June 2017
Volume 58, Issue 8
Open Access
ARVO Annual Meeting Abstract  |   June 2017
MicroRNA-302d promotes dedifferentiation of the retinal pigment epithelium by targeting the CDKN1A
Author Affiliations & Notes
  • chao jiang
    Jiangsu Province People's Hospital, NANJING, JIANGSU , China
  • Xue Chen
    Jiangsu Province People's Hospital, NANJING, JIANGSU , China
  • Chen Zhao
    Jiangsu Province People's Hospital, NANJING, JIANGSU , China
  • Footnotes
    Commercial Relationships   chao jiang, None; Xue Chen, None; Chen Zhao, None
  • Footnotes
    Support  2013CB967500 to Chen Zhao); National Natural Science Foundation of China (81525006); Jiangsu Province’s Innovation Team (to Chen Zhao); the Fundamental Research Funds of the State Key Laboratory of Ophthalmology (to Chen Zhao); and A Project Funded by the Priority Academic Program Development of Jiangsu Higher Education Institutions (PAPD).
Investigative Ophthalmology & Visual Science June 2017, Vol.58, 5358. doi:
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    • Get Citation

      chao jiang, Xue Chen, Chen Zhao; MicroRNA-302d promotes dedifferentiation of the retinal pigment epithelium by targeting the CDKN1A
      . Invest. Ophthalmol. Vis. Sci. 2017;58(8):5358.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose : MicroRNA plays a critical role in dedifferentiation of retinal pigment epithelium (RPE) cells. In our previous studies, we found the hsa-miR-302d-3p was down-regulated in RPE differentiation process. However, the microRNA mediated dedifferentiation in human RPE cells has not been thoroughly reported. In our study, we aim to analyze the role of miR-302d in RPE dedifferentiation

Methods : We respectively transfected hsa-miR-302d mimic and inhibitor into hiPSC-RPE at 30 day to see its role on cell differentiation. The related rpe genes and proteins expression in the rpe differentiation process were observed by Q-PCR and western blotting. We also observe the expression of ZO-1 in ips-rpe cells according to immunofluorescence. We next determined whether miR-302d would promote proliferation and migration in ARPE-19 cells. Furthermore, cell cycle were observed by flow cytometry after transfected hsa-miR-302d mimic and inhibitor. We confirm that CDKN1A as a direct target of miR-302d in ARPE-19 cells. Also, we established an AMD model for the treatment of hydrogen peroxide with H2O2. We observed the phagocytosis and apoptosis of RPE cells after transfected hsa-miR-302d mimic and inhibitor

Results : We found relative mRNA expressions were significantly decreased of RPE65, RLBP1, MERTK, BEST1, CTNNB1 and TJP1 in hiPS-RPE at 30 days after transfected with miR-302d mimic compared to NC mimic. Also the relative mRNA expressions were significantly increased of RPE65, RLBP1, MERTK, BEST1, and TJP1 in hiPSC-RPE at 30 days transfected with miR-302d inhibitor compared to NC inhibitor. Both proliferation and migration were promoted by miR-302d overexpression after transfection with ARPE19 cells. And the luciferase reporter gene assay showed that CDKN1A is the target gene of microRNA302d. After transfected with miR-302d minics and inhibitor, we then treated the cells with 200uM H2O2. We found that with the oxidative stress model, the cell apoptosis rates significantly increased after transfected with miR-302d inhibitor compared to NC inhibitor

Conclusions : Mir-302d is associated with RPE cells proliferation and migration as well as apoptosis under oxidative stress, indicating that Mir-302d negatively regulates RPE cells differentiation.

This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.

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