Glucose protects cultured retinal cells from oxidative injury via pentose phosphate pathway activation and maintenance of reduced glutathione

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.Purpose : Oxidative injury has been implicated to play a role in a range of retinal neurodegenerative conditions. Thus, protecting retinal cells in vivo from such an insult is extremely beneficial. We therefore sought to investigate whether glucose, acting via the pentose phosphate pathway (PPP) was able to counteract oxidative stress to retinal cells in culture. Methods : Mixed retinal neuron-glial cultures were prepared from 2 day old rat pups and used at 7 days in vitro. Neuron-only and primary Muller cell cultures were prepared from mixed cultures, by treatment with cytosine arabinoside to kill dividing cells, or, by regular medium changes for 30 days, respectively. At appropriate stages, cultures were treated with t-butyl hydroperoxide (tbH; 100nM-10mM) in energy substrate-free DMEM to induce oxidative stress. Some cultures were co-treated with glucose (100µM-25mM) or other metabolic substrates (5mM; pyruvate, lactate, glutamine, fructose-1,6-bisphosphate). Glycolysis was inhibited with iodoacetate (IOA; 10µM) or 2-deoxy-D-glucose (2-DG; 1mM). PPP was inhibited with 6-aminonicotinamide (6NA; 500µM) and glutathione biosynthesis with buthionine sulphoxamine (BSO; 100µM). Cell viability, immunocytochemistry and Western blot were employed to assess cellular outcomes after treatments. An antioxidant assay identified which, if any, of the metabolic substrates tested had intrinsic antioxidant properties. Results : Oxidative stress resulted in loss of viability to mixed retinal cells and primary Muller cells: the EC50 for tbH was approximately 35µM in each case. Glucose dose-dependently reduced the toxicity of tbH with a maximal effect at 5mM (EC50 of tbH elevated to approximately 250µM). Pyruvate was also partially protective, but had intrinsic antioxidant properties. Glycolytic blockade had no effect on the protective effect of glucose but both 6NA and BSO inhibited the protective response. When cultured alone, neurons were equally susceptible to tbH-toxicity but could not be protected by glucose. Conclusions : Glucose prevented oxidative stress to retinal cells via the PPP and the consequent generation of reduced glutathione. Neurons were not subjected to glucose-induced protection except when glial cells were present, implying the passage of a transmissible protective factor between the two cell types.John P M Wood, Glyn Chidlow, Teresa Mammone, Robert James Casso

Expression and activation of SAPK/JNK in the ONH in a rat model of ocular hypertension

Teresa Mammone, Glyn Chidlow, Robert J Casson, John PM Woo

Characterization of the novel spontaneously immortalized rat Müller cell line SIRMu-1

Müller cells (MCs) play a crucial role in the retina, and cultured MC lines are an important tool with which to study MC function. Transformed MC lines have been widely used; however, the transformation process can also lead to unwanted changes compared to the primary cells from which they were derived. To provide an alternative experimental tool, a monoclonal novel spontaneously immortalized rat Müller cell line, SIRMu-1, was derived from primary rat MCs and characterized. Immunofluorescence, western blotting and RNA sequencing demonstrate that the SIRMu-1 cell line retains similar characteristics to cultured primary MCs in terms of expression of the MC markers cellular retinaldehyde-binding protein, glutamine synthetase, S100, vimentin and glial fibrillary acidic protein at both the mRNA and protein levels. Both the cellular morphology and overall transcriptome of the SIRMu-1 cells are more similar to primary rat MCs than the commonly used rMC-1 cells, a well-described, transformed rat MC line. Furthermore, SIRMu-1 cells proliferate rapidly, have an effectively indefinite life span and a high transfection efficiency. The expression of Y chromosome specific genes confirmed that the SIRMu-1 cells are derived from male MCs. Thus, the SIRMu-1 cell line represents a valuable experimental tool to study roles of MCs in both physiological and pathological states.Thaksaon Kittipassorn, Cameron D. Haydinger, John P.M. Wood, Teresa Mammone, Robert J. Casson, Daniel J. Pee

RNA sequencing data of cultured primary rat Muller cells, the spontaneously immortalized rat Muller cell line, SIRMu-1, and the SV40-transformed rat Muller cell line, rMC-1

Available online 7 March 2019Müller cells (MCs), the major type of glial cell of the vertebrate retina, have a vital role in retinal physiology and pathology. They provide structural and functional support for retinal neurons, including photoreceptors, and are implicated in various retinal diseases. Primary and immortalized MCs are important experimental tools for MC research. Here we present high throughput RNA sequencing data of 3 populations of cultured rat MCs: primary cells, the spontaneously immortalized rat MC line, SIRMu-1, and the SV40-transformed rat MC line, rMC-1. These data were deposited in NCBI Gene Expression Omnibus (GEO ID: GSE123161). For data analysis, interpretation and discussion, please refer to the research article, “Characterization of the novel spontaneously immortalized rat Müller cell line SIRMu-1” (Kittipassorn et al., 2019). This dataset is valuable for gaining insight into gene expression profiles of different types of cultured MCs and the roles of MCs in health and disease.Thaksaon Kittipassorn, Cameron D. Haydinger, John P.M. Wood, Teresa Mammone, Robert J. Casson, Daniel J. Pee

