Physiological features of primary cultures and subcultures of human retinal pigment epithelial cells before and after cryopreservation for cell transplantation

被引:0
作者
Monika Valtink
K. Engelmann
Olaf Strauß
Renata Krüger
Cornelius Löliger
Angela Sobottka Ventura
Gisbert Richard
机构
[1] Department of Ophthalmology,
[2] Cornea Bank,undefined
[3] University of Hamburg,undefined
[4] Martinistrasse 52,undefined
[5] D-20246 Hamburg,undefined
[6] Germany Tel.: +49-40-428032334 Fax: +49-40-428034481,undefined
[7] Institute of Clinical Physiology,undefined
[8] Clinic Benjamin Franklin,undefined
[9] Freie Universität Berlin,undefined
[10] Berlin,undefined
[11] Germany,undefined
[12] Department of Transfusion Medicine and Transplantation Immunology,undefined
[13] University of Hamburg,undefined
[14] Hamburg,undefined
[15] Germany,undefined
[16] Department of Ophthalmology,undefined
[17] University of Bern,undefined
[18] Bern,undefined
[19] Switzerland,undefined
来源
Graefe's Archive for Clinical and Experimental Ophthalmology | 1999年 / 237卷
关键词
Cell Transplantation; Primary Culture; Retinal Pigment Epithelial; Retinal Pigment Epithelial Cell; Cell Culture Condition;
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摘要
· Background: One striking disadvantage of in vitro culturing of human retinal pigment epithelial (RPE) cells is the loss of epithelial differentiation and specific cell function during culture. This may be one of the main reasons for the failure of RPE cell transplantation. The aim of this study was to evaluate cell culture conditions ensuring the maintenance of differentiation and function of RPE cells after subcultivation and storage in liquid nitrogen. · Methods: Enzymatically isolated cells were seeded onto coated culture dishes, cultured with a specially formulated improved growth medium until confluence and then cryopreserved in liquid nitrogen for 16–66 months. HLA class I and II typing was performed before cryopreservation and after thawing. Expression of Ca2+ channels in primary, first-passage and cryopreserved RPE cells was studied using the patch-clamp technique. · Results: After cryopreservation no loss of any HLA antigen was detectable in 12 of 14 cell strains studied. Patch-clamp experiments demonstrated that high-threshold L-type Ca2+ channels, which are typical for freshly isolated cells, could be detected in first-passage and cryopreserved RPE cells only when improved culture conditions were employed, not in conventionally cultured cells. The characteristics of these channels showed little change in subcultured cells compared to primary cultures. · Conclusion: This is the first study showing the maintenance of adult human RPE-specific cell differentiation and characteristics in vitro after primary culture and after cryopreservation using improved cell culture methods. The optimization and quality control of cell culture is an important prerequisite for successful cell transplantation.
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页码:1001 / 1006
页数:5
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