Microarray analysis of iris gene expression in mice with mutations influencing pigmentation

Abstract

Purpose: Several ocular diseases involve the iris, notably including oculocutaneous albinism, pigment dispersion syndrome, and exfoliation syndrome. To screen for candidate genes that may contribute to the pathogenesis of these diseases, genome-wide iris gene expression patterns were comparatively analyzed from mouse models of these conditions.

Methods: Iris samples from albino mice with a Tyr mutation, pigment dispersion-prone mice with Tyrp1 and Gpnmb mutations, and mice resembling exfoliation syndrome with a Lyst mutation were compared with samples from wild-type mice. All mice were strain (C57BL/6J), age (60 days old), and sex (female) matched. Microarrays were used to compare transcriptional profiles, and differentially expressed transcripts were described by functional annotation clustering using DAVID Bioinformatics Resources. Quantitative real-time PCR was performed to validate a subset of identified changes.

Results: Compared with wild-type C57BL/6J mice, each disease context exhibited a large number of statistically significant changes in gene expression, including 685 transcripts differentially expressed in albino irides, 403 in pigment dispersion-prone irides, and 460 in exfoliative-like irides.

Conclusions: Functional annotation clusterings were particularly striking among the overrepresented genes, with albino and pigment dispersion-prone irides both exhibiting overall evidence of crystallin-mediated stress responses. Exfoliative-like irides from mice with a Lyst mutation showed overall evidence of involvement of genes that influence immune system processes, lytic vacuoles, and lysosomes. These findings have several biologically relevant implications, particularly with respect to secondary forms of glaucoma, and represent a useful resource as a hypothesis-generating dataset.

Figure 1.

Iris phenotypes of wild-type C57BL/6J and albino B6.Tyr^c-2J mice. Slit lamp images of eyes with broad-beam (rows 1, 3) and transilluminating (rows 2, 4) light. (A–C) At all ages, wild-type C57BL/6J irides had a smooth-appearing surface accentuated by numerous underlying vessels and a uniformly deep sienna-brown color. (D–F) With transilluminating illumination, C57BL/6J irides appeared black at all ages, indicating an intact healthy iris (the bright white circle is a reflection of the photographic flash and not an iris defect). (G–I) At all ages, B6.Tyr^c-2J irides had a complete lack of melanin pigment, but otherwise remained intact. (J–L) With transilluminating illumination, B6.Tyr^c-2J irides freely passed light across most areas. Because it is not transparent, the iridial vasculature was prominently visible.

Figure 2.

Iris phenotypes of pigment dispersion–prone B6.Tyrp1^b Gpnmb^R150X and exfoliative-like B6-Lyst^bg-J mice. Slit lamp images of eyes with broad beam (rows 1, 3) and transilluminating (rows 2, 4) light. (A, B) Through 5 months of age, the irides of B6.Tyrp1^b Gpnmb^R150X mice were very similar to wild-type. (C) With increasing age, the pigment-dispersing iris disease in B6.Tyrp1^b Gpnmb^R150X mice was evident by the presence of dispersed pigment across the iris, giving it a granular appearance, and within the pupil. (D, E) With transilluminating light, B6.Tyrp1^b Gpnmb^R150X irides from young mice showed mild transillumination defects (red areas). (F) With increasing age, the transillumination defects of B6.Tyrp1^b Gpnmb^R150X mice became more apparent as iris atrophy accompanied pigment dispersion. (G–I) As a consequence of an early-onset degenerative disease, the iris of B6-Lyst^bg-J mice appeared dark and granular. As observable in (I), cataracts were also common in B6-Lyst^bg-J eyes. (J–L) With transilluminating light, B6-Lyst^bg-J irides exhibited a distinct pattern of transillumination defects occurring in exfoliation syndrome characterized by concentric rings of transillumination.