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Down-regulation of microRNA-216a confers protection against yttrium aluminium garnet laser-induced retinal injury via the GDNF-mediated GDNF/GFRα1/RET signalling pathway

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Abstract

Retinal injury plays a leading role in the onset of visual impairment. Current forms of treatment are not able to ameliorate scarring, cell death and tissue and axon regeneration. Recently, microRNA-216a (miR-216a) has been reported to regulate snx5, a novel notch signalling pathway component during retinal development. This study aims to elucidate the role of miR-216a in yttrium aluminium garnet (YAG) laser-induced retinal injury by targeting glial cell line-derived neurotrophic factor (GDNF) via GDNF/GDNF family neurotrophic factor receptor α1 (GFRα1)/rearranged during transfection (RET) signalling pathway. Wistar male rats were first randomly assigned into control and model groups. Immunohistochemistry was performed to detect the GDNF positive expression rate and terminal deoxynucleotidyl transferase-mediated dUTP nick end labelling (TUNEL) staining for apoptotic index (AI) of retinal tissue. Retinal neurons were divided into normal, blank, negative control (NC), miR-216a mimic, miR-216a inhibitor, siRNA-GDNF and miR-216a inhibitor+siRNA-GDNF groups. Dual luciferase reporter assay was conducted in order to identify the targeting relationship between GDNF and miR-216a. Reverse transcription quantitative polymerase chain reaction (RT-qPCR) and western blot were used for the analysis of mRNA and protein levels of miR-216a and related genes. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was used to determine cell proliferation and flow cytometry was used to observe cell cycle and apoptosis. Results show that the model group had an increased GDNF positive rate, AI of retinal tissue and mRNA and protein levels of cellular oncogene fos (c-fos), vascular endothelial growth factor (VEGF), brain-derived neurotrophic factor (BDNF), GDNF, GFRα1 and bcl-2-associated X protein (bax), declined miR-216a level and mRNA and protein levels of RET and bcl-2 compared with the control group. GDNF was verified as the target gene for miR-216a. Compared with the blank and NC groups, the miR-216a mimic and siRNA-GDNF groups had higher mRNA and protein levels of c-fos, VEGF and bax, cell number in the G1 phase and increased cell apoptosis but reduced BDNF, GDNF, GFRα1, RET and bcl-2 expression, cell proliferation and cell numbers in the S phase, while the opposite trend was observed in the miR-216a inhibitor group. Taken together, our findings demonstrate that elevated GDNF levels can reduce the retinal injury, whereby down-regulated miR-216a aggravates the YAG laser-induced retinal injury by targeting the GDNF level through the GDNF/GFRα1/RET signalling pathway.

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Acknowledgments

We are grateful to the reviewers for helpful comments on this paper.

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Authors and Affiliations

  1. Department of Ophthalmology, Jiangxi Pingxiang People’s Hospital, Pingxiang, 337055, People’s Republic of China

    Xi-Bin Hu, Jian-Zhong He, Yan-Fei Qiu, Wei Lai & Min Zhong

  2. Department of Ophthalmology, The Second Affiliated Hospital of Nanchang University, Nanchang, 330006, People’s Republic of China

    Shu-Hua Fu

  3. Department of Histology and Embryology, Nanchang University School of Basic Medical Sciences, Nanchang, 330006, People’s Republic of China

    Qi Luo

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  1. Xi-Bin Hu
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  2. Shu-Hua Fu
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  3. Qi Luo
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Correspondence to Jian-Zhong He.

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Communicated by Aurnab Ghose.

Corresponding editor: Aurnab Ghose

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Hu, XB., Fu, SH., Luo, Q. et al. Down-regulation of microRNA-216a confers protection against yttrium aluminium garnet laser-induced retinal injury via the GDNF-mediated GDNF/GFRα1/RET signalling pathway. J Biosci 43, 985–1000 (2018). https://doi.org/10.1007/s12038-018-9795-9

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  • DOI: https://doi.org/10.1007/s12038-018-9795-9

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