Pigment epithelium-derived factor (PEDF) and derived peptides promote survival and differentiation of photoreceptors and induce neurite-outgrowth in amacrine neurons.

Pigment epithelium-derived factor (PEDF) is a cytoprotective protein for the retina. We hypothesize that this protein acts on neuronal survival and differentiation of photoreceptor cells in culture. The purpose of the present study was to evaluate the neurotrophic effects of PEDF and its fragments in an in vitro model of cultured primary retinal neurons that die spontaneously in the absence of trophic factors. We used Wistar albino rats. Cell death was assayed by immunofluorescence and flow cytometry through TUNEL assay, propidium iodide, mitotracker, and annexin V. Immunofluorescence of cells for visualizing rhodopsin, CRX, and antisyntaxin under confocal microscopy was performed. Neurite outgrowth was also quantified. Results show that PEDF protected photoreceptor precursors from apoptosis, preserved mitochondrial function and promoted polarization of opsin enhancing their developmental process, as well as induced neurite outgrowth in amacrine neurons. These effects were abolished by an inhibitor of the PEDF receptor or receptor-derived peptides that block ligand/receptor interactions. While all the activities were specifically conferred by short peptide fragments (17 amino acid residues) derived from the PEDF neurotrophic domain, no effects were triggered by peptides from the PEDF antiangiogenic region. The observed effects on retinal neurons imply a specific activation of the PEDF receptor by a small neurotrophic region of PEDF. Our findings support the neurotrophic PEDF peptides as neuronal guardians for the retina, highlighting their potential as promoters of retinal differentiation, and inhibitors of retinal cell death and its blinding consequences. Cover Image for this issue: https://doi.org/10.1111/jnc.15089.

The Role of Optical Coherence Tomography Angiography (OCTA) in Detecting Choroidal Neovascularization in Different Stages of Best Macular Dystrophy: A Case Series.

Best macular dystrophy (BMD) is an autosomal dominant macular dystrophy of childhood onset characterized by bilateral and symmetric vitelliform lesions. Several stages of disease have been well-described in the literature. Choroidal neovascularization (CNV) has traditionally been considered a hallmark of end-stage disease, and anti-vascular endothelial growth factor (anti-VEGF) agents have been used to improve visual prognosis. While CNV was historically detected with fluorescein angiography, optical coherence tomography angiography (OCTA) has recently been employed as a novel mechanism for identifying CNV in BMD. In this case series, we discuss our institutional experience with using OCTA to detect CNV in BMD and contextualize this experience within the broader emerging literature. While OCTA allows for the identification of CNV in less severe stages of BMD, the management of this CNV remains uncertain.

Protective effects of retinoid x receptors on retina pigment epithelium cells.

Age-related macular degeneration (AMD) is among the main pathologies leading to blindness in adults and has currently no cure or effective treatment. Selective apoptosis of retina pigment epithelial (RPE) cells results in the progressive loss of photoreceptor neurons, with the consequent gradual vision loss. Oxidative stress plays an important role in this process. We have previously determined that activation of RXRs protects rat photoreceptor neurons from oxidative stress-induced apoptosis. In this study we investigated whether RXR ligands prevented apoptosis in an RPE cell line, D407 cells, exposed to hydrogen peroxide (H2O2). H2O2 induced apoptosis of D407 cells, promoting p65NFκB nuclear translocation, increasing Bax mRNA expression, activating caspase-3 and altering cell morphology. We show, for the first time, that HX630, a RXR pan-agonist, protected D407 cells from H2O2-induced apoptosis, preventing p65NFκB nuclear translocation, increasing Bclxl and PPARγ mRNA levels and simultaneously decreasing Bax mRNA levels and caspase-3 activation. Pretreatment with a RXR antagonist blocked HX630 protection. LG100754, which binds RXRs but only activates heterodimers and is an antagonist of RXR homodimers, also had a protective effect. In addition, only agonists known to bind to RXR/PPARγ were protective. As a whole, our results suggest that RXR activation protects RPE cells from oxidative stress-induced apoptosis and this protection might involve signaling through a heterodimeric receptor, such as RXR/PPARγ. These data also imply that RXR agonists might provide potential pharmacological tools for treating retina degenerative diseases.

