Validation and shortcomings of the most common mouse model of chronic rhinosinusitis with nasal polyps

BACKGROUND: Chronic rhinosinusitis (CRS) is a highly prevalent airway disease worldwide. Whereas eosinophilic CRS with nasal polyps (eCRSwNP) represents its most severe phenotype, pathogenic mechanisms remain poorly understood despite a wide spectrum of in vitro and in vivo experimental models. A mouse model of experimental ovalbumin (OVA)-induced airway allergy with coadministration of Staphylococcus aureus enterotoxin B (SEB) has been widely used to study eosinophilic eCRSwNP. This study revisits the features of this model and its suitability for studying eCRS. METHODOLOGY: We implemented the most used eCRSwNP mouse model based on OVA+SEB intranasal challenges. Readouts including inflammatory features by (immuno)histology of the sinonasal epithelium (NP formation, eosinophils, epithelial and basement membrane thickness, fibrosis, goblet cells, Charcot-Leyden crystals (CLC)-like, tight junctions) and IgE production by enzyme-linked immunosorbent assay (ELISA), were compared to features of the corresponding human disease. RESULTS: The OVA+SEB model induced eosinophilic inflammation of upper and lower airways, with epithelial and basement membrane thickening, goblet cell hyperplasia and subepithelial fibrosis in the sinuses, along increased IgE production. Except local IgE in nasal lavage (NL), which was only increased in OVA+SEB group, all other features did not differ between OVA and OVA+SEB groups. Macro- or microscopic NP were not detected. CONCLUSIONS: With the notable exception of local IgE production, the addition of SEB did not induce additional inflammatory or structural change in the sinuses from mice exposed to and challenged with OVA. This model might represent a model for severe upper airway allergy rather than a specific model of human eCRSwNP.

Anaphylaxis: Mediators, Biomarkers, and Microenvironments.

The life-threatening nature of anaphylactic reactions has increased interest in discovering new biomarkers that could improve diagnosis and prevention. However, the diverse nature of the clinical features and the etiology and pathogenesis of anaphylaxis hinder the identification of valuable molecular indicators of disease. Most studies on anaphylaxis focus on the immune system. Anaphylactic reactions are characterized primarily by IgE-mediated activation of mast cells and basophils and release of mediators. Determination of serum tryptase levels is the main in vitro test used to confirm the reaction, although there are no biomarkers that can predict it. Nevertheless, recent research has postulated that alternative pathways, cell types, and systems are involved. Consequently, various molecular products have been explored and considered potential biomarkers, although none of them are yet used in clinical practice. The products that are altered in patients with anaphylaxis include vasoactive agents, proteases, proteoglycans, lipids, interleukins, cytokines, products of the complement-contact and coagulation systems, circulating proteins, extracellular vesicles, microRNAs, and metabolites. The recognition of biological processes and molecular pathways affecting the microenvironments involved in anaphylaxis will considerably improve clinical practice and the identification of better molecular markers. We offer a broad review of the various mediators described in anaphylaxis, consider their usefulness as potential biomarkers of this pathological event, and examine their role in the molecular basis of the reaction.

Microsurgical treatment of trigeminal neuralgia in patients older than 70 years: An efficacy and safety study. / Tratamiento microquirúrgico de la neuralgia trigeminal en mayores de 70 años, estudio de eficacia y seguridad.

