Selective and efficient culturing of retinal pigment epithelial cells using a feeder layer.
BACKGROUND
The techniques to isolate and purify retinal pigment epithelial (RPE) cells from small piece of autologous tissues are extremely difficult, and it is important to develop an efficient cell culture technique for RPE cells. The purpose of this study was to investigate the effect of 3T3-J2 cells and conditioned medium from 3T3-J2 cells on the proliferation of cultured RPE cells.


METHODS
RPE cells from pigmented rabbits and a human RPE-derived cell line, ARPE-19, were used. First, the effects of co-culturing RPE cells with 3T3-J2 cells on the growth of the cells were analyzed. Second, the effects of the conditioned medium from 3T3-J2 cells on the proliferation of both types of cells were investigated. And third, the effects of the conditioned medium on RPE cell culture from a surgically removed choroidal neovascular (CNV) membrane were investigated.


RESULTS
The 3T3-J2 cells increased the proliferation of both rabbit RPE cells and ARPE-19 cells. The number of rabbit RPE cells cultured in a mixture of the conditioned medium from 3T3-J2 cells was significantly higher than that in the reported optimal condition, and a similar tendency was observed for ARPE-19 cells. The results from enzyme-linked immunosorbent assay showed the presence of PDGF-AB, VEGF and IGF-I in the conditioned medium. The conditioned medium also promoted selective growth of human RPE cells from CNV.


CONCLUSIONS
The results from this study present the conditions for efficient and selective culture of primary RPE cells.
Kobayashi
C
C
Department of Ophthalmology, Nagoya University Graduate School of Medicine, 65 Tsuruma-cho, Showa-ku, Nagoya, Japan.
Kagami
H
H
Kito
K
K
Ishikawa
K
K
Ebisawa
K
K
Ueda
M
M
Terasaki
H
H
D016475:3T3 Cells; D000818:Animals; D002452:Cell Count; D018929:Cell Culture Techniques / Q000379:methods*; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D002478:Cells, Cultured; D002829:Choroid / Q000601:surgery; D020256:Choroidal Neovascularization / Q000378:metabolism; D018920:Coculture Techniques / Q000379:methods*; D017077:Culture Media, Conditioned / Q000494:pharmacology; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D006801:Humans; D007150:Immunohistochemistry; D008297:Male; D051379:Mice; D008875:Middle Aged; D010857:Pigment Epithelium of Eye / Q000166:cytology* / Q000378:metabolism*; D011817:Rabbits; D012160:Retina / Q000166:cytology*; D020133:Reverse Transcriptase Polymerase Chain Reaction
D017077:Culture Media, Conditioned