Synthesis of docosahexaenoic acid from eicosapentaenoic acid in retina neurons protects photoreceptors from oxidative stress.

Oxidative stress is involved in activating photoreceptor death in several retinal degenerations. Docosahexaenoic acid (DHA), the major polyunsaturated fatty acid in the retina, protects cultured retina photoreceptors from apoptosis induced by oxidative stress and promotes photoreceptor differentiation. Here, we investigated whether eicosapentaenoic acid (EPA), a metabolic precursor to DHA, had similar effects and whether retinal neurons could metabolize EPA to DHA. Adding EPA to rat retina neuronal cultures increased opsin expression and protected photoreceptors from apoptosis induced by the oxidants paraquat and hydrogen peroxide (H2 O2 ). Palmitic, oleic, and arachidonic acids had no protective effect, showing the specificity for DHA. We found that EPA supplementation significantly increased DHA percentage in retinal neurons, but not EPA percentage. Photoreceptors and glial cells expressed Δ6 desaturase (FADS2), which introduces the last double bond in DHA biosynthetic pathway. Pre-treatment of neuronal cultures with CP-24879 hydrochloride, a Δ5/Δ6 desaturase inhibitor, prevented EPA-induced increase in DHA percentage and completely blocked EPA protection and its effect on photoreceptor differentiation. These results suggest that EPA promoted photoreceptor differentiation and rescued photoreceptors from oxidative stress-induced apoptosis through its elongation and desaturation to DHA. Our data show, for the first time, that isolated retinal neurons can synthesize DHA in culture. Docosahexaenoic acid (DHA), the major polyunsaturated fatty acid in retina photoreceptors, and its precursor, eicosapentaenoic acid (EPA) have multiple beneficial effects. Here, we show that retina neurons in vitro express the desaturase FADS2 and can synthesize DHA from EPA. Moreover, addition of EPA to these cultures protects photoreceptors from oxidative stress and promotes their differentiation through its metabolization to DHA.

Light, lipids and photoreceptor survival: live or let die?

Due to its constant exposure to light and its high oxygen consumption the retina is highly sensitive to oxidative damage, which is a common factor in inducing the death of photoreceptors after light damage or in inherited retinal degenerations. The high content of docosahexaenoic acid (DHA), the major polyunsaturated fatty acid in the retina, has been suggested to contribute to this sensitivity. DHA is crucial for developing and preserving normal visual function. However, further roles of DHA in the retina are still controversial. Current data support that it can tilt the scale either towards degeneration or survival of retinal cells. DHA peroxidation products can be deleterious to the retina and might lead to retinal degeneration. However, DHA has also been shown to act as, or to be the source of, a survival molecule that protects photoreceptors and retinal pigment epithelium cells from oxidative damage. We have established that DHA protects photoreceptors from oxidative stress-induced apoptosis and promotes their differentiation in vitro. DHA activates the retinoid X receptor (RXR) and the ERK/MAPK pathway, thus regulating the expression of anti and pro-apoptotic proteins. It also orchestrates a diversity of signaling pathways, modulating enzymatic pathways that control the sphingolipid metabolism and activate antioxidant defense mechanisms to promote photoreceptor survival and development. A deeper comprehension of DHA signaling pathways and context-dependent behavior is required to understand its dual functions in retinal physiology.

Activation of retinoid X receptors protects retinal neurons and pigment epithelial cells from BMAA-induced death.