INTRODUCTION: The increasing incidence of trigeminal neuralgia (TN) with age together with population ageing call for reexamination of surgical treatment options for refractory TN in elderly patients. METHODS: Retrospective review of a consecutive series of patients older than 70 who underwent microvascular decompression (MVD) for refractory TN between 1997 and 2015. Outcomes based on the Barrow Neurological Institute pain intensity score (BNI score) and surgical complications were compared to those of patients younger than 70 undergoing MVD in the same period. RESULTS: Forty patients older than 70 (mean = 74.8 years) underwent interventions. At a mean follow-up time of 34 months, 73% of the patients presented complete absence of pain without medication (BNI I) and 85% had good pain control with or without medication (BNI I-III). A comparison of these patients with the 85 patients younger than 70 treated surgically during the same period did not find a significant association between age and achievement of pain control (BNI I-II). However, there was a significant association between age older than 70 and complete pain relief (BNI I; P=.03). The mean hospital stay in patients over 70 was also significantly longer (P=.04), although the postsurgical complication rate was similar to that in younger patients. CONCLUSIONS: Elderly patients with refractory TN may benefit from treatment with MVD and the probability of success and surgical risk are comparable to those in younger patients.

Three new sensitive and specific heat-shock protein 70 PCRs for global Leishmania species identification.

The heat-shock protein 70 gene (hsp70) has been exploited for Leishmania species identification in the New and Old World, using PCR followed by restriction fragment length polymorphism (RFLP) analysis. Current PCR presents limitations in terms of sensitivity, which hampers its use for analyzing clinical and biological samples, and specificity, which makes it inappropriate to discriminate between Leishmania and other trypanosomatids. The aim of the study was to improve the sensitivity and specificity of a previously reported hsp70 PCR using alternative PCR primers and RFLPs. Following in silico analysis of available sequences, three new PCR primer sets and restriction digest schemes were tested on a globally representative panel of 114 Leishmania strains, various other infectious agents, and clinical samples. The largest new PCR fragment retained the discriminatory power from RFLP, while two smaller fragments discriminated less species. The detection limit of the new PCRs was between 0.05 and 0.5 parasite genomes, they amplified clinical samples more efficiently, and were Leishmania specific. We succeeded in significantly improving the specificity and sensitivity of the PCRs for hsp70 Leishmania species typing. The improved PCR-RFLP assays can impact diagnosis, treatment, and epidemiological studies of leishmaniasis in any setting worldwide.

Microbialites show a distinct cyanobacterial phylogenetic structure and functional redundancy in Bacalar lagoon and Cenote Azul sinkhole, Yucatan Peninsula, Mexico.

Cyanobacterial components of microbialites from two geographically close systems, the Bacalar lagoon (BL) and the Cenote Azul sinkhole (CA) in Quintana Roo, Mexico, were characterized. BL and CA systems were studied along a longitudinal gradient (north to south) and a depth gradient (5-30 m), respectively. Microscopic observations, 16S rRNA amplicon sequencing, and shotgun metagenomics were used to characterize Cyanobacteria. Both systems showed similar metabolic/functional profiles but harbored completely different cyanobacterial taxa. BL was dominated by Nostocales, including a population of previously undescribed Chakia sp., while CA was dominated by an unknown taxon of Chroococcales, comprising 70% of relative abundance through all depths. Interestingly, cyanobacterial assemblages in microbialites exhibited phylogenetic overdispersion in most of the BL sites, while CA sites exhibited phylogenetic clustering, these differences were attributed to depth/light conditions and possibly different times of geological formation for BL and CA systems.

First detection of SARS-CoV-2 variant B.1.1.529 (Omicron) in Ecuador.

The National Institute of Research and Public Health reported the first local record of the Omicron variant detected in Ecuador. A fully vaccinated subject returned from South Africa with a negative RT-PCR. We present the cumulative frequency of the variants in Ecuador and a phylogenetic analysis of this new Omicron.

Response of RAW 264.7 and J774A.1 macrophages to particles and nanoparticles of a mesoporous bioactive glass: A comparative study.

