A Strange Affair

Jim Jarmusch's new film, The Dead Don't Die, gnaws at some Then again, Jarmusch is also making fun of this cute little town and its.

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Knox3, Edward N. Pugh, Jr. Electron microscopy was applied to fixed retinal tissue to assess cone morphology. Cone function was assessed with single cone recordings sunction pipette recording. Immunohistochemistry methods were employed to investigate S-opsin distribution throughout the cone cell. However our results indicate that the mutant S-opsin is less efficient at attaining a properly folded mature form in the ER, and a large fraction of nascent S-opsin is degraded by ERAD.

Our cis retinal supplementation results suggest that chromophore binding of nascent cone opsins is critical for efficient biosynthesis and export from the ER. Commercial Relationships: Lauren L. Nikonov, None; Barry E.

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Knox, None; Edward N. Purpose: Rabs are small GTP-binding proteins that regulate intracellular vesicular traffic. There are 60 known mammalian Rabs, some of which regulate general steps and are expressed ubiquitously, while others perform a specialised function and are expressed in a more restricted pattern. Rab38 was shown in the past to be expressed in pigmented cells such as melanocytes and retinal pigment epithelium RPE and be important for the delivery of tyrosinase to melanosomes. Deficiency in Rab38 leads to a significant reduction in a number of mature melanosomes in the RPE of the Rab38 mutant mice chocolate.

Conclusions: Rab38 is important for rhodopsin degradation. Cathepsin D is severely reduced in the absence of Rab38 which suggests that Rab38 might be involved in cathepsin D trafficking.

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Purpose: MerTK-mediated phagocytosis is crucial for professional phagocytes, e. This study was planned to test how MerTK affected phagosome uptaking; and whether the MerTK mutant phagocyte was able to uptake the large targets, if yes, by what mechanism.

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  • Methods: The macrophage and RPE were isolated from wild-type and MerTK knockout mice and cultured in vitro for 24 hours before the assays. The time course of the FITC-labeled apoptotic lymphocytes was compared between two groups. The pulling strength generated by cytoskeletal and dynamic proteins during engulfment and phagosome formation was measured and analyzed by atomic force microscope AFM.

    Results: The cultured MerTK mutant macrophages were able to uptake apoptotic lymphocytes, although at a significant low level compared to the wild-type counterparts. However, they adapted a macropinocytotic mechanism. In the wildtype phagocytes, the phagocytic cup formation was initiated by denting the membrane at the cell-cell contacting region and protruding the peripheral membrane to sheathe the target particle, during which process the inner membrane of the growing phagocytic cup was in close contact with the target surface.

    On the other hand, the MerTK mutant phagocytes ate the targets by membrane ruffling, and rolling the particles with membrane wave until engulfment was completed, a typical manifestation of macropinocytosis as described by Swanson et al. The AFM measurement of the particle-pulling strength suggested that the MerTK-mediated phagocytosis generated a vertical pulling force, which was necessary for up-taking the engulfed particles.

    Whereas the MerTK mutant cells decreased this type of vertical pulling power, but instead, generated Copyright by the Association for Research in Vision and Ophthalmology, Inc. Conclusions: Our study showed that the MerTK mutant cells were still able to uptake the targets although with inefficacy. The MerTK mutant cells preserved the macropinocytosis mechanism for enclosing the large bound-particles. While the wild-type phagocytes generated a vertical power for pulling the bound targets into the cytoplasm, the mutants were defective, but replace with a dominant horizontal pushing force to close-in the bound particles.

    Weber3, Eberhart Zrenner1, M. Dominik Fischer1,4. Purpose: X-linked juvenile retinoschisis XLRS is the most common juvenile maculopathy in men and is caused by mutations in the gene encoding retinoschisin RS1. Evidence in the literature on the therapeutic effect of carboanhydrase inhibitors CAIs to treat schisis formation in the retina has remained equivocal. Changes of retinal structure were monitored by confocal scanning laser ophthalmoscopy and spectral domain optical coherence tomography at the age of 4, 6, 8, 12 weeks and 7 months.