Exposure to the non-protein amino acid cyanotoxin ß-N-methylamino-L-alanine (BMAA), released by cyanobacteria found in many water reservoirs has been associated with neurodegenerative diseases. We previously demonstrated that BMAA induced cell death in both retina photoreceptors (PHRs) and amacrine neurons by triggering different molecular pathways, as activation of NMDA receptors and formation of carbamate-adducts was only observed in amacrine cell death. We established that activation of Retinoid X Receptors (RXR) protects retinal cells, including retina pigment epithelial (RPE) cells from oxidative stress-induced apoptosis. We now investigated the mechanisms underlying BMAA toxicity in these cells and those involved in RXR protection. BMAA addition to rat retinal neurons during early development in vitro increased reactive oxygen species (ROS) generation and polyADP ribose polymers (PAR) formation, while pre-treatment with serine (Ser) before BMAA addition decreased PHR death. Notably, RXR activation with the HX630 agonist prevented BMAA-induced death in both neuronal types, reducing ROS generation, preserving mitochondrial potential, and decreasing TUNEL-positive cells and PAR formation. This suggests that BMAA promoted PHR death by substituting Ser in polypeptide chains and by inducing polyADP ribose polymerase activation. BMAA induced cell death in ARPE-19 cells, a human epithelial cell line; RXR activation prevented this death, decreasing ROS generation and caspase 3/7 activity. These findings suggest that RXR activation prevents BMAA harmful effects on retinal neurons and RPE cells, supporting this activation as a broad-spectrum strategy for treating retina degenerations.

Retinoid X receptor activation is essential for docosahexaenoic acid protection of retina photoreceptors.

We have established that docosahexaenoic acid (DHA), the major polyunsaturated fatty acid in the retina, promotes survival of rat retina photoreceptors during early development in vitro and upon oxidative stress by activating the ERK/MAPK signaling pathway. Here we have investigated whether DHA turns on this pathway through activation of retinoid X receptors (RXRs) or by inducing tyrosine kinase (Trk) receptor activation. We also evaluated whether DHA release from phospholipids was required for its protective effect. Addition of RXR antagonists (HX531, PA452) to rat retinal neuronal cultures inhibited DHA protection during early development in vitro and upon oxidative stress induced with Paraquat or H2O2. In contrast, the Trk inhibitor K252a did not affect DHA prevention of photoreceptor apoptosis. These results imply that activation of RXRs was required for DHA protection whereas Trk receptors were not involved in this protection. Pretreatment with 4-bromoenol lactone, a phospholipase A2 inhibitor, blocked DHA prevention of oxidative stress-induced apoptosis of photoreceptors. It is noteworthy that RXR agonists (HX630, PA024) also rescued photoreceptors from H2O2-induced apoptosis. These results provide the first evidence that activation of RXRs prevents photoreceptor apoptosis and suggest that DHA is first released from phospholipids and then activates RXRs to promote the survival of photoreceptors.

Natural radioactivity in building materials, indoor radon measurements, and assessment of the associated risk indicators in some localities of the Centre Region, Cameroon.

The objective of the current study is to investigate the natural radioactivity of some building materials, the resulting long-term external and internal effective dose equivalents (EEDE and IEDE) analysis followed by indoor radon measurements, and the assessment of some radiological risk indicators associated with radon exposure. A total of 37 samples of building materials were analyzed with a sodium iodide detector (NaI (Tl)), and the computer code RESRAD-BUILD was used for the analysis of the EEDE and IEDE of the structural elements of the houses (walls and floor). For indoor radon measurements, 140 houses were selected, and in each of them was placed 01 RADTRAK dosimeter. Inhalation dose, total dose, and some radiological risk indicators were calculated. The specific activities of 226Ra, 232Th, and 40K for the overall sampled building materials were found to vary between 10 ± 2-52 ± 7, 10 ± 1-95 ± 10, and 31 ± 1-673 ± 20 Bq kg-1, respectively. The dwelling types with bare brick walls, cement mortar plastered walls, and concrete floors show EEDE and IEDE values well below the recommended limits. The corresponding dwelling type contributions to the measured average indoor radon concentration (42 ± 12 Bq m-3) are 22%, 13%, and 16%, respectively. Inhalation dose resulting from the measured indoor radon concentrations varies from 0.35 to 3.24 mSv y-1 with a mean value of 0.96 ± 0.55 mSv y-1, which represents about 65% of the total dose simulated (1.49 ± 0.88 mSv y-1) by the RESRAD-BUILD code. The overall analysis of indoor radon-related radiological risk indicators shows low levels of risk relative to permissible limits.

The use of multimodal imaging in the evaluation of a patient with central retinal artery occlusion in the setting of asteroid hyalosis: a case report.

