A retinal pigment epithelium-derived cell line from transgenic mouse harboring temperature-sensitive simian virus 40 large T-antigen gene.
We established a retinal pigment epithelium-derived cell line from transgenic mouse harboring temperature-sensitive simian virus 40 T-antigen gene (tsSV40T) and examined its characteristics. We enucleated both eyes from a 2-month-old transgenic mouse and removed the retinal pigment epithelial (RPE) cells and neuroretinal cells. After cloning the RPE cells, we obtained a cell line (RPET). RPET cells grew well at 33 degrees C but not at 37 degrees C, expressing on the temperature-sensitive character of tsSV40T, and maintained characters of RPE cells such as T1-tyrosinase production, phagocytosis of rod outer segments, and presence of cytokeratin, microvilli on the cell surface and lysosome-like granules around the Golgi apparatus in the cytoplasm. Conditioned medium (CM) from a culture of neuroretinal cells harboring tsSV40T was essentially required for growth. The factor(s) in CM was heat-and acid labile, but was resistant to trypsin digestion. In the presence of 3% CM, epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF) and strong effects on RPET cells, whereas insulin, insulin-like growth factor I (IGF-I), and IGF-II had moderate growth effects. Interestingly, none of these growth factors stimulated the RPET cells in the absence of CM. EHS-Matrix had growth effect, whereas laminin, collagen types I and IV, and fibronectin had no marked growth effects on RPET cells. RPET cells were morphologically changed on a laminin-coated dish. They could not spread on the coated dish, and the majority of the cells floated. But when the floating cells were transferred to non-coated dishes, they immediately attached themselves. These results suggest that RPET cells are a good model for for finding novel growth factor(s) and for investigating the mechanism of cell-laminin attachment.
Kato
K
K
Department of Ophthalmology, Tohoku University School of Medicine, Tohoku University, Sendai, Japan.
Ishiguro
S
S
Yamamoto
H
H
Yanai
N
N
Obinata
M
M
Tamai
M
M
D000818:Animals; D000952:Antigens, Polyomavirus Transforming / Q000235:genetics*; D002455:Cell Division; D002460:Cell Line; D004815:Epidermal Growth Factor / Q000494:pharmacology; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology; D007334:Insulin-Like Growth Factor I / Q000494:pharmacology; D007335:Insulin-Like Growth Factor II / Q000494:pharmacology; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D010857:Pigment Epithelium of Eye / Q000166:cytology / Q000187:drug effects / Q000821:virology*; D013539:Simian virus 40 / Q000235:genetics* / Q000302:isolation & purification; D013696:Temperature
D000952:Antigens, Polyomavirus Transforming; D016222:Fibroblast Growth Factor 2; D004815:Epidermal Growth Factor; D007334:Insulin-Like Growth Factor I; D007335:Insulin-Like Growth Factor II