Retinal Degeneration Progression Changes Lentiviral Vector Cell Targeting in the Retina

In normal mice, the lentiviral vector (LV) is very efficient to target the RPE cells, but transduces retinal neurons well only during development. In the present study, the tropism of LV has been investigated in the degenerating retina of mice, knowing that the retina structure changes during degeneration. We postulated that the viral transduction would be increased by the alteration of the outer limiting membrane (OLM). Two different LV pseudotypes were tested using the VSVG and the Mokola envelopes, as well as two animal models of retinal degeneration: light-damaged Balb-C and Rhodopsin knockout (Rho-/-) mice. After light damage, the OLM is altered and no significant increase of the number of transduced photoreceptors can be obtained with a LV-VSVG-Rhop-GFP vector. In the Rho-/- mice, an alteration of the OLM was also observed, but the possibility of transducing photoreceptors was decreased, probably by ongoing gliosis. The use of a ubiquitous promoter allows better photoreceptor transduction, suggesting that photoreceptor-specific promoter activity changes during late stages of photoreceptor degeneration. However, the number of targeted photoreceptors remains low. In contrast, LV pseudotyped with the Mokola envelope allows a wide dispersion of the vector into the retina (corresponding to the injection bleb) with preferential targeting of Müller cells, a situation which does not occur in the wild-type retina. Mokola-pseudotyped lentiviral vectors may serve to engineer these glial cells to deliver secreted therapeutic factors to a diseased area of the retina

Corneal Transduction by Intra-Stromal Injection of AAV Vectors In Vivo in the Mouse and Ex Vivo in Human Explants

The cornea is a transparent, avascular tissue that acts as the major refractive surface of the eye. Corneal transparency, assured by the inner stroma, is vital for this role. Disruption in stromal transparency can occur in some inherited or acquired diseases. As a consequence, light entering the eye is blocked or distorted, leading to decreased visual acuity. Possible treatment for restoring transparency could be via viral-based gene therapy. The stroma is particularly amenable to this strategy due to its immunoprivileged nature and low turnover rate. We assayed the potential of AAV vectors to transduce keratocytes following intra-stromal injection in vivo in the mouse cornea and ex vivo in human explants. In murine and human corneas, we transduced the entire stroma using a single injection, preferentially targeted keratocytes and achieved long-term gene transfer (up to 17 months in vivo in mice). Of the serotypes tested, AAV2/8 was the most promising for gene transfer in both mouse and man. Furthermore, transgene expression could be transiently increased following aggression to the cornea

Adenosine A2A receptors modulate BDNF both in normal conditions and in experimental models of Huntington’s disease

Brain-derived neurotrophic factor (BDNF), a member of the neurotrophin family, enhances synaptic transmission and regulates neuronal proliferation and survival. Functional interactions between adenosine A2A receptors (A2ARs) and BDNF have been recently reported. In this article, we report some recent findings from our group showing that A2ARs regulate both BDNF functions and levels in the brain. Whereas BDNF (10 ng/ml) increased the slope of excitatory postsynaptic field potentials (fEPSPs) in hippocampal slices from wild-type (WT) mice, it was completely ineffective in slices taken from A2AR knock-out (KO) mice. Furthermore, enzyme immunoassay studies showed a significant reduction in hippocampal BDNF levels in A2AR KO vs. WT mice. Having found an even marked reduction in the striatum of A2AR KO mice, and as both BDNF and A2ARs have been implicated in the pathogenesis of Huntington’s disease (HD), an inherited striatal neurodegenerative disease, we then evaluated whether the pharmacological blockade of A2ARs could influence striatal levels of BDNF in an experimental model of HD-like striatal degeneration (quinolinic acid-lesioned rats) and in a transgenic mice model of HD (R6/2 mice). In both QA-lesioned rats and early symptomatic R6/2 mice (8 weeks), the systemic administration of the A2AR antagonist SCH58261 significantly reduced striatal BDNF levels. These results indicate that the presence and the tonic activation of A2ARs are necessary to allow BDNF-induced potentiation of synaptic transmission and to sustain a normal BDNF tone. The possible functional consequences of reducing striatal BDNF levels in HD models need further investigation

Repurposing NGO data for better research outcomes: A scoping review of the use and secondary analysis of NGO data in health policy and systems research

Background Non-government organisations (NGOs) collect and generate vast amounts of potentially rich data, most of which are not used for research purposes. Secondary analysis of NGO data (their use and analysis in a study for which they were not originally collected) presents an important but largely unrealised opportunity to provide new research insights in critical areas including the evaluation of health policy and programmes. Methods A scoping review of the published literature was performed to identify the extent to which secondary analysis of NGO data has been used in health policy and systems research (HPSR). A tiered analytic approach provided a comprehensive overview and descriptive analyses of the studies which: 1) used data produced or collected by or about NGOs; 2) performed secondary analysis of the NGO data (beyond use of an NGO report as a supporting reference); 3) used NGO-collected clinical data. Results Of the 156 studies which performed secondary analysis of NGO-produced or collected data, 64% (n=100) used NGO-produced reports (e.g. to critique NGO activities and as a contextual reference) and 8% (n=13) analysed NGO-collected clinical data.. Of the studies, 55% investigated service delivery research topics, with 48% undertaken in developing countries and 17% in both developing and developed. NGO-collected clinical data enabled HPSR within marginalised groups (e.g. migrants, people in conflict-affected areas), with some limitations such as inconsistencies and missing data. Conclusion We found evidence that NGO-collected and produced data are most commonly perceived as a source of supporting evidence for HPSR and not as primary source data. However, these data can facilitate research in under-researched marginalised groups and in contexts that are hard to reach by academics, such as conflict-affected areas. NGO–academic collaboration could help address issues of NGO data quality to facilitate their more widespread use in research. Their use could enable relevant and timely research in the areas of health policy, programme evaluation and advocacy to improve health and reduce health inequalities, especially in marginalised groups and developing countries

Hyperactivation of retina by light in mice leads to photoreceptor cell death mediated by VEGF and retinal pigment epithelium permeability

International audienceLight toxicity is suspected to enhance certain retinal degenerative processes such as age-related macular degeneration. Death of photoreceptors can be induced by their exposure to the visible light, and although cellular processes within photoreceptors have been characterized extensively, the role of the retinal pigment epithelium (RPE) in this model is less well understood. We demonstrate that exposition to intense light causes the immediate breakdown of the outer blood–retinal barrier (BRB). In a molecular level, we observed the slackening of adherens junctions tying up the RPE and massive leakage of albumin into the neural retina. Retinal pigment epithelial cells normally secrete vascular endothelial growth factor (VEGF) at their basolateral side; light damage in contrast leads to VEGF increase on the apical side – that is, in the neuroretina. Blocking VEGF, by means of lentiviral gene transfer to express an anti-VEGF antibody in RPE cells, inhibits outer BRB breakdown and retinal degeneration, as illustrated by functional, behavioral and morphometric analysis. Our data show that exposure to high levels of visible light induces hyperpermeability of the RPE, likely involving VEGF signaling. The resulting retinal edema contributes to irreversible damage to photoreceptors. These data suggest that anti-VEGF compounds are of therapeutic interest when the outer BRB is altered by retinal stresses

CRISPR-LbCpf1 prevents choroidal neovascularization in a mouse model of age-related macular degeneration

The CRISPR endonuclease LbCpf1 is reported to have greater efficiency and specificity than Cas9. Here, the authors use LbCpf1 to target the angiogenesis-related genes VEGF and HIF1a, and show that delivery of the nuclease using AAV9 is effective in mouse models of macular degeneration