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Root hairs (RH) are excellent model systems for studying cell size and polarity since they elongate several hundred-fold their original size. Their tip growth is determined both by intrinsic and environmental signals. Although nutrient availability and temperature are key factors for a sustained plant growth, the molecular mechanisms underlying their sensing and downstream signaling pathways remain unclear. We use genetics to address the roles of the cell surface receptor kinase FERONIA (FER) and the nutrient sensing TOR Complex 1 (TORC) in RH growth. We identified that low temperature (10°C) triggers a strong RH elongation response in Arabidopsis thaliana involving FER and TORC. We found that FER is required to perceive limited nutrient availability caused by low temperature. FERONIA interacts with and activates TORC-downstream components to trigger RH growth. In addition, the small GTPase Rho of plants 2 (ROP2) is also involved in this RH growth response linking FER and TOR. We also found that limited nitrogen nutrient availability can mimic the RH growth response at 10°C in a NRT1.1-dependent manner. These results uncover a molecular mechanism by which a central hub composed by FER-ROP2-TORC is involved in the control of RH elongation under low temperature and nitrogen deficiency.
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Proteínas de Arabidopsis , Arabidopsis , Arabidopsis/metabolismo , Nitratos/farmacologia , Nitratos/metabolismo , Proteínas de Arabidopsis/metabolismo , Temperatura , Fosfotransferases/metabolismo , Nitrogênio/metabolismo , Raízes de Plantas/metabolismo , Proteínas de Plantas/metabolismo , Proteínas de Transporte de Ânions/metabolismoRESUMO
Three series of polyesters based on monomer combinations of ε-caprolactone (ε-CL), ethylene brassylate (EB), and l-Lactide (LLA) with the alkyl substituted lactone ε-decalactone (ε-DL) were synthesized at different molar ratios. Copolymers were obtained via ring opening polymerization (ROP) employing TBD (1,5,7-triazabicyclo-[4.4.0]-dec-5-ene), an organic catalyst which can be handled under normal conditions, avoiding the use of glove box equipment. The molar monomer composition of resulting copolymers differed from theoretical values due to lower ε-DL reactivity; their Mn and Mw values were up to 14 kDa and 22.8 kDa, respectively, and distributions were (Æ) ≤ 2.57. The thermal stability of these materials suffered due to variations in their ε-DL molar content. Thermal transitions such as melting (Tm) and crystallization (Tc) showed a decreasing tendency as ε-DL molar content increased, while glass transition (Tg) exhibited minor changes. It is worth mentioning that changes in monomer composition in these polyesters have a strong impact on their thermal performance, as well as in their crystallization degree. Consequently, variations in their chemical structure may have an effect on hydrolyic degradation rates. It should be noted that, in future research, some of these copolymers will be exposed to hydrolytic degradation experiments, including characterizations of their mechanical properties, to determine their adequacy in potential use in the development of soft medical devices.
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INTRODUCTION: Retinopathy of prematurity (ROP) is a condition that affects preterm infants, being the second leading cause of childhood blindness worldwide. The most commonly used treatments are cryotherapy or laser photocoagulation, but over the last few years, anti-vascular endothelial growth factor (anti-VEGF) drugs have gradually gained more adherents, mainly in the treatment of patients with type 1 retinopathy of prematurity (ET- ROP) Therefore, it is important to summarize the existing evidence to evaluate the efficacy and safety of anti-VEGF drugs in type 1 retinopathy of prematurity. METHODS: We searched in Epistemonikos, the largest database of systematic reviews in health, which is maintained by screening multiple information sources, including MEDLINE, EMBASE, Cochrane, among others. We extracted data from the systematic reviews, reanalyzed data of primary studies, conducted a meta-analysis and generated a summary of findings table using the GRADE approach. RESULTS AND CONCLUSIONS: We identified six systematic reviews including 15 studies overall, of which five were randomized trials. We conclude that the use of anti-VEGF compared to laser photocoagulation, probably reduces the risk of refractive errors. On the other hand, the use of anti-VEGF may result in little or no difference in the mortality at hospital discharge, lens or corneal opacity requiring surgery, and complete or partial retinal detachment, but the certainty of the evidence is low. Finally, it is not possible to clearly establish whether anti-VEGF compared with laser photocoagulation, increases the recurrence of retinopathy of prematurity, because the certainty of the existing evidence has been assessed as very low.
