Nancy Ransom

Ultrastructural features of retinal dystrophy in mutant vitiligo mice

Ultrastructural features of the retinal pigment epithelium and photoreceptor cells were studied in vitiligo (C57BL/6 mivit/mivit) mice. Eyes from 12-day- to 56-week-old animals were analysed. Abnormal photoreceptors were seen in 12-day-old mice. By 3 weeks malformed outer segments were evident in the posterior and equatorial retina, but normal photoreceptors were present in the periphery. By 28 weeks, a marked gradient in cell loss was evident, with a progressive increase in cell viability along the posterior-peripheral axis. Viable intact photoreceptors were still present in the peripheral retina of 56-week-old mice. Melanosome content varied between adjacent pigment epithelium cells in both the posterior and peripheral retina. In the choroid, however, a steep posterior-peripheral gradient in melanosome content was evident with highest pigmentation in the periphery. In the optic nerve head region abnormal development of photoreceptors was correlated with proliferation of abnormal pigment epithelium cells. Accumulation of rod outer segment debris in the posterior and peripheral subretinal space preceded photoreceptor cell death. Short pigment epithelial microvilli without proper attachment to photoreceptors are suggestive of alterations in pigment-epithelium-photoreceptor interaction, which might affect photoreceptor differentiation and phagocytosis of rod outer segments by pigment epithelium cells.

Alpha-fodrin is cleaved by caspase-3 in a chronic ocular hypertensive (COH) rat model of glaucoma

PURPOSE alpha-Fodrin is a neuronal cytoskeletal protein and a known caspase-3 target. We sought to determine whether caspase-3 cleaves alpha-fodrin in COH rat retinas and whether this process is reduced by adeno-associated virus (AAV)-induced retinal ganglion cell expression of baculovirus inhibitory repeat-containing 4 (BIRC4), a potent caspase-3 inhibitor. METHODS Ocular hypertension was induced unilaterally in five rat eyes by limbal injection of hypertonic saline. In a similar experiment, ocular hypertension was induced in four eyes pre-treated with an intravitreal injection of AAV-BIRC4 to assess alpha-fodrin cleavage. Western immunoblotting was performed on all retinas. RESULTS Caspase-3 cleavage of alpha-fodrin yields a specific 120kDa protein fragment. COH retina immunoblots indicated significantly more caspase-3 cleavage of alpha-fodrin than controls (P < 0.01, paired t-test). Inhibition of retinal caspase-3 activity with BIRC4 reduced caspase-3-mediated alpha-fodrin cleavage compared to controls. CONCLUSIONS This confirms our previous finding of caspase-3 cleavage of alpha-fodrin in COH retinas and parallels pathology seen in Alzheimers disease, in which neurons undergo chronic caspase activation, slow build-up of cleavage products, and delayed apoptosis. If caspase activation in glaucoma leads to protracted rather than rapid retinal ganglion cell apoptosis, a much longer therapeutic window exists for apoptosis inhibition with caspase inhibitors such as BIRC4.

APOPTOSIS OF PHOTORECEPTORS AND LENS FIBER CELLS WITH CATARACT AND MULTIPLE TUMOR FORMATION IN THE EYES OF TRANSGENIC MICE LACKING THE P53 GENE AND EXPRESSING THE HPV 16 E7 GENE UNDER THE CONTROL OF THE IRBP PROMOTER

Because of the clinical heterogeneity of human inherited retinal degenerations, we have initiated a study to seek common themes in the pathogenesis of cell death of photoreceptors by apoptosis. The interstitial retinol binding protein (IRBP) promoter was used to drive expression of the human papilloma virus 16 (HPV 16) E7 gene in the retina and other ocular tissues in mice. The result is the death of photoreceptors as they undergo terminal differentiation. Lens fiber cells also die after a period of inappropriate proliferation and abnormal differentiation to form cataracts. Cross-breeding these transgenic mice to mice lacking the p53 gene leads to formation of several ocular tumors by one month of age if both copies of the p53 gene are missing. With one copy of the gene, the mice develop retinal tumors after a much longer latency and at a lower incidence; the tumors that do arise have lost their normal copy of the gene. The lack of the p53 gene does not eliminate apoptosis of either the retina or the lens in these transgenic mice although the rate of destruction of photoreceptors is slightly delayed. The retinal tumors apparently arise from precursors that survive amid a dying cell layer.