    Changes in inner retinal thickness IRT, as measure of schisis formation and outer retinal thickness ORT, as measure of photoreceptor degeneration were quantified at all time-points. ERG was performed at the age of 8 and 12 weeks. At each time point two mice were sacrificed for histology. Results: Schisis formation peak at 3 months preceded photoreceptor degeneration and hyperfluorescence peak at 7 months. Structural pathology was most severe in the superior hemiretina.

    Quantitative analysis showed no significant differences regarding the inner or outer retinal thickness of the treated vs. Schisis formation in the inner retina peaked at the age of 3 months and was followed by photoreceptor degeneration and hyperfluorescence suggesting photoreceptor debris accumulation. Schisis formation and hyperfluorescent spots were more severe in the superior hemiretina. Chapman2, Simon Kaja1, Stephanie L. Purpose: Anandamide, N-Arachidonoylethanolamine, as the most extensively studied N-Acylethanolamine NAE , is an endogenous ligand for cannabinoid and vanilloid receptors.


    In response to injury, NAEs become upregulated to exert their protective function as lipid signaling molecules. With little knowledge available about the function and physiological targets of many NAEs, such as Npalmitoylethanolamine NAE , N-lauroylethanolamine NAE and Nlinoleoylethanolamine NAE , we measured their protective properties in a surgical model of glaucomatous retinopathy. We hypothesized that NAEs that do not bind cannabinoid and vanilloid receptors regulate intracellular calcium homeostasis thereby protecting retinal ganglion cells RGCs , optic nerve head astrocytes ONHAs and the optic nerve from dysfunction and cell death.

    Methods: Intraocular pressure was raised unilaterally in one eye of Brown Norway rats by episcleral vein injection of hypertonic saline. NAEs were delivered intravitreally or systemically intraperitoneally daily and visual acuity and contrast sensitivity were measured with visuo-spatial behavior testing of the optomotor reflex. The extent of optic nerve damage was determined with quantitative histology. Results: NAEs , and , NAEs that do not bind cannabinoid and vanilloid receptors, significantly reduced the loss in synaptic connectivity, loss of axons and cell numbers of RGCs after systemic or intravitreal delivery.

    At the same time, NAE administration also significantly reduced the decline in visual function as measured by visuo-spatial behavior testing. Treated animals had visual performance and histological phenotypes indistinguishable from age-matched and contralateral controls while visual performance, RGC and axon counts continued to decline with age in vehicle treated controls. Conclusions: Treatment with NAEs resulted in a highly significant, dosedependent reduction of glaucomatous retinopathy-induced RGC death and of the loss in visual performance indicating that NAEs that do not bind to cannabinoid and vanilloid receptors and lack endocannabinoid activity have significant neuroprotective potential.

    These results provide a rationale for the use of NAEs as potential therapeutic agents in primary open angle glaucoma and potentially other forms of glaucomatous retinopathy. Purpose: To evaluate the role of Fas apoptosis inhibitory molecule 2 Faim2 , an inhibitor of the Fas signaling pathway, and its regulation by stress kinase signaling during Fas-mediated apoptosis of photoreceptors. Methods: Photoreceptor W cells were treated with a Fas-receptor activating antibody and the levels of Faim2 were examined.

    Activation of the stress-kinase pathways was assayed by immunoblotting of kinases and their downstream targets. The levels of Faim2 were examined in the presence of stress-kinase inhibitors in Fas-treated W cells. The effect of stress-kinase inhibition on Fas-mediated photoreceptor apoptosis was evaluated by caspase 8 activation and measuring cell viability. Faim2 expression was inhibited by siRNA knock-down in W cells and caspase 8 activation was measured after Fas activating antibody treatment.

    Results: Treatment of W cells with a Fas-activating antibody led to increased levels of Faim2. Blocking the ERK pathway using pharmacological inhibitors increased the susceptibility of W cells to Fas-induced caspase activation and apoptosis. When the levels of Faim2 were reduced in W cells by siRNA knock-down, Fas activating antibody treatment resulted in earlier and more robust caspase 8 activation.