BACKGROUND: A central retinal artery occlusion (CRAO) is an ophthalmic emergency due to its strong association with cerebrovascular and cardiovascular morbidity and mortality. A timely diagnosis is necessary but difficult in the setting of dense asteroid hyalosis, as typical fundoscopic findings can be obscured. We present a case where multimodal imaging in an eye with an obscured fundus could lead to timely diagnosis and management of CRAO in a patient with acute vision loss. CASE PRESENTATION: A 94-year-old Caucasian woman with a history of exudative macular degeneration presented to the retina clinic with acute vision loss in one eye over the course of an afternoon. The patient had dense asteroid hyalosis, and a direct retinal exam was not possible. Multimodal imaging suggested a CRAO diagnosis. The patient received digital ocular massage directly prior to undergoing fluorescein angiography (FANG), which confirmed the diagnosis. The patient was transported from clinic to the emergency room for an emergency stroke workup, which revealed a spontaneous echo in the left atrial appendage, and the patient was started on antiplatelet therapy. When she presented for follow-up within a week, the patient noted that her vision had improved at the time of digital ocular massage and continued to improve thereafter. Her FANG showed marked reperfusion of the retina, and she subsequently has completely regained her baseline visual acuity. CONCLUSIONS: Multimodal imaging is useful in evaluating visual loss in patients with acute vision loss. In addition, ocular massage is a simple, low-risk intervention that may have benefit in the treatment of acute CRAO. Patients who present to ophthalmologists with an acute CRAO need an emergency referral for evaluation of cerebrovascular and cardiovascular comorbidities.

Retina stem cells, hopes and obstacles.

Retinal degeneration is a major contributor to visual dysfunction worldwide. Although it comprises several eye diseases, loss of retinal pigment epithelial (RPE) and photoreceptor cells are the major contributors to their pathogenesis. Early therapies included diverse treatments, such as provision of anti-vascular endothelial growth factor and many survival and trophic factors that, in some cases, slow down the progression of the degeneration, but do not effectively prevent it. The finding of stem cells (SC) in the eye has led to the proposal of cell replacement strategies for retina degeneration. Therapies using different types of SC, such as retinal progenitor cells (RPCs), embryonic SC, pluripotent SCs (PSCs), induced PSCs (iPSCs), and mesenchymal stromal cells, capable of self-renewal and of differentiating into multiple cell types, have gained ample support. Numerous preclinical studies have assessed transplantation of SC in animal models, with encouraging results. The aim of this work is to revise the different preclinical and clinical approaches, analyzing the SC type used, their efficacy, safety, cell attachment and integration, absence of tumor formation and immunorejection, in order to establish which were the most relevant and successful. In addition, we examine the questions and concerns still open in the field. The data demonstrate the existence of two main approaches, aimed at replacing either RPE cells or photoreceptors. Emerging evidence suggests that RPCs and iPSC are the best candidates, presenting no ethical concerns and a low risk of immunorejection. Clinical trials have already supported the safety and efficacy of SC treatments. Serious concerns are pending, such as the risk of tumor formation, lack of attachment or integration of transplanted cells into host retinas, immunorejection, cell death, and also ethical. However, the amazing progress in the field in the last few years makes it possible to envisage safe and effective treatments to restore vision loss in a near future.

Retinoid X receptor activation promotes photoreceptor survival and modulates the inflammatory response in a mouse model of retinitis pigmentosa.

