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H.L. Deissler, G.K. Lang, G.E. Lang; Generation and Characterization of iBREC: A Novel hTERT Immortalized Bovine Retinal Endothelial Cell Line . Invest. Ophthalmol. Vis. Sci. 2005;46(13):4180.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: Primary bovine retinal endothelial cells (BREC) have proven extremely useful to study molecular mechanisms underlying development of diabetic retinopathy (DR). Several studies revealed that growth factors IGF–1 and VEGF are deregulated in DR which leads to an increased proliferation rate of retinal endothelial cells and to induction of neovascularization in vivo. However, the major drawback has been to prepare primary BREC without contaminating cells, e.g. pericytes which inhibit proliferation of endothelial cells. Further studies require high numbers of pure BREC which have a phenotype closely related to the primary cells. So far, BREC with an extended life span have only been described once. Methods: The cDNA for the catalytic domain of the human telomerase under the control of a CMV–promotor was introduced into primary BREC using lipofection technique. Immortalized cells (iBREC) were characterized by immunostaining and influence on proliferation by IGF–1, VEGF and TGFß2 was tested by [³H]–thymidine–incorporation and liquid scintilation counting. Results: iBREC have now been proliferating for more than 80 passages without changes in proliferation rate and cobble–stone morphology. Immunostaining revealed expression of the endothelial marker von Willebrand Factor. iBREC–cultures do not express αSMA indicating the absence of pericytes. hTERT expression is high in iBREC whereas it is low or not detectable in primary BREC. Proliferation of iBREC was stimulated moderately by VEGF (2x at 5 ng/ml) as well as by IGF–1 (2x at 5 ng/ml) in a similar manner found in primary BREC. In contrast, proliferation of iBREC was dramatically inhibited by TGFß2 (0.1x at 10 ng/ml) which is most likely due to transdifferentiation of endothelial cells to mesenchymal cells and not to apoptosis. Conclusions: iBREC can be used as a model not only to investigate molecular processes involved in DR but also for studies involving potential therapeutic drugs.
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