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FX Yu, X Gu; Characterization of hTERT-Immortalized Bovine Retinal and Human Brain Vascular Endothelial Cell Lines . Invest. Ophthalmol. Vis. Sci. 2002;43(13):3313.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: To develop an in vitro model for the study of blood retinal barrier (BRB) regulation, we sought to generate human tolemerase-immortalized bovine retinal and human brain vascular endothelial cell (VEC) lines that retain normal growth control and biological functions. Methods: Primary cultures of human brain and bovine retinal vascular EC were transfected with the catalytic component of human telomerase, human telomerase reverse transcriptase (hTERT), and colonies selected with puromycin. The endothelial origin of these cells was confirmed by immunocytochemistry. Matrigel was used to induce the formation of tubule-like structures. Results: Both bovine retinal and human brain microvascular endothelial cells expressing hTERT resembled young primary ECs in their morphology and growth response. More than 100 of population doubling attained thus far for the hTERT-containing cells and at present, the hTERT cells are still proliferating. Both bovine and human hTERT cell lines express von Willebrand factor, a key markers distinguishing ECs from other cell types both in vivo and in vitro, and uptake LDL. They form tubule-like structures in Metrigel and the stability of the «tubes» appears to be modulated by serum and growth factors. Conclusion: hTERT-immortalized ECs bypass replicative senescence and exhibit functional and morphogenetic characteristics of parental cells. Such characteristics are highly desirable for the molecular and genetic studies that are not permitted by primary culture. Our future studies will use these cell lines to elucidate the mechanisms underlying maintenance and breakdown of the BRB, in relation to the development of diabetic macular edema.
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