Mesoporous bioactive glasses (MBGs) are bioceramics designed to induce bone tissue regeneration and very useful materials with the ability to act as drug delivery systems. MBGs can be implanted in contact with bone tissue in different ways, as particulate material, in 3D scaffolds or as nanospheres. In this work, we assessed the effects of particles of mesoporous bioactive glass MBG-75S and mesoporous nanospheres NanoMBG-75S on RAW 264.7 and J774A.1 macrophages, which present different sensitivity and are considered as ideal models for the study of innate immune response. After evaluating several cellular parameters (morphology, size, complexity, proliferation, cell cycle and intracellular content of reactive oxygen species), the action of MBG-75S particles and NanoMBG-75S on the polarization of these macrophages towards the pro-inflammatory (M1) or reparative (M2) phenotype was determined by the expression of specific M1 (CD80) and M2 (CD206, CD163) markers. We previously measured the adsorption of albumin and fibrinogen on MBG-75S particles and the production of pro-inflammatory cytokines as TNF-α and IL-6 by macrophages in response to these particles. This comparative study demonstrates that particles of mesoporous bioactive glass MBG-75S and mesoporous nanospheres NanoMBG-75S allow the appropriated development and function of RAW 264.7 and J774A.1 macrophages and do not induce polarization towards the M1 pro-inflammatory phenotype. Therefore, considering that these mesoporous biomaterials offer the possibility of loading drugs into their pores, the results obtained indicate their high potential for use as drug-delivery systems in bone repair and osteoporosis treatments without triggering an adverse inflammatory response.

Heat-shock protein 70 PCR-RFLP: a universal simple tool for Leishmania species discrimination in the New and Old World.

INTRODUCTION: Species typing in leishmaniasis gains importance in diagnostics, epidemiology, and clinical studies. A restriction fragment length polymorphism (RFLP) assay of PCR amplicons from a partial heat-shock protein 70 gene (hsp70) had been described for the New World, allowing identification of some species. METHODS: Based on an initial in silico analysis of 51 hsp70 sequences, most of which we recently determined in the frame of a phylogenetic study, species-specific restriction sites were identified. These were tested by PCR-RFLP on 139 strains from 14 species, thereby documenting both inter- and intra-species variability. RESULTS: Our assay could identify Leishmania infantum, L. donovani, L. tropica, L. aethiopica, L. major, L. lainsoni, L. naiffi, L. braziliensis, L. peruviana, L. guyanensis, and L. panamensis by applying 2 subsequent digests. L. mexicana, L. amazonensis, and L. garnhami did not generate species-specific restriction fragment patterns. CONCLUSION: Currently no assay is available for global Leishmania species discrimination. We present a universal PCR-RFLP method allowing identification of most medically relevant Old and New World Leishmania species on the basis of a single PCR, obviating the need to perform separate PCRs. The technique is simple to perform and can be implemented in all settings where PCR is available.

Hydroxylation of demethoxy-Q6 constitutes a control point in yeast coenzyme Q6 biosynthesis.

Coenzyme Q is a lipid molecule required for respiration and antioxidant protection. Q biosynthesis in Saccharomyces cerevisiae requires nine proteins (Coq1p-Coq9p). We demonstrate in this study that Q levels are modulated during growth by its conversion from demethoxy-Q (DMQ), a late intermediate. Similar conversion was produced when cells were subjected to oxidative stress conditions. Changes in Q(6)/DMQ(6) ratio were accompanied by changes in COQ7 gene mRNA levels encoding the protein responsible for the DMQ hydroxylation, the penultimate step in Q biosynthesis pathway. Yeast coq null mutant failed to accumulate any Q late biosynthetic intermediate. However, in coq7 mutants the addition of exogenous Q produces the DMQ synthesis. Similar effect was produced by over-expressing ABC1/COQ8. These results support the existence of a biosynthetic complex that allows the DMQ(6) accumulation and suggest that Coq7p is a control point for the Q biosynthesis regulation in yeast.

In vitro and in vivo cytokeratin patterns of expression in bioengineered human periodontal mucosa.

