Retinal gene therapy with a large MYO7A cDNA using adeno-associated virus.

Usher 1 patients are born profoundly deaf and then develop retinal degeneration. Thus they are readily identified before the onset of retinal degeneration, making gene therapy a viable strategy to prevent their blindness. Here, we have investigated the use of adeno-associated viruses (AAVs) for the delivery of the Usher 1B gene, MYO7A, to retinal cells in cell culture and in Myo7a-null mice. MYO7A cDNA, under control of a smCBA promoter, was packaged in single AAV2 and AAV5 vectors and as two overlapping halves in dual AAV2 vectors. The 7.9-kb smCBA-MYO7A exceeds the capacity of an AAV vector; packaging of such oversized constructs into single AAV vectors may involve fragmentation of the gene. Nevertheless, the AAV2 and AAV5 single vector preparations successfully transduced photoreceptor and retinal pigment epithelium cells, resulting in functional, full-length MYO7A protein and correction of mutant phenotypes, suggesting successful homologous recombination of gene fragments. With discrete, conventional-sized dual AAV2 vectors, full-length MYO7A was detected, but the level of protein expression was variable, and only a minority of cells showed phenotype correction. Our results show that MYO7A therapy with AAV2 or AAV5 single vectors is efficacious; however, the dual AAV2 approach proved to be less effective

DWSB in heterotic flux compactifications

We address the construction of non-supersymmetric vacua in heterotic compactifications with intrinsic torsion and background fluxes. In particular, we implement the approach of domain-wall supersymmetry breaking (DWSB) previously developed in the context of type II flux compactifications. This approach is based on considering backgrounds where probe NS5-branes wrapping internal three-cycles and showing up as four-dimensional domain-walls do not develop a BPS bound, while all the other BPS bounds characterizing the N=1 supersymmetric compactifications are preserved at tree-level. Via a scalar potential analysis we provide the conditions for these backgrounds to solve the ten-dimensional equations of motion including order \alpha' corrections. We also consider backgrounds where some of the NS5-domain-walls develop a BPS bound, show their relation to no-scale SUSY-breaking vacua and construct explicit examples via elliptic fibrations. Finally, we consider backgrounds with a non-trivial gaugino condensate and discuss their relation to supersymmetric and non-supersymmetric vacua in the present context.Comment: 56 pages, 1 figur

Rpgrip1 is required for rod outer segment development and ciliary protein trafficking in zebrafish

The authors would like to thank the Royal Society of London, the National Eye Research Centre, the Visual Research Trust, Fight for Sight, the W.H. Ross Foundation, the Rosetrees Trust, and the Glasgow Children’s Hospital Charity for supporting this work. This work was also supported by the Deanship of Scientific Research at King Saud University for funding this research (Research Project) grant number ‘RGP – VPP – 219’.Mutations in the RPGR-interacting protein 1 (RPGRIP1) gene cause recessive Leber congenital amaurosis (LCA), juvenile retinitis pigmentosa (RP) and cone-rod dystrophy. RPGRIP1 interacts with other retinal disease-causing proteins and has been proposed to have a role in ciliary protein transport; however, its function remains elusive. Here, we describe a new zebrafish model carrying a nonsense mutation in the rpgrip1 gene. Rpgrip1homozygous mutants do not form rod outer segments and display mislocalization of rhodopsin, suggesting a role for RPGRIP1 in rhodopsin-bearing vesicle trafficking. Furthermore, Rab8, the key regulator of rhodopsin ciliary trafficking, was mislocalized in photoreceptor cells of rpgrip1 mutants. The degeneration of rod cells is early onset, followed by the death of cone cells. These phenotypes are similar to that observed in LCA and juvenile RP patients. Our data indicate RPGRIP1 is necessary for rod outer segment development through regulating ciliary protein trafficking. The rpgrip1 mutant zebrafish may provide a platform for developing therapeutic treatments for RP patients.Publisher PDFPeer reviewe

Negative discriminant states in N=4 supersymmetric string theories

Single centered BPS black hole solutions exist only when the charge carried by the black hole has positive discriminant. On the other hand the exact dyon spectrum in heterotic string theory compactified on T^6 is known to contain states with negative discriminant. We show that all of these negative discriminant states can be accounted for as two centered black holes. Thus after the contribution to the index from the two centered black holes is subtracted from the total microscopic index, the index for states with negative discriminant vanishes even for finite values of charges, in agreement with the results from the black hole side. Bound state metamorphosis -- which requires us to identify certain apparently different two centered configurations according to a specific set of rules -- plays a crucial role in this analysis. We also generalize these results to a class of CHL string theories.Comment: LaTeX file, 32 pages; v2: reference added; v3: added new section 3.

