Acyclovir Attenuates Stress-Induced miRNA-146a Levels in Human Retinal Pigment Epithelial (RPE) Cells

Abstract

Purpose: : Retinal pigment epithelial (RPE) cells rely on a defined population of micro RNAs (miRNAs) to shape their homeostatic patterns of gene expression. In these studies we analyzed the effects of physiologically relevant cytokine stressors (TNF alpha and IL-1 beta) on miRNA speciation in human RPE cells to study what specific gene expression pathways were altered. As acyclovir has been previously reported to quench elective pro-inflammatory miRNA signaling (Lukiw et al., Neuroreport 21:922-927, 2010), we also tested the effects of this potent acyclic nucleoside anti-viral drug under the same experimental conditions.

Methods: : ARPE-19 cell culture; IL-1 beta- and/or TNF alpha-induced stress; bioinformatics; DNA array; human retinal tissue; human retinal and brain cells in primary culture; gel shift assay; micro-RNA array; Northern micro-dot blot analysis; RT-PCR; transfection assay; Western immunohistochemistry

Results: : In IL-1 beta- and/or TNF alpha-stressed human RPE cells we identified several retinal-abundant miRNA species that are consistently up-regulated including the pro-inflammatory miRNA-146a species. Up-regulation of this inducible, NF-kB-regulated, 22 nucleotide small non-coding RNA (sncRNA) is associated with altered innate-immune responses in stressed human brain cells in Alzheimer’s and prion-linked neurodegenerative disease and in age-related macular degeneration (AMD) [p<0.01, ANOVA]. Acyclovir was found to quench this effect restoring miRNA-146a back to homeostatic levels.

Conclusions: : Dysfunctional inflammatory signaling in RPE cells appears to be involved in a number of progressive human retinal disorders including AMD. These data provide the first evidence of altered miRNA signaling in cytokine-stressed human RPE cells. Dissection of the interactive molecular contributions of NF-ΚB-, and miRNA-146a-mediated signaling mechanisms in stressed RPE cells should provide further therapeutic insight into potential inhibitors that will be useful in retinal cell pharmacotherapy and RPE-associated disease.

Keywords: retinal pigment epithelium • cytokines/chemokines • stress response