Journal of Cell and Molecular Research

Ferdowsi University of Mashhad

Current Issue

By Issue

By Author

By Subject

Author Index

Keyword Index

About Journal

Aims and Scope

Editorial Board

Publication Ethics

Indexing and Abstracting

Related Links

FAQ

Peer Review Process

Journal Metrics

News

Guidelines

JCMR Forms

Peer Review Process at a Glance

List of Reviewers

JCMR on Publons

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

Document Type : Research Articles

Authors

1 Ferdowsi University of Mashhad, Mashhad, Iran

2 National Institutes of Genetic Engineering and Biotechnology

3 University of Social Welfare and Rehabilitation Sciences, Tehran, Iran

4 High Institute for Research and Education in Transfusion Medicine,Tehran, Iran

5 Shahid Beheshti University of Medical Sciences, Tehran, Iran

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.

Keywords

References
1. Aruta C., Giordano F., De Marzo A., Comitato A., Raposo G., Nandrot E. F. and Marigo V. (2011) In vitro differentiation of retinal pigment epithelium from adult retinal stem cells. Pigment Cell and Melanoma Research 24: 233-40.
2. Baas D., Bumsted K., Martinez J., Vaccarino F., Wikler K. and Barnstable C. (2000) The subcellular localization of Otx2 is cell-type specific and developmentally regulated in the mouse retina. Molecular Brain Research 78: 26-37.
3. Bharti K., Liu W., Csermely T., Bertuzzi S. and Arnheiter H. (2008) Alternative promoter use in eye development: the complex role and regulation of the transcription factor MITF. Development 135: 1169-78.
4. Boulton M. and Dayhaw-Barker P. (2001) The role of the retinal pigment epithelium: topographical variation and ageing changes. Eye 15: 384-9.
5. Boulton M. (2014) Studying melanin and lipofuscin in RPE cell culture models. Experimental Eye Research 126: 61-67.
6. Cai X., Conley S. M. and Naash M. I. (2009) RPE65: role in the visual cycle, human retinal disease, and gene therapy. Ophthalmic Genetics 30: 57-62.
7. Capowski E., Simonett J., Clark E., Wright L., Howden S, Wallace K., Petelinsek A., Pinilla I, Phillips M. and Meyer J. (2014) Loss of MITF expression during human embryonic stem cell differentiation disrupts retinal pigment epithelium development and optic vesicle cell proliferation. Human Molecular Genetics 23: 6332-44
8. Cheli Y., Ohanna M., Ballotti R. and Bertolotto C. (2010) Fifteen‐year quest for microphthalmia‐associated transcription factor target genes. Pigment Cell and Melanoma Research 23: 27-40.
9. Chowers I., Kim Y., Farkas R. H., Gunatilaka T. L., Hackam A. S., Campochiaro P. A., Finnemann S. C. and Zack D. J. (2004) Changes in retinal pigment epithelial gene expression induced by rod outer segment uptake. Investigative Ophthalmology and Visual Science 45: 2098-106.
10. Housset M., Samuel A., Ettaiche M., Bemelmans A., Beby F., Billon N. and Lamonerie T. (2013) Loss of Otx2 in the adult retina disrupts retinal pigment epithelium function, causing photoreceptor degeneration. The Journal of Neuroscience 33: 9890-9904.
11. Hu J. and Bok D. (2001) A cell culture medium that supports the differentiation of human retinal pigment epithelium into functionally polarized monolayers. Molecular Vision 7: 14-9.
12. Johnson S. L., Nguyen A. N. and Lister J. A. (2011) Mitfa is required at multiple stages of melanocyte differentiation but not to establish the melanocyte stem cell. Developmental Biology 350: 405-13.
13. Julien S., Kociok N., Kreppel F., Kopitz J., Kochanek S., Biesemeier A., Blitgen-Heinecke P., Heiduschka P. and Schraermeyer, U. (2007) Tyrosinase biosynthesis and trafficking in adult human retinal pigment epithelial cells. Graefe's archive for clinical and experimental ophthalmology 245: 1495-505.
14. Klimanskaya I., Rosenthal N. and Lanza R. (2008) Derive and conquer: sourcing and differentiating stem cells for therapeutic applications. Nature Reviews Drug Discovery 7: 131-42.
15. Le K., Schachter J., Quigley M. (2014) The role of melanin in protecting the skin and the retina from light damage: A comparative biological framework for Age-Related Macular Degeneration. Investigative Ophthalmology and Visual Science 55: 627-627.