In Vitro Antiophidian Properties of Dipteryx alata Vogel Bark Extracts

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Extracts from Dipteryx alata bark obtained with different solvents (hexane, dichloromethane, ethyl acetate and methanol) were mixed in vitro with Bothrops jararacussu (Bjssu, 40 mu g/mL) and Crotalus durissus terrificus (Cdt, 15 mu g/mL) snake venoms, and applied to a mouse phrenic nerve-diaphragm preparation to evaluate the possible neutralization of venom effects. Cdt venom neurotoxic effect was not inhibited by any of the extracts, while the neurotoxic and myotoxic actions of Bjssu venom were decreased by the methanolic extract. This inhibition appears to be augmented by tannins. Dichloromethane bark extract inhibited similar to 40% of Bjssu venom effects and delayed blockade induced by Cdt. The methodology used to determine which extract was active allows inferring that: (i) phenolic acids and flavonoids contained in the methanolic extract plus tannins were responsible mostly for neutralization of Bjssu effects; (ii) terpenoids from the dichloromethane extract may participate in the anti-Cdt and anti-Bjssu venom effects; (iii) a given extract could not inhibit venoms from different species even if those belong to the same family, so it is improper to generalize a certain plant as antiophidian; (iv) different polarity extracts do not present the same inhibitory capability, thus demonstrating the need for characterizing both venom pharmacology and the phytochemistry of medicinal plant compounds.15959565970Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)PROBIC/UNISOConselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)FAPESP [Proc. FAPESP 04/09705-8, 07/53883-6]FAPESP [07/51414-9, 08/05459-3]CNPq [Proc. 302206/2008-6

The Correlation between Mixing Length and Metallicity on the Giant Branch: Implications for Ages in the Gaia Era

In the updated APOGEE-Kepler catalog, we have asteroseismic and spectroscopic data for over 3000 first ascent red giants. Given the size and accuracy of this sample, these data offer an unprecedented test of the accuracy of stellar models on the post-main-sequence. When we compare these data to theoretical predictions, we find a metallicity dependent temperature offset with a slope of around 100 K per dex in metallicity. We find that this effect is present in all model grids tested, and that theoretical uncertainties in the models, correlated spectroscopic errors, and shifts in the asteroseismic mass scale are insufficient to explain this effect. Stellar models can be brought into agreement with the data if a metallicity-dependent convective mixing length is used, with Delta alpha(ML), YREC similar to 0.2 per dex in metallicity, a trend inconsistent with the predictions of three-dimensional stellar convection simulations. If this effect is not taken into account, isochrone ages for red giants from the Gaia data will be off by as much as a factor of two even at modest deviations from solar metallicity ([Fe/H]- -0.5)

Neuronal circuitry for pain processing in the dorsal horn

Neurons in the spinal dorsal horn process sensory information, which is then transmitted to several brain regions, including those responsible for pain perception. The dorsal horn provides numerous potential targets for the development of novel analgesics and is thought to undergo changes that contribute to the exaggerated pain felt after nerve injury and inflammation. Despite its obvious importance, we still know little about the neuronal circuits that process sensory information, mainly because of the heterogeneity of the various neuronal components that make up these circuits. Recent studies have begun to shed light on the neuronal organization and circuitry of this complex region

Toward a better definition of focal cortical dysplasia: An iterative histopathological and genetic agreement trial.

OBJECTIVE: Focal cortical dysplasia (FCD) is a major cause of difficult-to-treat epilepsy in children and young adults, and the diagnosis is currently based on microscopic review of surgical brain tissue using the International League Against Epilepsy classification scheme of 2011. We developed an iterative histopathological agreement trial with genetic testing to identify areas of diagnostic challenges in this widely used classification scheme. METHODS: Four web-based digital pathology trials were completed by 20 neuropathologists from 15 countries using a consecutive series of 196 surgical tissue blocks obtained from 22 epilepsy patients at a single center. Five independent genetic laboratories performed screening or validation sequencing of FCD-relevant genes in paired brain and blood samples from the same 22 epilepsy patients. RESULTS: Histopathology agreement based solely on hematoxylin and eosin stainings was low in Round 1, and gradually increased by adding a panel of immunostainings in Round 2 and the Delphi consensus method in Round 3. Interobserver agreement was good in Round 4 (kappa = .65), when the results of genetic tests were disclosed, namely, MTOR, AKT3, and SLC35A2 brain somatic mutations in five cases and germline mutations in DEPDC5 and NPRL3 in two cases. SIGNIFICANCE: The diagnoses of FCD 1 and 3 subtypes remained most challenging and were often difficult to differentiate from a normal homotypic or heterotypic cortical architecture. Immunohistochemistry was helpful, however, to confirm the diagnosis of FCD or no lesion. We observed a genotype-phenotype association for brain somatic mutations in SLC35A2 in two cases with mild malformation of cortical development with oligodendroglial hyperplasia in epilepsy. Our results suggest that the current FCD classification should recognize a panel of immunohistochemical stainings for a better histopathological workup and definition of FCD subtypes. We also propose adding the level of genetic findings to obtain a comprehensive, reliable, and integrative genotype-phenotype diagnosis in the near future