Expression of melanin-related genes in cultured adult human retinal pigment epithelium and uveal melanoma cells.
OBJECTIVE
Controversy exists over melanogenesis of adult human RPE cells in vitro. This study investigated melanin content and production and expression of tyrosinase (TYR), tyrosinase-related-protein-1 (TRP1), tyrosinase-related-protein-2 (TRP2), and P gene in cultured human RPE cells and uveal melanoma cells.


METHODS
RPE cells were isolated and cultured from three adult donor eyes. A continuous human uveal melanoma cell line was established from primary choroidal melanoma. Melanin content and production were measured, and the expression of TYR, TRP1, TRP2, and P gene at the mRNA and protein levels were detected by RT-PCR and western blot, respectively.


RESULTS
Melanin content per cell of cultured RPE decreased rapidly and in proportion to cell division. No melanin production could be demonstrated in any passages. In cultured RPE cells, mRNA expression of TYR, TRP1, TRP2, and P-gene and protein expression of TYR, TRP1, and TRP2 could not be detected. In uveal melanoma cells, melanin content per cell remained stable, and melanin production could be detected in each passage. Expression of mRNA of TYR, TRP1, TRP2, and P-gene and protein of TYR, TRP1, and TRP2 could be detected in melanoma cells.


CONCLUSIONS
Human RPE cells under standard culture circumstances do not produce melanin and do not express the four key genes required in melanin biosynthesis pathway. In contrast, human uveal melanoma cells produce melanin and express all of these melanogenic genes in vitro.
Lu
Fan
F
Eye Hospital, School of Ophthalmology and Optometry, Wenzhou Medical College and Key Laboratory of Vision Science, Ministry of Health PR China, Wenzhou China.
Yan
Dongsheng
D
Zhou
Xiangtian
X
Hu
Dan-Ning
DN
Qu
Jia
J
D000328:Adult; D015153:Blotting, Western; D002478:Cells, Cultured; D015870:Gene Expression; D006801:Humans; D008543:Melanins / Q000096:biosynthesis* / Q000378:metabolism; D008545:Melanoma / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D026901:Membrane Transport Proteins / Q000235:genetics; D014442:Monophenol Monooxygenase / Q000235:genetics / Q000378:metabolism; D010857:Pigment Epithelium of Eye / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014357:Trypsin; D014362:Trypsinogen / Q000235:genetics / Q000378:metabolism; D014604:Uveal Neoplasms / Q000378:metabolism*
D008543:Melanins; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; C489600:OCA2 protein, human; D012333:RNA, Messenger; C113139:Trp2 protein, vertebrate; D014362:Trypsinogen; D014442:Monophenol Monooxygenase; C489303:PRSS1 protein, human; D014357:Trypsin