INTRODUCCIÓN: La retinopatía del prematuro es una patología que afecta a recién nacidos de pretérmino, siendo la segunda causa de ceguera infantil a nivel mundial. Los tratamientos más utilizados son la crioterapia o la fotocoagulación láser, pero en el último tiempo, los fármacos anti factor del crecimiento vascular endotelial (anti-VEGF) gradualmente han ganado más adeptos, principalmente en el tratamiento de pacientes con retinopatía del prematuro tipo 1 (ET-ROP). Por lo anterior, es importante realizar un resumen de la evidencia existente para evaluar la eficacia y seguridad de los fármacos anti-VEGF en retinopatía del prematuro tipo 1. MÉTODOS: Para responder esta pregunta utilizamos Epistemonikos, la mayor base de datos de revisiones sistemáticas en salud, la cual es mantenida mediante búsquedas en múltiples fuentes de información, incluyendo MEDLINE, EMBASE, Cochrane, entre otras. Extrajimos los datos desde las revisiones identificadas, analizamos los datos de los estudios primarios, realizamos un metaanálisis y preparamos una tabla de resumen de los resultados utilizando el método GRADE. RESULTADOS Y CONCLUSIONES: Identificamos seis revisiones sistemáticas que en conjunto incluyeron 15 estudios primarios, de los cuales cinco corresponden a ensayos aleatorizados. Concluimos que el uso de anti-VEGF en comparación con la fotocoagulación con láser, probablemente disminuye el riesgo de errores refractivos. Por otro lado, el uso de anti-VEGF podría resultar en poca o nula diferencia en la mortalidad al alta hospitalaria, opacidad de cristalino o córnea que requiera cirugía y desprendimiento completo o parcial de retina, pero la certeza de la evidencia es baja. Finalmente, no es posible establecer con claridad si los anti-VEGF comparados con la fotocoagulación láser aumenta la recurrencia de la retinopatía del prematuro, debido a que la certeza de la evidencia existente ha sido evaluada como muy baja.
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Preparações Farmacêuticas , Retinopatia da Prematuridade , Inibidores da Angiogênese/uso terapêutico , Humanos , Lactente , Recém-Nascido , Recém-Nascido Prematuro , Injeções Intravítreas , Lasers , Fotocoagulação , Revisões Sistemáticas como Assunto , Fator A de Crescimento do Endotélio VascularRESUMO
Legumes form root mutualistic symbioses with some soil microbes promoting their growth, rhizobia, and arbuscular mycorrhizal fungi (AMF). A conserved set of plant proteins rules the transduction of symbiotic signals from rhizobia and AMF in a so-called common symbiotic signaling pathway (CSSP). Despite considerable efforts and advances over the past 20 years, there are still key elements to be discovered about the establishment of these root symbioses. Rhizobia and AMF root colonization are possible after a deep cell reorganization. In the interaction between the model legume Lotus japonicus and Mesorhizobium loti, this reorganization has been shown to be dependent on a SCAR/Wave-like signaling module, including Rho-GTPase (ROP in plants). Here, we studied the potential role of ROP3 in the nitrogen-fixing symbiosis (NFS) as well as in the arbuscular mycorrhizal symbiosis (AMS). We performed a detailed phenotypic study on the effects of the loss of a single ROP on the establishment of both root symbioses. Moreover, we evaluated the expression of key genes related to CSSP and to the rhizobial-specific pathway. Under our experimental conditions, rop3 mutant showed less nodule formation at 7- and 21-days post inoculation as well as less microcolonies and a higher frequency of epidermal infection threads. However, AMF root colonization was not affected. These results suggest a role of ROP3 as a positive regulator of infection thread formation and nodulation in L. japonicus. In addition, CSSP gene expression was neither affected in NFS nor in AMS condition in rop3 mutant. whereas the expression level of some genes belonging to the rhizobial-specific pathway, like RACK1, decreased in the NFS. In conclusion, ROP3 appears to be involved in the NFS, but is neither required for intra-radical growth of AMF nor arbuscule formation.
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The allelic combination of ROP18/ROP5 genes of Toxoplasma gondii has been shown to be highly predictive of mouse virulence in canonical isolates and strains. The aims of this study were to analyze the alleles present in the ROP18/ROP5 genes from T. gondii isolates obtained in Argentina, to associate the results with the virulence registered in mouse model, and to compare with other isolates and reference strains using a phylogenetic network. Fourteen T. gondii isolates from Argentina were analyzed by nPCR-RFLP for ROP18/ROP5. Phylogenetic network analysis was inferred using the ToxoDB genotypes and the ROPs molecular markers. All isolates and reference strains were categorized as lethal or non-lethal. As results, combinations 2/2, 3/3 and 4/3 for ROP18/ROP5 were detected in 12 isolates, whereas only alleles 1 and 2 of ROP5 were detected in 2 isolates. The majority of the isolates had a mouse virulence matching to that predicted by the ROP18/ROP5 allele combination. The 3 isolates that differed from the expected virulence presented non-clonal genotypes. ROPs incorporation increased the accuracy of the phylogenetic network relations among the T. gondii samples, prevailing the clustering according to regions. Our results indicate a predominance of type 3 allele in both ROP18 and ROP5 markers and an association of allelic profiles 3/3 and 4/3 of non-clonal genotypes from Argentina, both with virulent and avirulent profiles in mice.