BACKGROUND AND OBJECTIVE: Development of human oral mucosa substitutes by tissue engineering may provide new therapeutic tools for the management of periodontal diseases. In this study we evaluated a fibrin-agarose human oral mucosa substitute both in vitro and in vivo. MATERIAL AND METHODS: In vitro bioengineered oral mucosa substitutes were developed from irrelevant biopsy samples of human oral gingiva. In vivo evaluation of the constructed tissues was performed by implantation into athymic nude mice. The expression of several epithelial markers was assessed by microarray analysis and immunohistochemistry. RESULTS: Bioengineered oral mucosa samples kept in vitro developed a multilayered epithelium that expressed several cytokeratins, including some markers of simple epithelia (cytokeratins 7, 8 and 18), along with markers of stratified epithelia (cytokeratins 5 and 13) and of cell proliferation (proliferating cell nuclear antigen). Bioengineered tissues grafted in vivo onto nude mice exhibited very good biointegration with the host, showing a cytokeratin expression pattern that was very similar to that of normal native oral mucosa controls. Histological analysis of the artificial tissues demonstrated that oral mucosa substitutes evaluated in vivo were structurally mature, showing some typical structures of human native oral mucosa such as rete ridges and chorial papillae, along with numerous blood vessels at the fibrin-agarose stromal substitute. These structures were absent in samples evaluated in vitro. CONCLUSION: The results indicate that this model of human oral mucosa, constructed using fibrin-agarose scaffolds, shows similarities to native oral mucosa controls and imply that bioengineered oral mucosa substitutes could eventually be used clinically.

Molecular and functional characterization of eight novel GAA mutations in Italian infants with Pompe disease.

We characterized 29 unrelated patients presenting with the severe form of Pompe disease (Glycogen Storage Disease Type II, acid maltase deficiency) and identified 26 pathogenic mutations divided over 28 different genotypes. Among the eight new mutations, five were exonic point mutations (c.572A>G, c.1124G>T, c.1202A>G, c.1564C>G and c.1796C>A) leading to codon changes (p.Y191C, p.R375L, p.Q401R, p.P522A and p.S599Y); two were intronic point mutations (c.-32-3C>A and c.1636+5G>C) affecting mRNA processing; one was a single base deletion (c.742delC) generating a truncated protein (p.L248PfsX20). A comprehensive evaluation, based on different methodological approaches, confirmed the detrimental effect of the eight mutations on the protein and its function. Structural alterations potentially induced by the five missense mutations were also predicted through visual inspection of the atomic model of the GAA protein, in terms of both function and spatial orientation of specific residues as well as disturbance generated by amino acid substitutions. Although the remarkable heterogeneity of the mutational spectrum in Pompe disease was already known, our data demonstrate and confirm the power of molecular and functional analysis in predicting the natural course of Pompe disease.

Funtional characterization of four novel MAN2B1 mutations causing juvenile onset alpha-mannosidosis.

Alpha-mannosidosis is a recessively inherited disorder due to the deficiency of the lysosomal alpha-mannosidase. We report the molecular analysis performed in two patients with the late onset form of alpha-mannosidosis. Four new alleles were identified: three missense mutations involving highly conserved residues, c.597 C>A (p.H200N), c.1553 T>C (p.L518P) and c.2746 C>A (p.R916S) and a single nucleotide deletion, c.2660delC. In vitro expression studies in COS-1 cells demonstrated that pH200N, p.L518P and p.R916S proteins are expressed but retained no residual enzyme activity. These data are supported by structural 3D analysis which predicted that both p.L518P and p.R916S could affect the interaction of the small E-domain with the active site domain or the main body of the structure while the pH200N might alter substrate binding or other catalytic properties. Finally, the c.2660delC causes a frameshift introducing a premature stop codon (p.T887SfsX45), presuming to be a severe mutation.

Mutation profile of the GAA gene in 40 Italian patients with late onset glycogen storage disease type II.