16. Levy C., Khaled M. and Fisher D. E. (2006) MITF: master regulator of melanocyte development and melanoma oncogene. Trends in Molecular Medicine 12: 406-14.
17. Lu F., Yan D., Zhou X., Hu D-N. and Qu J. (2007) Expression of melanin-related genes in cultured adult human retinal pigment epithelium and uveal melanoma cells. Molecular Vision 13: 2066-72.
18. Ludwig A., Rehberg S. and Wegner M. (2004) Melanocyte-specific expression of dopachrome tautomerase is dependent on synergistic gene activation by the Sox10 and Mitf transcription factors. FEBS letters 556: 236-44.
19. Maminishkis A., Chen S., Jalickee S., Banzon T., Shi G., Wang F. E., Ehalt T., Hammer J. A. and Miller S. S. (2006) Confluent monolayers of cultured human fetal retinal pigment epithelium exhibit morphology and physiology of native tissue. Investigative Ophthalmology and Visual Science 47: 3612-24.
20. Martinez-Morales J. R., Dolez V., Rodrigo I., Zaccarini R., Leconte L., Bovolenta P. and Saule S. (2003) OTX2 activates the molecular network underlying retina pigment epithelium differentiation. Journal of Biological Chemistry 278: 21721-31.
21. Nussenblatt R. B. and Ferris III F. (2007) Age-related macular degeneration and the immune response: implications for therapy. American Journal of Ophthalmology 144: 618-26.
22. Panda-Jonas S., Jonas J. B. and Jakobczyk-Zmija M. (1996) Retinal pigment epithelial cell count, distribution, and correlations in normal human eyes. American Journal of Ophthalmology 121: 181-9.
23. Pfeffer B. and Philp N. (2014) Cell culture of retinal pigment epithelium: Special Issue. Experimental Eye Research 126: 1-4.
24. Rak D. J., Hardy K. M., Jaffe G. J. and McKay B. S. (2006) Ca++-switch induction of RPE differentiation. Experimental Eye Research 82: 648-56.
25. Raviv S., Bharti K., Rencus-Lazar S., Cohen-Tayar Y., Schyr R., Evantal N., Meshorer E., Zilberberg A., Idelson M., Reubinoff B. (2014) PAX6 regulates melanogenesis in the retinal pigmented epithelium through feed-forward regulatory interactions with MITF. PLoS Genet. 29: 10.
26. Reinisalo M., Putula J., Mannermaa E., Urtti A. and Honkakoski P. (2012) Regulation of the human tyrosinase gene in retinal pigment epithelium cells: the significance of transcription factor orthodenticle homeobox 2 and its polymorphic binding site. Molecular Vision 18: 38.
27. Schraermeyer U., Kopitz J., Peters S., Henke-Fahle S., Blitgen-Heinecke P., Kokkinou D., Schwarz T. and Bartz-Schmidt K. U. (2006) Tyrosinase biosynthesis in adult mammalian retinal pigment epithelial cells. Experimental Eye Research 83: 315-21.
28. Simeone A., Acampora D., Gulisano M., Stornaiuolo A. and Boncinelli E. (1992) Nested expression domains of four homeobox genes in developing rostral brain. Nature 358: 687-90.
29. Strunnikova N., Maminishkis A., Barb J., Wang F., Zhi C., Sergeev Y., Chen W., Edwards A.O., Stambolian D., Abecasis G., Swaroop A., Munson P. J. and Miller S. S. (2010) Transcriptome analysis and molecular signature of human retinal pigment epithelium. Human Molecular Genetics 19: 2468-86.
30. Takeda K., Yokoyama S., Yasumoto K-i., Saito H., Udono T., Takahashi K. and Shibahara S. (2003) OTX2 regulates expression of DOPAchrome tautomerase in human retinal pigment epithelium. Biochemical and Biophysical Research Communications 300: 908-14.
31. Tsukiji N., Nishihara D., Yajima I., Takeda K., Shibahara S. and Yamamoto H. (2009) Mitf functions as an in ovo
regulator for cell differentiation and proliferation during development of the chick RPE. Developmental Biology 326: 335.
32. Yasumoto K., Yokoyama K., Takahashi K., Tomita Y. and Shibahara S. (1997) Functional analysis of microphthalmia-associated transcription factor in pigment cell-specific transcription of the human tyrosinase family genes. Journal of Biological Chemistry 272: 503-9.
33. Zarbin M., Casaroli-Marano R. and Rosenfeld P. (2014) Age-related macular degeneration: Clinical findings, histopathology and imaging techniques. Developments in Ophthalmology 53: 1-32.
Send comment about this article
CAPTCHA Image
Journal of Cell and Molecular Research
Volume 7, Issue 2 - Serial Number 14
December 2015
Pages 76-85
Files
History
  • Receive Date: 03 May 2015
  • Accept Date: 03 May 2015
Share
How to cite
Statistics