Photoreceptor cell (PHR) death is a hallmark of most retinal neurodegenerative diseases, in which inflammation plays a critical role. Activation of retinoid X receptors (RXR) modulates and integrates multiple cell functions, and has beneficial effects in animal models of chronic inflammatory diseases. Nonetheless, the mechanisms involved and their role in retina neuroprotection are poorly understood. In this work we assessed whether RXR activation prevents inflammation and/or PHR death in retinitis pigmentosa, an inherited retina neurodegeneration, using as an ex vivo model, retinas from the rd1 mice, a murine model of this disease. We demonstrated that rd1 retinas had lower levels of RXR alpha isoform than their wt counterparts at early developmental times, whereas its distribution pattern remained similar. In mixed neuro-glial cultures obtained from either rd1 or wt retinas, both PHR and Müller glial cells (MGC) expressed RXRalpha, and RXR activation by its synthetic pan-agonist PA024 selectively increased mRNA levels of RXRgamma isoform. PA024 decreased PHR death in rd1 mixed cultures; it reduced the amount of non-viable neurons, delayed the onset of PHR apoptosis, and decreased Bax mRNA levels. PA024 also reduced MGC reactivity in vitro before and at the onset of degeneration, decreasing GFAP expression, increasing glutamine synthetase mRNA levels, and promoting the transcription of the anti-inflammatory cytokine, Il-10. These results suggest that RXR activation rescues rd1 PHR and decreases MGC reactivity, promoting an anti-inflammatory environment in the rd1 retina, thus supporting the potential of RXR agonists as pharmacological tools for treating retina degenerative diseases.

Self-Reported Awareness of Retinopathy Severity in Diabetic Patients.

BACKGROUND: Diabetic retinopathy (DR) may be asymptomatic in both mild and advanced stages. A patient's accurate perception of their DR severity may therefore be critical for effective self-management behaviors and understanding the need for timely intervention and follow-up. PURPOSE: To evaluate the relationship between self-reported and actual retinopathy severity in diabetic patients. METHODS: This study was a single-center cross-sectional survey. Diabetic patients identified by enterprise data warehouse were sent an online questionnaire where they were asked to self-assess for presence of DR and grade their severity. Actual DR grading was determined via chart review. The primary outcome measures were patient-assessed DR severity and agreement with actual DR severity. RESULTS: Of 3208 invitations sent, 324 (10%) patients responded and 319 responses were analyzed. The data showed that 39 of 253 (15%) with no DR, 26 of 40 (65%) with mild/moderate DR, and 24 of 26 (92%) with severe DR believed they had DR (p<0.001). Of those with no DR, 214 of 253 (85%) accurately assessed absence of DR. Of those with mild/moderate DR, 25 of 40 (63%) accurately assessed their severity, 14 of 40 (35%) believed they had no DR, and 1 of 40 (3%) believed they had severe DR. In patients with severe DR, 9 of 26 (35%) correctly assessed their severity, 15 of 26 (58%) believed they had mild/moderate DR, and 2 of 26 (8%) believed they had no DR. CONCLUSION: Patients with severe DR were the most likely to report presence of DR, but often underestimated their disease severity. Many with mild/moderate DR did not realize they had DR. This consistent underestimation of severity across all a significant barrier to timely follow-up and treatment necessary to prevent future visual impairment.

Indoor Radon Measurements Using Radon Track Detectors and Electret Ionization Chambers in the Bauxite-Bearing Areas of Southern Adamawa, Cameroon.

The current work deals with indoor radon (222Rn) concentrations and ambient dose-equivalent rate measurements in the bauxite-bearing areas of the Adamawa region in Cameroon before mining from 2022. In total, 90 Electret Ionization Chambers (EIC) (commercially, EPERM) and 175 Radon Track Detectors (commercially, RADTRAK2) were used to measure 222Rn concentrations in dwellings of four localities of the above region. A pocket survey meter (RadEye PRD-ER, Thermo Scientific, Waltham, MA, USA) was used for the ambient dose-equivalent rate measurements. These measurements were followed by calculations of annual doses from inhalation and external exposure. 222Rn concentrations were found to vary between 36 ± 8-687 ± 35 Bq m-3 with a geometric mean (GM) of 175 ± 16 Bq m-3 and 43 ± 12-270 ± 40 Bq m-3 with a geometric mean of 101 ± 21 Bq m-3 by using EPERM and RADTRAK, respectively. According to RADTRAK data, 51% of dwellings have radon concentrations above the reference level of 100 Bq m-3 recommended by the World Health Organization (WHO). The ambient dose equivalent rate ranged between 0.04-0.17 µSv h-1 with the average value of 0.08 µSv h-1. The inhalation dose and annual external effective dose to the public were assessed and found to vary between 0.8-5 mSv with an average value of 2 mSv and 0.3-1.8 mSv with an average value of 0.7 mSv, respectively. Most of the average values in terms of concentration and radiation dose were found to be above the corresponding world averages given by the United Nations Scientific Commission on the Effects of Atomic Radiation (UNSCEAR). Even though the current exposure of members of the public to natural radiation is not critical, the situation could change abruptly when mining starts.