Glycogen storage disease type II (GSDII) is a recessively inherited disorder due to the deficiency of acid alpha-glucosidase (GAA) that results in impaired glycogen degradation and its accumulation in the lysosomes. We report here the complete molecular analysis of the GAA gene performed on 40 Italian patients with late onset GSDII. Twelve novel alleles have been identified: missense mutations were functionally characterized by enzyme activity and protein processing in a human GAA-deficient cell line while splicing mutations were studied by RT-PCR and in silico analysis. A complex allele was also identified carrying three different alterations in cis. The c.-32-13T > G was the most frequent mutation, present as compound heterozygote in 85% of the patients (allele frequency 42.3%), as described in other late onset GSDII Caucasian populations. Interestingly, the c.-32-13T > G was associated with the c.2237G > A (p.W746X) in nine of the 40 patients. Genotype-phenotype correlations are discussed with particular emphasis on the subgroup carrying the c.-32-13T > G/c.2237G > A genotype.

MicroRNA changes through Müller glia dedifferentiation and early/late rod photoreceptor differentiation.

Cell-type determination is a complex process driven by the combinatorial effect of extrinsic signals and the expression of transcription factors and regulatory genes. MicroRNAs (miRNAs) are non-coding RNAs that, generally, inhibit the expression of target genes and have been involved, among other processes, in cell identity acquisition. To search for candidate miRNAs putatively involved in mice rod photoreceptor and Müller glia (MG) identity, we compared miRNA expression profiles between late-stage retinal progenitor cells (RPCs), CD73-immunopositive (CD73+) rods and postnatal MG. We found a close similarity between RPCs and CD73+ miRNA expression profiles but a divergence between CD73+ and MG miRNA signatures. We validated preferentially expressed miRNAs in the CD73+ subpopulation (miR-182, 183, 124a, 9(∗), 181c and 301b(∗)) or MG (miR-143, 145, 214, 199a-5p, 199b(∗), and 29a). Taking advantage of the unique capacity of MG to dedifferentiate into progenitor-like cells that can be differentiated to a rod phenotype in response to external cues, we evaluated changes of selected miRNAs in MG-derived progenitors (MGDP) during neuronal differentiation. We found decreased levels of miR-143 and 145, but increased levels of miR-29a in MGDP. In MGDPs committed to early neuronal lineages we found increased levels of miR-124a and upregulation of miR-124a, 9(∗) and 181c during MGDP acquisition of rod phenotypes. Furthermore, we demonstrated that ectopic miR-124 expression is sufficient to enhance early neuronal commitment of MGDP. Our data reveal a dynamic regulation of miRNAs in MGDP through early and late neuronal commitment and miRNAs that could be potential targets to exploit the silent neuronal differentiation capacity of MG in mammals.

Perspectiva de mujeres mexicanas sobre lactancia materna asociada al cambio del modelo cultural / Mexican women breastfeeding perspective associated with the cultural model change / Perspectiva das mulheres mexicanas sobre a aleitamento materno associado ao câmbio do modelo cultural

RESUMEN Introducción: La lactancia materna es importante para el desarrollo saludable del recién nacido. A pesar de sus beneficios, la decisión de no amamantar se incrementa entre las mujeres de la Ciudad de México debido a la influencia de la globalización y del cambio de modelo cultural moderno al posmoderno. Objetivo: Comprender la perspectiva de las mujeres mexicanas sobre el ofrecer y abandonar la lactancia materna asociada al cambio del modelo cultural. Metodología: Se llevó a cabo un estudio cualitativo con enfoque exploratorio, descriptivo: participaron 12 diadas (madre-hijo lactante), residentes de la Ciudad de México, quienes ofrecieron y abandonaron la lactancia materna. Se realizaron entrevistas en profundidad. Los datos obtenidos se grabaron, transcribieron y codificaron para garantizar el rigor de la investigación. Resultados: Se reconocieron dos categorías que sustentan el por qué las mujeres brindan o abandonan la lactancia materna, vistas desde el modelo cultural. En el modelo moderno las mujeres consideran una convicción sentida el ofrecer lactancia materna, reconocen los beneficios de dicho alimento para su hijo; mientras que, en el modelo posmoderno, las mujeres tienen una convicción negativa de amamantar. Conclusiones: Al asociar la lactancia materna con modelos culturales, en el moderno se considera la lactancia como un deber, mientras que en el posmoderno un derecho. Al encontrarnos en transición entre ambos modelos, las mujeres están influenciadas por discursos y prácticas culturales que en ocasiones son contradictorias. Comprender estos fenómenos permitirá diseñar estrategias efectivas desde la enfermería para fomentar la lactancia materna.

