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Rachel L. Harding, Scott Hammond, David Hyde; miRNAs Regulate Retinal Regeneration in Zebrafish. Invest. Ophthalmol. Vis. Sci. 2011;52(14):2249.
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Damage of the adult zebrafish retina induces the Muller glia to begin proliferating, which leads to regeneration of the damaged neurons. We asked if microRNAs (miRNAs), which regulate translation of target genes by basepairing with the corresponding mRNAs, played a role in coordinating this regenerative response. Because the enzyme Dicer cleaves the pre-miRNAs to their mature 22 bp length, we examined if Dicer, and by extrapolation miRNAs, were required for regenerating photoreceptors in the light-damaged retina.
Electroporation of intravitreally injected anti-dicer morpholinos was used to knockdown Dicer protein expression and was confirmed by immunoblots. Immunohistochemistry of PCNA expression was used to determine the extent of Muller glia proliferation in the light-damaged morphant and control retinas. A microarray of zebrafish miRNAs was conducted to determine what miRNAs increased in expression during the light-damage paradigm and would be candidates for regulating retinal regeneration. The expression profiles of candidate miRNAs were verified by qRT-PCR.
Injection of dicer morpholinos prior to the start of light treatment statistically reduced the number of PCNA-positive Muller glia in the regenerating retinas relative to control retinas beginning at 51 hours of light treatment. A microarray identified 106 miRNAs that increased in expression during the light treatment, making them candidates to be affected in the dicer morphant retina. The expression profiles of two of these miRNAs, which putatively regulate the expression of candidate proteins known to be involved in retinal regeneration, were verified by qRT-PCR.
This data demonstrates that miRNAs are required for regeneration of photoreceptors in the light-damaged zebrafish retina. Additionally, several miRNAs were identified that may regulate expression of candidate proteins that are essential for retinal regeneration. These individual miRNAs will be further examined to determine their specific function during retinal regeneration.
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