The expression of basic fibroblast growth factor and its receptor in cell lines derived from normal human mammary gland and a benign mammary lesion

1993 ◽  
Vol 106 (1) ◽  
pp. 135-143
Author(s):  
Y. Ke ◽  
D.G. Fernig ◽  
M.C. Wilkinson ◽  
J.H. Winstanley ◽  
J.A. Smith ◽  
...  

mRNA for basic Fibroblast Growth Factor (bFGF) was expressed in a series of SV40-transformed human mammary cell lines as molecules of 7.1, 3.6, 2.0 and 1.2 kb. This expression was much weaker in those lines of epithelial morphology than in myoepithelial-like cell lines derived from them. It was confirmed, using northern hybridization to single-stranded RNA probes, that the multiple mRNAs were transcribed from the coding strand for bFGF. bFGF activity was detected in extracts of the cells and the relative amounts of activity corresponded in general to the amounts of mRNA found. Similar results were obtained from spontaneously transformed cell lines derived from a human benign breast lesion. The presence of bFGF protein in the extracts was confirmed by western blotting, which showed a band of 18–19 kDa, migrating in the same position as authentic bFGF; in addition, the myoepithelial-like cells showed prominent bands of bFGF at 24 and 26 kDa. No FGF receptor was detectable by the binding of 125I-bFGF to the SV40-transformed cell lines or to the epithelial cell lines from the benign breast lesion, but both high- and low-affinity receptors were found on myoepithelial-like cells derived from the latter. The results indicate that differentiation to the human myoepithelial-like phenotype in culture is associated with the enhanced expression of bFGF, and it is suggested that bFGF, immunocytochemically detected in the basement membrane of the human breast, may arise, at least in part, from the myoepithelial cells of the mammary parenchyma.

Development ◽  
1992 ◽  
Vol 115 (4) ◽  
pp. 1059-1069 ◽  
Author(s):  
G. Brill ◽  
N. Vaisman ◽  
G. Neufeld ◽  
C. Kalcheim

We present evidence that basic fibroblast growth factor (bFGF)-producing cells stimulate primary differentiation of neurons from neural crest progenitors. Baby hamster kidney (BHK-21) cells were stably cotransfected with plasmid pSV2/neo, which contains the gene conferring resistance to the neomycin analog G418 and expression vectors containing the human bFGF cDNA. Various clones, which differed in their bFGF production levels, were isolated. Homogeneous neural crest cells were cultured on monolayers of bFGF-producing, BHK-21-derived cell lines. While the parental BHK-21 cells, which do not produce detectable bFGF, had poor neurogenic ability, the various bFGF-producing clones promoted a 1.5- to 4-fold increase in neuronal cell number compared to the parental cells. This increase was correlated with the levels of bFGF produced by the different transfected clones, which ranged between 2.3 and 140 ng/mg protein. In contrast, no stimulation of neuronal differentiation was observed when neural crest cells were grown on monolayers of parental BHK cells transfected with plasmid pSV2/neo alone, or on a parental BHK-derived clone, which secretes high amounts of recombinant vascular endothelial growth factor (VEGF). Furthermore, the neuron-promoting ability of bFGF-producing cells could be mimicked by addition of exogenous bFGF to neural crest cells grown on the parental BHK line. A similar treatment of neural crest cells grown on laminin substrata, instead of BHK cells, resulted in increased survival of non-neuronal cells, but not of neurons (see also Kalcheim, C. 1989, Dev. Biol. 134, 1–10). Taken together, these results suggest that bFGF stimulates neuronal differentiation of neural crest cells by a cell-mediated signalling mechanism.


1991 ◽  
Vol 27 ◽  
pp. S24
Author(s):  
W. Macheiner ◽  
E. Kokron ◽  
K. Patocka ◽  
C. Cerni ◽  
M. Vetterlein ◽  
...  

1996 ◽  
Vol 16 (9) ◽  
pp. 4842-4851 ◽  
Author(s):  
A Caré ◽  
A Silvani ◽  
E Meccia ◽  
G Mattia ◽  
A Stoppacciaro ◽  
...  

Homeobox (HOX) genes control axial specification during mammalian development and also regulate skin morphogenesis. Although selected HOX genes are variably expressed in leukemias and kidney and colon cancer cell lines, their relationship with the neoplastic phenotype remains unclear. In both normal development and neoplastic transformation, HOX target genes are largely unknown. We investigated the expression and function of HOXB cluster genes in human melanoma. The HOXB7 gene was constitutively expressed in all 25 melanoma cell lines and analyzed under both normal and serum-starved conditions, as well as in in vivo primary and metastatic melanoma cells; conversely, HOXB7 was expressed in proliferating but not quiescent normal melanocytes. Treatment of melanoma cell lines with antisense oligomers targeting HOXB7 mRNA markedly inhibited cell proliferation and specifically abolished expression of basic fibroblast growth factor (bFGF) mRNA. Band shift and cotransfection experiments showed that HOXB7 directly transactivates the hFGF gene through one out of five putative homeodomain binding sites present in its promoter. These novel findings indicate a key role for constitutive HOXB7 expression in melanoma cell proliferation via bFGF. The results also raise the possibility that growth factor genes are critical HOX target genes in other developmental and/or neoplastic cell systems.


1990 ◽  
Vol 127 (1) ◽  
pp. 39-46 ◽  
Author(s):  
E. G. Black ◽  
A. Logan ◽  
J. R. E. Davis ◽  
M. C. Sheppard

ABSTRACT We have used a recombinant human basic fibroblast growth factor (basic FGF) to study its effects on cell proliferation, gene expression and accumulation of cyclic AMP (cAMP) and inositol phosphates in two well-characterized endocrine cell lines, FRTL-5 rat thyroid and GH3 rat pituitary cells. Basic FGF induced a dose-dependent increase in mitogenesis (assessed by measuring incorporation of [3H]thymidine) in FRTL-5 cells (40 ng basic FGF/ml increased mitogenesis above the control value by 2148±108% (mean ± s.e.m.), but inhibited mitogenesis in GH3 cells at all doses (85±4% of control with 40 ng basic FGF/ml)). Thyroglobulin mRNA concentration was increased in FRTL-5 cells (126±6% of control with 40 ng basic FGF/ml) as was prolactin mRNA in GH3 cells (246±11% of control with 40 ng basic FGF/ml), but GH mRNA in GH3 cells was not significantly affected by any dose of basic FGF. Intracellular cAMP was reduced by basic FGF in both FRTL-5 and GH3 cells (40 ng bFGF/ml giving 80±5% of the control value in FRTL-5, and 67±15% of the control value in GH3 cells) despite increased levels when FRTL-5 cells were stimulated with 150 μU TSH/ml (5645±484% of control) or GH3 cells were stimulated by 10 μmol forskolin/1 (3347±396% of control). In both FRTL-5 and GH3 cells, accumulation of [3H]inositol phosphates was increased by 40 ng basic FGF/ml (201±6 and 330±51% of control values respectively). We have shown that basic FGF has different effects on mitogenesis in the two cell lines; gene expression and accumulation of inositol phosphates were increased in both, whereas the intracellular concentration of cAMP was decreased. The actions of basic FGF may be mediated through both inhibition of adenylate cyclase and hydrolysis of phosphatidyl inositol bisphosphate as has been proposed for 3T3 fibroblasts. Our data suggest that there may be a physiological role for basic FGF in both thyroid and pituitary tissue. Journal of Endocrinology (1990) 127, 39–46


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