 and WERI cells. WERI apoptosis was significantly enhanced by bFGF (0.1ng), while no significant effect of bFGF was detected on Y-79 apoptosis levels.
Human RB cell lines, primary tumors, and a transgenic mouse model of RB express bFGF. Similar to findings in PNET tumors, bFGF is pro-apoptotic in the less aggressive WERI cell line, but has a minimal effect on the more aggressive Y-79 line. Thus, exogenous bFGF may be a useful adjunct to chemotherapy in some cases of RB.
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