ABSTRACT Introduction: Breastfeeding is an important activity for the healthy development of the newborn, but nevertheless the benefits, the decision of women to avoid breastfeeding is becoming more prevalent in Mexico City. Perhaps this decision is the result of the influence of the globalization process with its cultural model change from the modern to the postmodern. Objective: To better understand the perspective of Mexican women regarding breastfeeding or not breastfeeding and the relationship of this decision with the cultural model change. Methodology: This is a qualitative, descriptive, and exploratory study on 12 mothers in Mexico City, who decided to terminate breastfeeding their babies. In-Depth interviews were conducted. The gathered data were recorded, transcribed, and coded in order to procure rigour of research. Results: Two categories related to the cultural model that reflect why women breastfeed or not were identified. In the modern model, women consider breastfeeding as a solid conviction which has benefits to their babies, while in the postmodern model, women tend to have a negative image of breastfeeding. Conclusions: From the association of breastfeeding with the cultural models, in the modern model, women consider breastfeeding as a duty, while in the postmodern one, women consider breastfeeding as a right. Currently, we are in a transition between these possibly contradicting cultural models. Therefore, better understanding these phenomena can help nursing professionals design strategies which can foster a healthy decision on breastfeeding.

RESUMO Introdução: O aleitamento materno é importante para o desenvolvimento saudável do recém-nascido. Apesar de seus benefícios, a decisão de não amamentar aumenta entre as mulheres da Cidade do México devido à influência da globalização e da mudança do modelo cultural moderno para o pós-moderno. Objetivo: Compreender a perspectiva das mulheres mexicanas sobre a oferta e o abandono do aleitamento materno associado à mudança do modelo cultural. Metodologia: Realizou-se um estudo qualitativo com abordagem exploratória e descritiva: participaram 12 díades (mãe-filho em lactação), residentes na Cidade do México, que ofereceram e abandonaram a amamentação. Entrevistas em profundidade foram realizadas. Os dados obtidos foram gravados, transcritos e codificados para garantir o rigor da pesquisa. Resultados: Foram reconhecidas duas categorias que sustentam porque as mulheres oferecem ou abandonam a amamentação, visto a partir do modelo cultural. No modelo moderno, as mulheres consideram o fato da amamentação uma convicção sincera, reconhecem os benefícios dessa alimentação para o filho; enquanto no modelo pós-moderno as mulheres têm uma convicção negativa de amamentar. Conclusões: Ao associar o aleitamento materno aos modelos culturais, no moderno amamentar é considerado um dever, enquanto na pós-moderna um direito. Ao encontrarmos em transição entre os dois modelos, as mulheres são influenciadas por discursos e práticas culturais que às vezes são contraditórios. Compreender esses fenômenos permitirá o desenho de estratégias de enfermagem eficazes para promover o aleitamento materno.

Synthesis and turnover rates of four rat liver mitochondrial RNA species.

The synthesis and turnover rates of the two 12 S and 16 S mt rRNAs and of the mt mRNAs for subunits I and III of cytochrome oxidase have been determined by measuring the kinetics of incorporation of [3H]uridine in the mtRNA of rat hepatocytes. All the RNA species examined have approximately the same turnover (t1/2 approximately 100 min) and therefore the rate of synthesis, which is about 10-times higher for the rRNAs, seems to be the factor responsible for the different mt rRNA and mRNA steady-state levels.

Toward a general-purpose analog VLSI neural network with on-chip learning.

