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Establishment of a lens epithelial cell line from cataract dog
Author(s) -
OCHIAI H
Publication year - 2011
Publication title -
acta ophthalmologica
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.534
H-Index - 87
eISSN - 1755-3768
pISSN - 1755-375X
DOI - 10.1111/j.1755-3768.2011.246.x
Subject(s) - transfection , western blot , cell culture , ascorbic acid , biology , microbiology and biotechnology , capsulorhexis , phacoemulsification , medicine , ophthalmology , gene , genetics , food science , visual acuity
Purpose The aim of this study is to establish a lens epithelial cells (LECs) line originated from cataract dog. Methods Anterior capsulorrhexis specimen from a dog (8‐year‐old male Wire Fox Terrier) naturally developing mature cataracts was obtained prior to routine phacoemulsification cataract extraction. The primary lens epithelial cells were transfected with the expression plasmid DNA encoding replication origin defective simian virus 40 (SV40) large T antigen and then cloned a colony by using glass syrinder. Results The primary cells proliferated to confluent until three passages. However, the immortalized cells remained proliferative, and this cloned cell line, termed as cdLEC, grew well and could be propagated over 200 times by splitting at 1:20. Functional analysis of Na‐dependent vitamin C transporter (SVCT) indicated that the Km value toward ascorbic acid (vitamin C) was 19.9 + 2.8 M, and RT‐PCR analysis showed that SVCT2 was observed in this cell line while SVCT1 was not, which was the characteristic of LECs. Western blot analysis and cyto‐immunochemistry indicated immortalized cells produced a protein with a molecular weight of 25 kDa, which reacted with an antibody to B‐crystallin within the whole cytosol. Conclusion These results indicate that cdLEC may provide a useful in vitro system for the study of path‐physiology of canine cataract.