Sirt1 Involvement in Mouse and Human Retinal Development and Function

Abstract

Purpose: : Recent evidence suggests that SIRT1 may play a significant role in development of the central nervous system as well as during pathological neurodegeneration. The purpose of this study was to investigate the expression of SIRT1 in developing, adult and degenerating mouse and human retinas and to study the effects of SIRT1 knockdown on retinal development genes.

Methods: : Immunostaining for SIRT1 was performed on the following tissues: two post-natal day 1 (P1) mouse eyes, 14 normal adult B57BL6 mouse eyes (P41-P347), 20 Rhodopsin-knockout (RHO^-/-) mouse eyes (P42-P336), two human fetal eyes (10 weeks gestation), six normal adult human eyes from four donors, and peripheral retinal sections from three human retinitis pigmentosa (RP) patients. Knockdown of SIRT1 via siRNA in mouse retinal progenitor cells (mRPCs) was performed followed by PCR to evaluate changes in the following retinal development genes: Nestin, PAX6, CRX, and Nrl.

Results: : Fetal human, adult human, and adult mouse retinas all had cytoplasmic staining for SIRT1 in all layers, with the exception of P176 mouse eyes, which were negative in the ONL and photoreceptors. Additionally, P1 mouse eyes had two distinct nuclear layers, with the inner layer staining positive and the outer layer negative. Two of the three human RP cases showed staining in remaining retinal layers, while the third patient showed staining only in the inner plexiform and ganglion cell layers. RHO^-/- mouse eyes demonstrated expression of SIRT1 in all remaining retinal layers at all time points, with the exception of the P302 eyes which revealed a predominantly negative inner nuclear layer with sparse positive cells. SIRT1 siRNA studies showed significant decreases in CRX, PAX6 and Nestin in cells treated with SIRT1 siRNA compared to controls. No significant changes were observed in Nrl expression.

Conclusions: : Expression of SIRT1 in the developing and adult retinas, coupled with siRNA results from mRPCs, suggests a probable role for this protein in normal retinal development and function. The possible role of SIRT1 in retinal degeneration is less clear and warrants further investigation.

Keywords: retinal development • retinal degenerations: cell biology • microscopy: light/fluorescence/immunohistochemistry