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Pranab K Mukherjee, Nicolas G Bazan; Sirtuin1 (SIRT1) is a Neuroprotectin D1 (NPD1) target in retinal pigment epithelial cell survival signaling. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):3618.
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© ARVO (1962-2015); The Authors (2016-present)
SIRT1 belongs to a family of highly conserved proteins that has been linked to caloric restriction beneficial outcomes and aging by modulating energy metabolism, genomic stability and stress resistance. Sirtuin1 is a potential therapeutic target in a variety of diseases including cancer, diabetes, inflammatory disorders and neurodegenerations. NPD1, a DHA-derived mediator, induced cell survival through the upregulation of the BCL2 class of survival proteins and downregulation of pro-apoptotic Bad and Bax under oxidative stress (OS) in RPE cells. The purpose of this research is to test whether NPD1 modulates SIRT1 in RPE cells under OS.
We have used ARPE-19 and primary human RPE cells. Cells grown for 72h were serum-starved for 8h. Then cells were treated with TNF-α (10ng/ml) plus H2O2 (600µM) to induce OS and were further incubated with 50 to 500nM concentrations of NPD1 for 12h (apoptosis) and 6, 12, and 24h. Primary human RPE cells were exposed to TNF-α (10ng/ml) plus H2O2 (100µM). Cell extracts were prepared, protein contents were adjusted, and SIRT1 protein was detected by Western blot using anti-SIRT1 antibody. Hoechst and Tunnel staining was used to score apoptotic cells.
NPD1 at 50 to 500 nM concentrations upregulated SIRT1 abundance in ARPE-19 cells and in primary human RPE cells undergoing OS. However, OS or NPD1 alone had little or no effect on the SIRT1 expression. Moreover, the NPD1-mediated induction of SIRT1 is time- and concentration-dependent. The upregulation of SIRT1 by NPD1 peaked 6h after OS onset, decreased at 12h, and plateaued at 24h in ARPE-19 cells. This enhancement peaked at 100nM, after which it started to decline. Additionally, upregulation of SIRT1 is specific for NPD1 since nine other structurally-related lipid mediators were devoid of bioactivity on SIRT1 under similar conditions in RPE cells. Moreover, the NPD1-mediated upregulation of SIRT1 attenuated OS-induced apoptosis in RPE cells.
NPD1 stereoselectively upregulates SIRT1 abundance in human RPE cells when confronted with OS. As a consequence, remarkable cell survival takes place. This new target for the lipid mediator allows us to propose an integrated DHA/NPD1 lipidome survival signaling that results in enhanced photoreceptor cell integrity.
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