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Jorgelina Muriel Calandria, Khanh Do, Nicolas Guillermo Bazan; DHA-NPD1 signaling via cREL downregulates pro-inflammatory Wnt5a in RPE cells. Invest. Ophthalmol. Vis. Sci. 2017;58(8):1050.
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© ARVO (1962-2015); The Authors (2016-present)
Neuroprotectin D1 (NPD1) inhibits the expression of pro-inflammatory genes and enhances the expression of anti-apoptotic proteins of the Bcl-2 family (Mukherjee et al. 2004, PNAS 101:8491 ) that, along with other translational and post-translational events, become instructive signals that decide RPE cell fate. Wnt5a, a secreted protein from the Wnt family of ligands that promote cell fate specification, was found to be involved in homeostatic circuitry activation of NFkB during innate immune response (Naskar et al, 2014. J Immunol 192:4386). We hypothesized that DHA/NPD1 signaling regulates Wnt5a expression and activity. Thus we studied here NPD1 bioactivity in the regulation of the expression and release of Wnt5a.
cREL was silenced or overexpressed in human RPE cells, and Wnt5a mRNA was measured using SYBR green real-time PCR assay. The time course of Wnt5a secretion was tested by Western blot in medium and hRPE cells undergoing uncompensated oxidative stress (UOS) triggered by 1200 µM H2O2 and 10 ng/ml TNFα, in the presence or absence of NPD1. The role of FZD5 and ROR2 was assessed by silencing the co-receptors and measuring the activity of a NF-κB luciferase reporter.
The silencing of cREL resulted in upregulation of Wnt5a transcription, and the overexpression of the transcription factor elicited the opposite effect, suggesting that NPD1 is regulating Wnt5a via cREL. Wnt5a was released from cells undergoing UOS only in the absence of NPD1. The peak for Wnt5a release was observed at 10 hours after the onset of UOS. NF-κB induction upon addition of Wnt5a was altered by the combination of FZD5 and ROR2 silencing, but not separately. NPD1 selectively halts the increase of FZD5 expression triggered by UOS.
The evidence presented here points to a systemic inflammatory role of Wnt5a in which expression is controlled by NPD1 via cREL. Wnt5a is released and is involved in a positive feedback loop that leads to inflammatory signaling amplification. The mechanism proposed here may be a target of new therapies aiming to decrease proinflammatory processes in diverse conditions, including age-related macular degeneration (AMD).
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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