Bestrophinopathies: perspectives on clinical disease, Bestrophin-1 function and developing therapies

Bestrophinopathies are a group of clinically distinct inherited retinal dystrophies that typically affect the macular region, an area synonymous with central high acuity vision. This spectrum of disorders is caused by mutations in bestrophin1 (BEST1), a protein thought to act as a Ca2+-activated Cl- channel in the retinal pigment epithelium (RPE) of the eye. Although bestrophinopathies are rare, over 250 individual pathological mutations have been identified in the BEST1 gene, with many reported to have various clinical expressivity and incomplete penetrance. With no current clinical treatments available for patients with bestrophinopathies, understanding the role of BEST1 in cells and the pathological pathways underlying disease has become a priority. Induced pluripotent stem cell (iPSC) technology is helping to uncover disease mechanisms and develop treatments for RPE diseases, like bestrophinopathies. Here, we provide a comprehensive review of the pathophysiology of bestrophinopathies and highlight how patient-derived iPSC-RPE are being used to test new genomic therapies in vitro

Annexin A8 regulates Wnt signaling to maintain the phenotypic plasticity of retinal pigment epithelial cells

Wnt signalling mediates complex cell-cellinteractions during development and proliferation. Annexin A8 (AnxA8), a calcium-dependent phospholipid-binding protein, and canonical Wnt signalling mechanisms have both been implicated in retinal pigment epithelial (RPE) cell differentiation. The aim here was to examine the possibility of cross-talk between AnxA8 and Wnt signalling, as both are down-regulated upon fenretinide (FR)-mediated RPE transdifferentiation. AnxA8 suppression in RPE cells via siRNA or administration of FR induced neuronal-like cell transdifferentiation and reduced expression of Wnt-related genes, as measured by real-time PCR and western blotting. AnxA8 gene expression, on the other hand, remained unaltered upon manipulating Wnt signalling, suggesting Wnt-related genes to be downstream effectors of AnxA8. Co-immunoprecipitation revealed an interaction between AnxA8 and β-catenin, which was reduced in the presence of activated TGF-β1. TGF-β1 signalling also reversed the AnxA8 loss-induced cell morphology changes, and induced β-catenin translocation and GSK-3β phosphorylation in the absence of AnxA8. Ectopic over-expression of AnxA8 led to an increase in active β-catenin and GSK-3β phosphorylation. These data demonstrate an important role for AnxA8 as a regulator of Wnt signalling and a determinant of RPE phenotype, with implications for regenerative medicine approaches that utilise stem cell-derived RPE cells to treat conditions such as age-related macular degeneration

Regulation of retinal pigment epithelial cell phenotype by Annexin A8

The retinoic acid derivative fenretinide (FR) is capable of transdifferentiating cultured retinal pigment epithelial (RPE) cells towards a neuronal-like phenotype, but the underlying mechanisms are not understood. To identify genes involved in this process we performed a microarray analysis of RPE cells pre- and post-FR treatment, and observed a marked down-regulation of AnnexinA8 (AnxA8) in transdifferentiated cells. To determine whether AnxA8 plays a role in maintaining RPE cell phenotype we directly manipulated AnxA8 expression in cultured and primary RPE cells using siRNA-mediated gene suppression, and over-expression of AnxA8-GFP in conjunction with exposure to FR. Treatment of RPE cells with AnxA8 siRNA recapitulated exposure to FR, with cell cycle arrest, neuronal transdifferentiation, and concomitant up-regulation of the neuronal markers calretinin and calbindin, as assessed by real-time PCR and immunofluorescence. In contrast, AnxA8 transient over-expression in ARPE-19 cells prevented FR-induced differentiation. Ectopic expression of AnxA8 in AnxA8-depleted cells led to decreased neuronal marker staining, and normal cell growth as judged by phosphohistone H3 staining, cell counting and cleaved caspase-3 levels. These data show that down-regulation of AnxA8 is both necessary and sufficient for neuronal transdifferentiation of RPE cells and reveal an essential role for AnxA8 as a key regulator of RPE phenotype