A Defective Crosstalk Between Neurons and Müller Glial Cells in the rd1 Retina Impairs the Regenerative Potential of Glial Stem Cells.

Müller glial cells (MGC) are stem cells in the retina. Although their regenerative capacity is very low in mammals, the use of MGC as stem cells to regenerate photoreceptors (PHRs) during retina degenerations, such as in retinitis pigmentosa, is being intensely studied. Changes affecting PHRs in diseased retinas have been thoroughly investigated; however, whether MGC are also affected is still unclear. We here investigated whether MGC in retinal degeneration 1 (rd1) mouse, an animal model of retinitis pigmentosa, have impaired stem cell properties or structure. rd1 MGC showed an altered morphology, both in culture and in the whole retina. Using mixed neuron-glial cultures obtained from newborn mice retinas, we determined that proliferation was significantly lower in rd1 than in wild type (wt) MGC. Levels of stem cell markers, such as Nestin and Sox2, were also markedly reduced in rd1 MGC compared to wt MGC in neuron-glial cultures and in retina cryosections, even before the onset of PHR degeneration. We then investigated whether neuron-glial crosstalk was involved in these changes. Noteworthy, Nestin expression was restored in rd1 MGC in co-culture with wt neurons. Conversely, Nestin expression decreased in wt MGC in co-culture with rd1 neurons, as occurred in rd1 MGC in rd1 neuron-glial mixed cultures. These results imply that MGC proliferation and stem cell markers are reduced in rd1 retinas and might be restored by their interaction with "healthy" PHRs, suggesting that alterations in rd1 PHRs lead to a disruption in neuron-glial crosstalk affecting the regenerative potential of MGC.

Retinal pigment epithelial cells promote spatial reorganization and differentiation of retina photoreceptors.

Retina differentiation involves the acquisition of a precise layered arrangement, with RPE cells in the first layer in intimate contact with photoreceptors in the second layer. Here, we developed an in vitro coculture model, to test the hypothesis that RPE cells play a pivotal role in organizing the spatial structure of the retina. We cocultured rat retinal neurons with ARPE-19 epithelial cells under various experimental conditions. Strikingly, when seeded over RPE cells, photoreceptors attached to their apical surfaces and proceeded with their development, including the increased synthesis of rhodopsin. Conversely, when we seeded RPE cells over neurons, the RPE cells rapidly detached photoreceptors from their substrata and positioned themselves underneath, thus restoring the normal in vivo arrangement. Treatment with the metalloproteinase inhibitor TIMP-1 blocked this reorganization, suggesting the involvement of metalloproteinases in this process. Reorganization was highly selective for photoreceptors because 98% of photoreceptors but very few amacrine neurons were found to redistribute on top of RPE cells. Interestingly, RPE cells were much more efficient than other epithelial or nonepithelial cells in promoting this reorganization. RPE cells also promoted the growth of photoreceptor axons away from them. An additional factor that contributed to the distal arrangement of photoreceptor axons was the migration of photoreceptor cell bodies along their own neurites toward the RPE cells. Our results demonstrate that RPE and photoreceptor cells interact in vitro in very specific ways. They also show that in vitro studies may provide important insights into the process of pattern formation in the retina.

