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An abnormal response of retinoblastoma cells (Y-79) to neurotrophins

AuthorsWagner, N.; Wagner, K. D.; Sefton, Mark; Rodríguez-Tebar, Alfredo ; Grantyn, R.
Issue Date2000
PublisherAssociation for Research in Vision and Ophthalmology
CitationInvestigative Ophthalmology and Visual Science 41: 1932-1939 (2000)
AbstractPURPOSE. To clarify the expression of neurotrophins and their receptors in retinoblastoma (Rb) cells, to elucidate their potential role in the proliferation of neuroectodermal tumor cells, and to establish conditions for Rb cell differentiation. METHODS. The Rb-derived cell line Y-79 was grown in serum-free suspension or monolayer culture. Proliferating and differentiated cells were isolated and submitted to semiquantitative reverse transcription- polymerase chain reaction (RT-PCR) analysis, immunostaining, and flow cytometry. The proliferation rate of the cells was estimated by 5-bromo-2'- deoxyuridine (BrdU) incorporation, and the effects of neurotrophins and laminin on BrdU-incorporation, process outgrowth, or immunostaining were determined. RESULTS. In contrast to previously studied normal retinal precursor cells, Y-79 cells not only express nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3), and p75, but also the corresponding high affinity receptors TrkA, TrkB, and TrkC. Proliferation was stimulated by exogenous and endogenous neurotrophin receptor ligands. Inhibition of protein kinase phosphorylation with K252a blocked proliferation and promoted differentiation. The effect of K252a on differentiation was enhanced by the addition of soluble laminin. After 9 days of combined treatment, the fraction of differentiated cells amounted to 30%, differentiation being characterized by improved attachment, neurite outgrowth, expression of NF-68, and a loss of glial fibrillary acidic protein (GFAP) and parvalbumin immunoreactivity. These changes were accompanied by a downregulation of TrkB and TrkC, but not TrkA or p75. Differentiated cells were isolated and further grown in the absence of K252a. However, despite the high level of TrKA expression in differentiated cells, the addition of NGF had no effect on their survival. CONCLUSIONS. A mitogenic action of neurotrophins could contribute to retinal tumor growth.
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