    Conclusions: Faim2 acts as a neuroprotectant during Fas-mediated apoptosis of photoreceptors. The expression of Faim2 is regulated, at least in part, by the ERK signaling pathway. Modulation of ERK signaling to increase Faim2 expression may be a potential therapeutic option to prevent photoreceptor death. Commercial Relationships: Cagri G. Calandria, Nicolas G. Purpose: Poly-glutamine-dependent misfolding induces toxicity and consequently contributes to degeneration in the central nervous system.

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    In Spinocerebellar ataxia 1 SCA1 , Ataxin-1 mutation causes the accumulation and partial loss of function of the protein leading to cytotoxicity. Ataxin-1 interacts with transcription factors and spliceosome components. In addition to an involvement of the cerebellum in Copyright by the Association for Research in Vision and Ophthalmology, Inc.

    Psychiatry , , thus providing the basis for applying this model to retinal studies. Akt phosphorylation status on NPD1 treated cells was determined by western blot. Localization and accumulation of Ataxin-1 was determined by immunocytochemistry. This was evidenced by the increase in the percentage of TUNEL and Hoechst positive cells when compared with the controls expressing wild-type protein. Caspase-3 also showed the same pattern of activation.

    The expression of the mutant Ataxin-1 also induced accumulation of the Serine phosphorylated form in the nucleus.

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    The translocation into the nucleus and phosphorylation of Serine of Akt, the main kinase that phosphorylates Ataxin-1, was increased differentially when the 82Q form was expressed even in the presence of NPD1, suggesting that NPD1 does not prevent the phosphorylation of Ataxin-1 and may be acting on its dephosphorylation instead. Conclusions: These results demonstrate that: 1 the RPE cellular model depicts similar features to the changes triggered by the mutation in Purkinje cells, laying the groundwork for unveiling new pathways and therapeutic approaches in other misfolded protein-induced pathologies in the retina and 2 NPD1 rescues the deleterious effects of proteotoxic stress caused by extended polyQ tract Ataxin-1 expression in RPE cells due to the increase in turnover.

    These findings may lead to new therapeutic applications for DHA and NPD1 for preventing and attenuating the initiation of retinal degenerations.

    Commercial Relationships: Jorgelina M. Calandria, None; Nicolas G. Bogea, Beatrice T. Tam, Orson L. Purpose: Type II cell death, also described as cell death with autophagy, has been recently implicated in the regulation of photoreceptor degeneration in several mouse models. In the present study, we report the occurrence of type II cell death in a light induced model of retinal degeneration where Xenopus laevis tadpoles express a bovine form of the rhodopsin mutant P23H bP23H.

    Methods: Transgenic tadpoles expressing bP23H were transferred to a 12L: 12D regimen to induce retinal degeneration. They were euthanized at various time points. Transgenic tadpoles expressing an inducible form of caspase 9 iCasp9 were reared similarly, and retinal degeneration was induced by administration of AP Tadpole eyes were subsequently processed for high resolution electron microscopy according to standard procedures. Results: We observed defects in rod outer and inner segment membranes in tadpoles expressing bP23H, which were aggravated by exposure to cyclic light.

    Rod outer segments were shorter and exhibited vesiculations and altered disk morphology, and were rapidly phagocytosed by the retinal pigment epithelium. Phagocytosis was mostly complete by 36 hours. In rod inner segments, we observed autophagic compartments within the rough endoplasmic reticulum.

    kamishiro-hajime.info/voice/camera-de/mouchard-iphone-5c.php In contrast, these defects were not observed in rod outer and inner segment membranes of tadpoles expressing iCasp9, where the photoreceptors completely degenerated within 24 hours. Conclusions: Our results indicate the presence of ultrastructural defects in both the outer and inner segment membranes of bP23H expressing photoreceptors.

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