Müller glia activation in response to inherited retinal degeneration is highly varied and disease-specific

Despite different aetiologies, most inherited retinal disorders culminate in photoreceptor loss, which induces concomitant changes in the neural retina, one of the most striking being reactive gliosis by Müller cells. It is typically assumed that photoreceptor loss leads to an upregulation of glial fibrilliary acidic protein (Gfap) and other intermediate filament proteins, together with other gliosis-related changes, including loss of integrity of the outer limiting membrane (OLM) and deposition of proteoglycans. However, this is based on a mix of both injury-induced and genetic causes of photoreceptor loss. There are very few longitudinal studies of gliosis in the retina and none comparing these changes across models over time. Here, we present a comprehensive spatiotemporal assessment of features of gliosis in the degenerating murine retina that involves Müller glia. Specifically, we assessed Gfap, vimentin and chondroitin sulphate proteoglycan (CSPG) levels and outer limiting membrane (OLM) integrity over time in four murine models of inherited photoreceptor degeneration that encompass a range of disease severities (Crb1rd8/rd8, Prph2+/Δ307, Rho-/-, Pde6brd1/rd1). These features underwent very different changes, depending upon the disease-causing mutation, and that these changes are not correlated with disease severity. Intermediate filament expression did indeed increase with disease progression in Crb1rd8/rd8 and Prph2+/Δ307, but decreased in the Prph2+/Δ307 and Pde6brd1/rd1 models. CSPG deposition usually, but not always, followed the trends in intermediate filament expression. The OLM adherens junctions underwent significant remodelling in all models, but with differences in the composition of the resulting junctions; in Rho-/- mice, the adherens junctions maintained the typical rod-Müller glia interactions, while in the Pde6brd1/rd1 model they formed predominantly between Müller cells in late stage of degeneration. Together, these results show that gliosis and its associated processes are variable and disease-dependent

Care provision during termination of pregnancy following diagnosis of a severe congenital anomaly – a qualitative study of what is important to parents.

Objective: To understand the experiences of women and their partners following the decision to terminate a pregnancy affected by a severe congenital anomaly Design: Qualitative semi-structured interviews with a purposive sample of women and their partners who underwent a termination of pregnacy following diagnosis of a severe congenital anomaly. Setting: Women referred to four fetal medicine centres across two hospital trusts. Analysis: Data analysis was based on the constant comparative approach. Findings: The over-arching theme emerging from the data was that of ‘falling through the gap’, where the care received did not adequately meet the needs of women and their partners. This was particularly salient at three specific points in the care pathway: enacting the decision to have a termination of pregnancy and subsequent initiation of the process; care during labour and birth, where parents describe being caught in ‘no-man’s land’ between the antenatal and postnatal settings; and post-birth, where parents made sense of and came to terms with their decision. Conclusions: The diagnosis of a severe congenital anomaly and the subsequent pathway that parents face is a traumatic event. Responsibility for the decision to terminate the pregnancy intensifies emotions and adds to the complexity of caring for this group. These findings point to the need for a specific care pathway for parents undergoing this difficult experience. Recommendations include the need for a greater understanding of the views of midwives caring for these parents, review of specific training needs, and examination of processes to better support both parents

Rapid growth of an intact human liver transplanted into a recipient larger than the donor

Two individuals undergoing orthotopic hepatic transplantation received livers from donors who were on average 10 kg smaller than themselves based on recipient ideal body weight. As a result, the donor livers in these 2 cases were 29%-59% smaller than would be expected had the donor liver and recipient been matched ideally. The liver grafts in the recipients steadily increased in size, as determined by serial computed tomography scanning, to achieve new volumes consistent with those that would have been expected in a normal individual of the recipient's size, sex, and age. Fasting plasma levels of amino acids, glucagon, insulin, and standard liver injury tests were monitored to determine which measure best reflected the changes observed in the size of the grafts over time. No relationship between the changes observed in any of these parameters and hepatic growth was apparent. In both cases, the liver increased in volume at a rate of ~70 ml/day. These data demonstrate that a small-for-size liver transplanted into a larger recipient increases in size at a rate of ~70 ml/day until it achieves a liver volume consistent with that expected given the recipient's size, age, and sex. © 1987

Gene therapy restores vision in rd1 mice after removal of a confounding mutation in Gpr179

The rd1 mouse with a mutation in the Pde6b gene was the first strain of mice identified with a retinal degeneration. However, AAV-mediated gene supplementation of rd1 mice only results in structural preservation of photoreceptors, and restoration of the photoreceptor-mediated a-wave, but not in restoration of the bipolar cell-mediated b-wave. Here we show that a mutation in Gpr179 prevents the full restoration of vision in rd1 mice. Backcrossing rd1 with C57BL6 mice reveals the complete lack of b-wave in a subset of mice, consistent with an autosomal recessive Mendelian inheritance pattern. We identify a mutation in the Gpr179 gene, which encodes for a G-protein coupled receptor localized to the dendrites of ON-bipolar cells. Gene replacement in rd1 mice that are devoid of the mutation in Gpr179 successfully restores the function of both photoreceptors and bipolar cells, which is maintained for up to 13 months. Our discovery may explain the failure of previous gene therapy attempts in rd1 mice, and we propose that Grp179 mutation status should be taken into account in future studies involving rd1 mice

A comprehensive atlas of Aggrecan, Versican, Neurocan and Phosphacan expression across time in wildtype retina and in retinal degeneration

