Investigation of Melanogenic Factors Gene Expression in Human Adult and Neonate Retinal Pigment Epithelium Cell Cultures

Abstract

Retinal pigment epithelium is responsible for maintaining the structural integrity of the retina by an efficient defense against free radicals, photo-oxidative exposure and light energy. For this purpose the main RPE line of defense is melanosomes. Melanin content of retinal pigment epithelium cells in adults and neonates reveals remarkable variations. In the current study we compared melanogenic factors gene expression levels in human adult and neonate RPE cells in culture. RPE cells from adult and neonate human eye globes were isolated and then cultured in DMEM: F12 (1:1) containing 10% FBS. Expression of RPE65 and Cytokeratin 8/18 markers in isolated cells was confirmed by immunocytochemistry. To evaluate the responsible factors in the pathway of melanin biosynthesis, gene expression of orthodenticle homeobox 2, microphthalmia-associated transcription factor A and H as prominent transcription factors, tyrosinase and dopachrome tautomerase as effective enzymes in the melanin biosynthetic pathway were examined in human neonate derived RPE cells compared to human adult derived RPE cells in culture. According to the Real-Time RT-PCR data, gene expression of MITF-H, OTX2, TYR and DCT in RPE cells of the neonates showed a significant increase compared to the adults. With increasing passage number, gene expression of MITF-H, OTX2, TYR and DCT showed remarkable decline. According to the role of OTX2 and MITF-H as the main transcription factor effectors on the TYR and DCT, restoration of OTX2 and MITF-H gene expression may be retain melanin content in RPE cells.