Reproductive cycle and gonadal output of the Lessepsian jellyfish Cassiopea andromeda in NW Sicily (Central Mediterranean Sea)

Knowledge of the reproductive strategy is a key prerequisite to predict population dynamics and potential invasiveness of both native and non-indigenous outbreak-forming species. In 2014 the Lessepsian upside-down jellyfish Cassiopea andromeda reached the harbor of Palermo (NW Sicily, Thyrrenian Sea), to date its established westernmost outpost in the Mediterranean Sea. To predict C. andromeda reproductive success in its novel habitat, gonad histology was carried out to record the number and size of mature and immature oocytes. Both male and female simultaneously presented gametes at all stages of development suggesting an asynchronous, yet apparently continuous, reproduction strategy. Indeed, oogenesis was observed throughout the year from pre-vitellogenic, vitellogenetic, and late-vitellogenetic to mature oocytes suggesting multiple reproductive events, as known in other Mediterranean Rhizostomeae. Oocytes were found from May to December, with two seasonal peaks of abundance (late spring = 392 and autumn = 272), suggesting imminent spawning events. Further, jellyfish size varied significantly throughout the year, with maximum diameter (up to 24 cm) in summer, and minimum diameter (6 cm) in winter. Small-sized jellyfish in winter belong to the new cohort, most probably arising from intense summer strobilation of polyps. Late spring fertilization, planula development, and metamorphosis, followed by polyp strobilation in the summer months, may explain the late appearance of a new jellyfish cohort, likely coincident with that recorded throughout winter

Open Problems on Central Simple Algebras

We provide a survey of past research and a list of open problems regarding central simple algebras and the Brauer group over a field, intended both for experts and for beginners.Comment: v2 has some small revisions to the text. Some items are re-numbered, compared to v

Through-Thickness Residual Stress Profiles in Austenitic Stainless Steel Welds: A Combined Experimental and Prediction Study

Economic and safe management of nuclear plant components relies on accurate prediction of welding-induced residual stresses. In this study, the distribution of residual stress through the thickness of austenitic stainless steel welds has been measured using neutron diffraction and the contour method. The measured data are used to validate residual stress profiles predicted by an artificial neural network approach (ANN) as a function of welding heat input and geometry. Maximum tensile stresses with magnitude close to the yield strength of the material were observed near the weld cap in both axial and hoop direction of the welds. Significant scatter of more than 200 MPa was found within the residual stress measurements at the weld center line and are associated with the geometry and welding conditions of individual weld passes. The ANN prediction is developed in an attempt to effectively quantify this phenomenon of ‘innate scatter’ and to learn the non-linear patterns in the weld residual stress profiles. Furthermore, the efficacy of the ANN method for defining through-thickness residual stress profiles in welds for application in structural integrity assessments is evaluated

First-principles calculations of the phase stability of TiO2

Published versio

HIV incidence estimate combining HIV/AIDS surveillance, testing history information and HIV test to identify recent infections in Lazio, Italy

<p>Abstract</p> <p>Background</p> <p>The application of serological methods in HIV/AIDS routine surveillance systems to identify persons with recently acquired HIV infection has been proposed as a tool which may provide an accurate description of the current transmission patterns of HIV. Using the information about recent infection it is possible to estimate HIV incidence, according to the model proposed by Karon et al. in 2008, that accounts for the effect of testing practices on the number of persons detected as recently infected.</p> <p>Methods</p> <p>We used data from HIV/AIDS surveillance in the period 2004-2008 to identify newly diagnosed persons. These were classified with recent/non-recent infection on the basis of an avidity index result, or laboratory evidence of recently acquired infection (i.e., previous documented negative HIV test within 6 months; or presence of HIV RNA or p24 antigen with simultaneous negative/indeterminate HIV antibody test). Multiple imputation was used to impute missing information. The incidence estimate was obtained as the number of persons detected as recently infected divided by the estimated probability of detection. Estimates were stratified by calendar year, transmission category, gender and nationality.</p> <p>Results</p> <p>During the period considered 3,633 new HIV diagnoses were reported to the regional surveillance system. Applying the model, we estimated that in 2004-2008 there were 5,465 new infections (95%CI: 4,538-6,461); stratifying by transmission category, the estimated number of infections was 2,599 among heterosexual contacts, 2,208 among men-who-have-sex-with-men, and 763 among injecting-drug-users. In 2008 there were 952 (625-1,229) new HIV infections (incidence of 19.9 per 100,000 person-years). In 2008, for men-who-have-sex-with-men (691 per 100,000 person-years) and injecting drug users (577 per 100,000 person-years) the incidence remained comparatively high with respect to the general population, although a decreasing pattern during 2004-2008 was observed for injecting-drug-users.</p> <p>Conclusions</p> <p>These estimates suggest that the transmission of HIV infection in Lazio remains frequent and men-who-have-sex-with men and injecting-drug-users are still greatly affected although the majority of new infections occurs among heterosexual individuals.</p