As photoreceptor cells die during retinal degeneration, the surrounding microenvironment undergoes significant changes that are increasingly recognized to play a prominent role in determining the efficacy of therapeutic interventions. Chondroitin Sulphate Proteoglycans (CSPGs) are a major component of the extracellular matrix that have been shown to inhibit neuronal regrowth and regeneration in the brain and spinal cord, but comparatively little is known about their expression in retinal degeneration. Here we provide a comprehensive atlas of the expression patterns of four individual CSPGs in three models of inherited retinal degeneration and wildtype mice. In wildtype mice, Aggrecan presented a biphasic expression, while Neurocan and Phosphacan expression declined dramatically with time and Versican expression remained broadly constant. In degeneration, Aggrecan expression increased markedly in Aipl1-/- and Pde6brd1/rd1, while Versican showed regional increases in the periphery of Rho-/- mice. Conversely, Neurocan and Phosphacan broadly decrease with time in all models. Our data reveal significant heterogeneity in the expression of individual CSPGs. Moreover, there are striking differences in the expression patterns of specific CSPGs in the diseased retina, compared with those reported following injury elsewhere in the CNS. Better understanding of the distinct distributions of individual CSPGs will contribute to creating more permissive microenvironments for neuro-regeneration and repair

Role of HIF1α and HIF2α in Cre Recombinase–Induced Retinal Pigment Epithelium Pathology and Its Secondary Effect on Choroidal Neovascularization

CreTrp1 mice are widely used for conditional retinal pigment epithelium (RPE) gene function studies. Like other Cre/LoxP models, phenotypes in CreTrp1 mice can be affected by Cre-mediated cellular toxicity, leading to RPE dysfunction, altered morphology and atrophy, activation of innate immunity, and consequent impairment of photoreceptor function. These effects are common among the age-related alterations of RPE that feature in early/intermediate forms of age-related macular degeneration. This article characterizes Cre-mediated pathology in the CreTrp1 line to elucidate the impact of RPE degeneration on both developmental and pathologic choroidal neovascularization. Nonredundant roles of the two major components of the hypoxia-inducible factor (HIF) family of transcription regulators, HIF1α and HIF2α, were identified. Genetic ablation of Hif1a protected against Cre-induced degeneration of RPE and choroid, whereas ablation of Hif2a exacerbated this degeneration. Furthermore, HIF1α deficiency protected CreTrp1 mice against laser-induced choroidal neovascularization, whereas HIF2α deficiency exacerbated the phenotype. Cre-mediated degeneration of the RPE in CreTrp1 mice offers an opportunity to investigate the impact of hypoxia signaling in the context of RPE degeneration. These findings indicate that HIF1α promotes Cre recombinase–mediated RPE degeneration and laser-induced choroidal neovascularization, whereas HIF2α is protective

Harnessing the Potential of Human Pluripotent Stem Cells and Gene Editing for the Treatment of Retinal Degeneration

PURPOSE OF REVIEW: A major cause of visual disorders is dysfunction and/or loss of the light-sensitive cells of the retina, the photoreceptors. To develop better treatments for patients, we need to understand how inherited retinal disease mutations result in the dysfunction of photoreceptors. New advances in the field of stem cell and gene editing research offer novel ways to model retinal dystrophies in vitro and present opportunities to translate basic biological insights into therapies. This brief review will discuss some of the issues that should be taken into account when carrying out disease modelling and gene editing of retinal cells. We will discuss (i) the use of human induced pluripotent stem cells (iPSCs) for disease modelling and cell therapy; (ii) the importance of using isogenic iPSC lines as controls; (iii) CRISPR/Cas9 gene editing of iPSCs; and (iv) in vivo gene editing using AAV vectors. RECENT FINDINGS: Ground-breaking advances in differentiation of iPSCs into retinal organoids and methods to derive mature light sensitive photoreceptors from iPSCs. Furthermore, single AAV systems for in vivo gene editing have been developed which makes retinal in vivo gene editing therapy a real prospect. SUMMARY: Genome editing is becoming a valuable tool for disease modelling and in vivo gene editing in the retina

Apparent mass of small children: Experimental measurements

A test facility and protocol were developed for measuring the seated, vertical, whole-body vibration response of small children of less than 18 kg in mass over the frequency range from 1 to 45 Hz. The facility and protocol adhered to the human vibration testing guidelines of BS7085 and to current codes of ethics for research involving children. Additional procedures were also developed which are not currently defined in the guidelines, including the integral involvement of the parents and steps taken to maximize child happiness. Eight children were tested at amplitudes of 0.8 and 1.2 m/s2 using band-limited, Gaussian, white noise acceleration signals defined over the frequency interval from 1 to 50 Hz. Driving point apparent mass modulus and phase curves were determined for all eight children at both test amplitudes. All results presented a single, principal, anti-resonance, and were similar to data reported for primates and for adult humans seated in an automotive posture which provided backrest support. The mean frequency of the apparent mass peak was 6.25 Hz for the small children, as compared to values between 6.5 - 8.5 Hz for small primates and values between 6.5 - 8.6 Hz for adults seated with backrest support. The peak value of the mean, normalized, apparent mass was 1.54 for the children, which compares to values from 1.19 to 1.45 reported in the literature for small primates and 1.28 for adults seated with backrest support. ISO standard 5982, which specifies a mean, normalized, apparent mass modulus peak of 1.50 at a frequency of 4.0 Hz for adults seated without backrest support, provides significant differences

Bioinformatics advances in saliva diagnostics

There is a need recognized by the National Institute of Dental & Craniofacial Research and the National Cancer Institute to advance basic, translational and clinical saliva research. The goal of the Salivaomics Knowledge Base (SKB) is to create a data management system and web resource constructed to support human salivaomics research. To maximize the utility of the SKB for retrieval, integration and analysis of data, we have developed the Saliva Ontology and SDxMart. This article reviews the informatics advances in saliva diagnostics made possible by the Saliva Ontology and SDxMart