This paper describes elements necessary for a general-purpose low-cost very large scale integration (VLSI) neural network. By choosing a learning algorithm that is tolerant of analog nonidealities, the promise of high-density analog VLSI is realized. A 64-synapse, 8-neuron proof-of-concept chip is described. The synapse, which occupies only 4900 mum(2) in a 2-mum technology, includes a hybrid of nonvolatile and dynamic weight storage that provides fast and accurate learning as well as reliable long-term storage with no refreshing. The architecture is user-configurable in any one-hidden-layer topology. The user-interface is fully microprocessor compatible. Learning is accomplished with minimal external support; the user need only present inputs, targets, and a clock. Learning is fast and reliable. The chip solves four-bit parity in an average of 680 ms and is successful in about 96% of the trials.

Reaction of Ten Cultivars of Watermelon (Citrullus lanatus) to a Puerto Rican population of Meloidogyne incognita.

Ten cultivars of watermelon were evaluated for their response to a Puerto Rican population of Meloidogyne incognita under greenhouse conditions in a 2-year study (1989 and 1990). Ten-day-old seedlings were planted in steam-sterilized soil in 15-cm-d plastic pots. The nematode inoculum consisted of 10,000 eggs and (or) second-stage juveniles (J2)/plant. The cultivars were Sugar Baby, Charleston Gray, Seedless, Prince Charles, Charleston 76, Jubilee, Florida Giant, Royal Charleston, Royal Sweet, and Royal Jubilee, with tomato cv. Rutgers included as a susceptible check. A completely randomized design with 10 replications was used. Fifty-five days after soil infestation, root-gall indices, numbers of nematode eggs per root system, and J2 per 250 cm(3) of soil were recorded. All cultivars were susceptible. Sugar Baby had the lowest root-gall index, egg and J2 numbers, and a reproductive factor (Rf) of 2.89. Rf differed (P

Teachers' perceptions of occupational stress factors.

Perceptions of factors in occupational stress were examined using the 17 elementary and 25 high school teachers' gender, age, experience, and grade taught. Statistically significant differences were reported.

Glutamate-induced epigenetic and morphological changes allow rat Müller cell dedifferentiation but not further acquisition of a photoreceptor phenotype.

Müller cells are not only the main glial cell type in the retina but also latent progenitor/stem cells, which in pathological conditions can transdifferentiate to a neuronal phenotype and regenerate the neurons lost in a mature retina. Several signal transduction pathways can induce the dedifferentiation of mature Müller cells to a progenitor-like state, including that stimulated by glutamate. However, the precise molecular mechanisms by which terminally differentiated cells are initially primed to acquire multipotency remain unclear. In the present study, we have characterized early genetic and epigenetic events that occur immediately after glutamate-induced dedifferentiation of fully differentiated Müller cells is initiated. Using Müller cell-enriched cultures from postnatal rats, we demonstrate that glutamate triggers a rapid dedifferentiation response characterized by changes in cell morphology coupled to the induction of progenitor cell marker gene expression (e.g., nestin, lin28 and sox2) within 1h. Dedifferentiation involved the activation of N-methyl-d-aspartate and type II metabotropic glutamate receptors, as well as global DNA demethylation (evident through the decrease in methyl-CpG-binding protein 2 immunoreactivity) and an increase in gadd45-ß gene expression; although, early progenitor gene expression was only partially inhibited by pharmacological impairment of DNA methylation. Importantly, the expression of Müller glia identity genes (i.e., glutamine synthetase; cellular retinaldehyde binding protein, CRALBP) is retained through the process. Dedifferentiated Müller cells held an early neuronal differentiation potential similar to that observed in retinal progenitor-enriched cultures but, contrary to the latter, dedifferentiated Müller cells failed to further differentiate into mature photoreceptor lineages. We speculate that, in spite of the initial triggering of the dedifferentiation pathways, these cells may exhibit a certain degree of epigenetic memory that precludes them from further differentiation.