Diffuse small bowel thickening in aids patient - a case report

<p>Abstract</p> <p>Background</p> <p>Diarrhea is common in HIV/AIDS patients, caused by both classic enteric pathogens and different opportunistic agents. <it>Infection with these different pathogens may lead to similar radiological findings, thus causing diagnostic confusion</it>.</p> <p>Case presentation</p> <p>A 30-yr-old female with AIDS presented with chronic diarrhea of 4 months duration. She had diffuse small bowel thickening present on CT scan of her abdomen, with stool examination showing no parasites. She was erroneously diagnosed as abdominal tuberculosis and given antituberculosis drugs with which she showed no improvement. Repeat stool examination later at a specialized laboratory revealed <it>Cryptosporidium parvum </it>infection.</p> <p>The patient was given an extended course of nitazoxanide treatment, as her stool examination was positive for <it>Cryptosporidium parvum </it>even after 2 weeks of drug consumption. Parasite clearance was documented after 10 weeks of treatment. Interestingly, the bowel thickening reversed with parasitological clearance.</p> <p>Conclusions</p> <p><it>Cryptosporidium parvum </it>may lead to small bowel thickening in AIDS patients. This small bowel thickening may reverse following parasitological clearance.</p

Protective effects of curcumin against gamma radiation-induced ileal mucosal damage

The major objective of this study was to test curcumin as a potential radioprotectant for the ileum goblet cells of the rat. Wistar albino rats were used in the study. Group A was the control group and group B was the single dose radiation group. Group C was the two dose radiation group (4 days interval). The rats in groups D and E were given a daily dose of 100 mg/kg of curcumin for 14 and 18 days, respectively. During the curcumin administration period, the rats in group D were exposed to abdominal area gamma (γ)-ray dose of 5 Gy on the 10th day and group E was exposed to same dose radiation on the 10th and 14th day. Irradiation and treatment groups were decapitated on the 4th day after exposure to single or two-dose irradiation and ileum tissues were removed for light and electron microscopic investigation. Single or two dose 5 Gy γ-irradiation caused a marked intestinal mucosal injury in rats on the 4th day. Radiation produced increases in the number of goblet cells. Curcumin appears to have protective effects against radiation-induced damage, suggesting that clinical transfer is feasible

The illusion of competency versus the desirability of expertise: Seeking a common standard for support professions in sport

In this paper we examine and challenge the competency-based models which currently dominate accreditation and development systems in sport support disciplines, largely the sciences and coaching. Through consideration of exemplar shortcomings, the limitations of competency-based systems are presented as failing to cater for the complexity of decision making and the need for proactive experimentation essential to effective practice. To provide a better fit with the challenges of the various disciplines in their work with performers, an alternative approach is presented which focuses on the promotion, evaluation and elaboration of expertise. Such an approach resonates with important characteristics of professions, whilst also providing for the essential ‘shades of grey’ inherent in work with human participants. Key differences between the approaches are considered through exemplars of evaluation processes. The expertise-focused method, although inherently more complex, is seen as offering a less ambiguous and more positive route, both through more accurate representation of essential professional competence and through facilitation of future growth in proficiency and evolution of expertise in practice. Examples from the literature are also presented, offering further support for the practicalities of this approach

Identification and Correction of Mechanisms Underlying Inherited Blindness in Human iPSC-Derived Optic Cups

Leber congenital amaurosis (LCA) is an inherited retinal dystrophy that causes childhood blindness. Photoreceptors are especially sensitive to an intronic mutation in the cilia-related gene CEP290, which causes missplicing and premature termination, but the basis of this sensitivity is unclear. Here, we generated differentiated photoreceptors in three-dimensional optic cups and retinal pigment epithelium (RPE) from iPSCs with this common CEP290 mutation to investigate disease mechanisms and evaluate candidate therapies. iPSCs differentiated normally into RPE and optic cups, despite abnormal CEP290 splicing and cilia defects. The highest levels of aberrant splicing and cilia defects were observed in optic cups, explaining the retinal-specific manifestation of this CEP290 mutation. Treating optic cups with an antisense morpholino effectively blocked aberrant splicing and restored expression of full-length CEP290, restoring normal cilia-based protein trafficking. These results provide a mechanistic understanding of the retina-specific phenotypes in CEP290 LCA patients and potential strategies for therapeutic intervention