Vigilancia epidemiológica de COVID-19 mediante testeos agrupados focalizada en trabajadores de salud asintomáticos en Bahía Blanca / Epidemiological surveillance of COVID-19 through group testing focused on asymptomatic health workers in Bahía Blanca

INTRODUCCIÓN: Uno de los desafíos más relevantes al comienzo de la pandemia consistió en implementar estrategias dirigidas a prevenir la transmisión del virus SARS-CoV-2 y mitigar el impacto de la COVID-19. El propósito de este estudio fue contribuir a reducir la transmisión comunitaria a través de una iniciativa interinstitucional, cuyos objetivos fueron validar un método de detección de ARN del SARS-CoV-2 e implementar y evaluar la vigilancia en trabajadores de salud asintomáticos de instituciones de salud pública de Bahía Blanca. MÉTODOS: Se validó una prueba de detección del gen E del coronavirus mediante RT-PCR a partir de ARN aislado de hisopados nasofaríngeos. Para aumentar la capacidad de testeo se validó la detección del ARN viral en muestras agrupadas (pooles). Se realizó un estudio de cohorte prospectiva entre el 15/09/20 y el 15/09/21. RESULTADOS: La sensibilidad y especificidad de la prueba en muestras individuales fue del 95% (IC 95%: 85%-100%). La sensibilidad de la detección en pooles fue del 73% (IC 95%: 46%-99%) y la especificidad, del 100%. A lo largo de la vigilancia se incluyeron 855 trabajadores y 1764 hisopados, con una incidencia acumulada anual de 2,3% (IC 95%: 1,2%-3,4%). Se detectaron 20 casos asintomáticos positivos. DISCUSIÓN: El tamizaje de trabajadores de salud asintomáticos en la pandemia contribuyó a reducir el riesgo de brotes hospitalarios. Asimismo, se generó un marco de trabajo interdisciplinario aplicable a otros problemas de salud.

COVID-19 epidemiological surveillance through pool testing focused on asymptomatic healthcare workers in the city of Bahía Blanca / Vigilancia epidemiológica de covid-19 mediante testeos agrupados focalizada en trabajadores de salud asintomáticos en bahía blanca

RESUMEN INTRODUCCIÓN: Uno de los desafíos más relevantes al comienzo de la pandemia consistió en implementar estrategias dirigidas a prevenir la transmisión del virus SARS-CoV-2 y mitigar el impacto de la COVID-19. El propósito de este estudio fue contribuir a reducir la transmisión comunitaria a través de una iniciativa interinstitucional, cuyos objetivos fueron validar un método de detección de ARN del SARS-CoV-2 e implementar y evaluar la vigilancia en trabajadores de salud asintomáticos de instituciones de salud pública de Bahía Blanca. MÉTODOS: Se validó una prueba de detección del gen E del coronavirus mediante RT-PCR a partir de ARN aislado de hisopados nasofaríngeos. Para aumentar la capacidad de testeo se validó la detección del ARN viral en muestras agrupadas (pooles). Se realizó un estudio de cohorte prospectiva entre el 15/09/20 y el 15/09/21. RESULTADOS: La sensibilidad y especificidad de la prueba en muestras individuales fue del 95% (IC 95%: 85%-100%). La sensibilidad de la detección en pooles fue del 73% (IC 95%: 46%-99%) y la especificidad, del 100%. A lo largo de la vigilancia se incluyeron 855 trabajadores y 1764 hisopados, con una incidencia acumulada anual de 2,3% (IC 95%: 1,2%-3,4%). Se detectaron 20 casos asintomáticos positivos. DISCUSIÓN: El tamizaje de trabajadores de salud asintomáticos en la pandemia contribuyó a reducir el riesgo de brotes hospitalarios. Asimismo, se generó un marco de trabajo interdisciplinario aplicable a otros problemas de salud.

ABSTRACT INTRODUCTION: One of the most important challenges at the beginning of the pandemic was to implement strategies to prevent SARS-CoV-2 virus transmission and reduce the impact of COVID-19. The purpose of this study was to contribute to the reduction of community transmission through an interinstitutional initiative, aimed at validating a SARS-CoV-2 RNA detection method, and at implementing and assessing the surveillance of asymptomatic infected healthcare workers (HCWs) at public health institutions in the city of Bahía Blanca. METHODS: To validate a coronavirus RNA detection method, RNA was extracted from nasopharyngeal swabs and identification of the viral E gene was done by RT-PCR. Validation of sample pooling was performed to increase the testing capacity. A prospective cohort study was conducted from 15 September 2020 to 15 September 2021. RESULTS: The sensitivity and specificity of the test in individual samples was 95% (CI 95%: 85%-100%). The sensitivity of the pooling strategy was 73% (CI 95%: 46%-99%) and the specificity was 100%. A total of the 855 HCWs were included in the surveillance and 1764 swabs were performed, with an annual cumulative incidence of 2.3% (CI 95%: 1,2%-3,4%), and 20 positive asymptomatic cases were detected. DISCUSSION: The screening of asymptomatic HCWs during the pandemic contributed to reduce the risk of outbreaks in hospital settings. Moreover, an interdisciplinary team framework applicable to other health problems was generated.

