Abstract
Purpose: :
The rd1 mouse is among the best characterized animal models of Retinitis Pigmentosa, distinguished by early–onset degeneration of rod photoreceptors beginning around postnatal day 10 (P10). A mutation in the cGMP phosphodiesterase beta gene results in cGMP accumulation leading to cell death. Cellular and morphological alterations consistent with a delay in photoreceptor development have been reported prior to cell death. In addition to its role in phototransduction, cGMP is an important second messenger. Little is known about the role of cGMP in developing photoreceptors. Here we investigate differential gene expression in the rd1 mouse retina during early photoreceptor differentiation: from P2, prior to any identified morphological or biochemical differences, through P8, when early pathological changes become apparent.
Methods: :
Retinas from P2, P4, P6, and P8 wild type and rd1 mice on a C57BL/6 background were rapidly isolated and frozen. Affymetrix mouse GeneChips were used to evaluate gene expression with samples of 8–14 retinas run in quadruplicate at each time point. Selected genes were verified using real time PCR. Selected proteins were analyzed by Western blot and localized by immunohistochemistry.
Results: :
Comparative data analysis produced 162 probesets differentially expressed for at least one of the four time points. Of these genes, 27 were differentially expressed at more than one time point. Differential gene expression is apparent much earlier than previously known with Pde6b expression reduced by 4.5–fold at P2. Genes with functions in cell structure, transcription, and signaling transduction are downregulated at P2. Other genes, including some known to play a role in neuronal differentiation, are also downregulated at early time points.
Conclusions: :
Our results provide clues to understanding molecular mechanisms involved in photoreceptor differentiation and degeneration. This data may help us to unravel the role of cGMP signaling pathways prior to its role in phototransduction.
Keywords: gene microarray • retinal degenerations: cell biology • retinal development