Ceramide is a mediator of apoptosis in retina photoreceptors.

PURPOSE: The precise mechanisms involved in photoreceptor apoptosis are still unclear. In the present study, the role of ceramide, a sphingolipid precursor that induces apoptosis on cellular stress, was investigated in relation to the activation of cell death in photoreceptors. METHODS: Rat retina neuronal cultures, with or without docosahexaenoic acid (DHA), were treated with the ceramide analogue acetylsphingosine (C2-ceramide), and with a glucosylceramide synthase inhibitor. Ceramide synthesis in cultures treated with the oxidant paraquat was evaluated with [3H]palmitate. The effect of inhibitors of ceramide de novo synthesis, fumonisin B1 and cycloserine, on photoreceptor apoptosis was investigated. Apoptosis, mitochondrial membrane potential, and Bcl-2 expression were determined. RESULTS: Addition of C2-ceramide induced photoreceptor apoptosis. Paraquat increased formation of [3H]ceramide in photoreceptors, compared with the control, whereas inhibition of ceramide synthesis, immediately before paraquat treatment, prevented paraquat-induced photoreceptor apoptosis. Fumonisin also reduced photoreceptor apoptosis during early development in vitro. DHA, the retina major polyunsaturated fatty acid, which protects photoreceptors from oxidative stress-induced apoptosis, completely blocked C2-ceramide-induced photoreceptor death, simultaneously increasing Bcl-2 expression. Inhibiting glucosylceramide synthase, which catalyzes ceramide glucosylation, before ceramide or paraquat treatment blocked DHA's protective effect. CONCLUSIONS: The results suggest that oxidative stress stimulated an increase in ceramide levels that induced photoreceptor apoptosis. DHA prevented oxidative stress and ceramide damage by upregulating Bcl-2 expression and glucosylating ceramide, thus decreasing its intracellular concentration. This shows for the first time that ceramide is a critical mediator for triggering photoreceptor apoptosis in mammalian retina and suggests that modulating ceramide levels may provide a therapeutic tool for preventing photoreceptor death in neurodegenerative diseases.

Pro-apoptotic effects of parathyroid hormone in intestinal cells.

Apoptosis, a form of programmed cell death, is a process fundamental to normal growth and development, immune response, tissue remodeling after injury or insult, and homeostasis of the intestinal epithelium. Recently, it has become apparent that apoptosis is a crucial process in skeletal development and homeostasis, and that signaling by the parathyroid hormone (PTH) receptor can either promote or suppress apoptosis depending on the cellular context. In this study, we evaluated the role of PTH in intestinal apoptosis using human colonic Caco-2 cells. To that end, Caco-2 cells were exposed to PTH (10-8 mol/L) for 1-5 days. Evaluation of cell survival by use of resazurin staining, 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) staining, and crystal violet staining revealed that PTH treatment diminishes the number of viable cells. Assessment of cells after PTH treatment by use of propidium iodide showed that the hormone increased the number of red-stained (dead) cells (178%, 5 days). Moreover, we found that the hormone induced disruption of actin filaments with changes to cellular shape, alteration of cell-to-cell junctions, externalization of membrane phosphatidylserine, chromatin condensation, and DNA fragmentation of the nucleus, which are morphological features consistent with apoptosis. In addition, PTH treatment revealed a cytosolic staining pattern of 14-3-3. However, the significance of such differential localization for 14-3-3 function remains unknown. Taken together, the present study suggests that PTH promotes apoptosis in Caco-2 intestinal cells.