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Proteínas Serina-Treonina Quinases/genética , Toxoplasma/genética , Toxoplasma/patogenicidade , Toxoplasmose Animal/parasitologia , Toxoplasmose/parasitologia , Alelos , Animais , Modelos Animais de Doenças , Camundongos , Camundongos Endogâmicos ICR , Filogenia , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas de Protozoários , Virulência/genéticaRESUMO
Well-defined amphiphilic, biocompatible and partially biodegradable, thermo-responsive poly(N-vinylcaprolactam)-b-poly(ε-caprolactone) (PNVCL-b-PCL) block copolymers were synthesized by combining reversible addition-fragmentation chain transfer (RAFT) and ring-opening polymerizations (ROP). Poly(N-vinylcaprolactam) containing xanthate and hydroxyl end groups (X-PNVCL-OH) was first synthesized by RAFT/macromolecular design by the interchange of xanthates (RAFT/MADIX) polymerization of NVCL mediated by a chain transfer agent containing a hydroxyl function. The xanthate-end group was then removed from PNVCL by a radical-induced process. Finally, the hydroxyl end-capped PNVCL homopolymer was used as a macroinitiator in the ROP of ε-caprolactone (ε-CL) to obtain PNVCL-b-PCL block copolymers. These (co)polymers were characterized by Size Exclusion Chromatography (SEC), Fourier-Transform Infrared spectroscopy (FTIR), Proton Nuclear Magnetic Resonance spectroscopy (1H NMR), UV-vis and Differential Scanning Calorimetry (DSC) measurements. The critical micelle concentration (CMC) of the block copolymers in aqueous solution measured by the fluorescence probe technique decreased with increasing the length of the hydrophobic block. However, dynamic light scattering (DLS) demonstrated that the size of the micelles increased with increasing the proportion of hydrophobic segments. The morphology observed by cryo-TEM demonstrated that the micelles have a pointed-oval-shape. UV-vis and DLS analyses showed that these block copolymers have a temperature-responsive behavior with a lower critical solution temperature (LCST) that could be tuned by varying the block copolymer composition.
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The high prevalence of Toxoplasma gondii in the human population in Colombia has been linked to the existence of a high density of urban stray cats, exposing the whole population to a high density of oocysts. The goal of this study was to determine the DNA prevalence of T. gondii by conventional PCR and to phylogenetically analyze ROP18 sequences from positive samples in domestic cat (Felis catus) fecal samples in the city of Armenia, Quindío. Fecal samples from 140 cats were collected from 10 districts around the city. Samples were concentrated using Ritchie's method and analyzed through optical microscopy. Concentrates were used for DNA extraction followed by nested PCR amplification for T. gondii gene B1. PCR for ROP18 was performed on all B1 positive samples; the ROP18 sequences obtained were related to the Archetype I Brazilian and Chinese strains. No oocysts were detected by optical microscopy; however, 17.8% (25/140) B1 and 24% (6/25) ROP18 PCR-positive samples were detected. Phylogenetic analyses showed that isolates clustered into a single group. We assessed whether associations existed between T. gondii positive fecal samples and survey variables such as cat healthcare and socioeconomic characteristics of owners, but no statistically significant associations were found. The presence of T. gondii in cat feces is an important factor contributing to the high prevalence in the human population of this city.
TITLE: Détection d'ADN et génotypes de Toxoplasma gondii dans les fèces de chats domestiques en Colombie. ABSTRACT: La forte prévalence de Toxoplasma gondii dans la population humaine en Colombie a été liée à l'existence d'une forte densité de chats errants urbains, exposant l'ensemble de la population à une forte densité d'oocystes. Le but de ce travail était de déterminer la prévalence de l'ADN de T. gondii par PCR conventionnelle et d'analyser phylogénétiquement les séquences ROP18 d'échantillons positifs dans des échantillons fécaux de chat domestique (Felis catus) dans la ville d'Armenia, Quindío. Des échantillons fécaux de 140 chats ont été collectés dans 10 districts de la ville. Les échantillons ont été concentrés en utilisant la méthode de Ritchie et analysés par microscopie optique. Des concentrés ont été utilisés pour l'extraction d'ADN suivie d'une amplification par PCR nichée pour le gène B1 de T. gondii. La PCR pour ROP18 a été réalisée sur tous les échantillons positifs pour B1 ; les séquences ROP18 obtenues étaient apparentées aux souches Archétype I brésiliennes et chinoises. Aucun oocyste n'a été détecté par microscopie optique mais les échantillons étaient positifs par PCR pour 17,8 % (25/140) pour B1 et 24 % (6/25) pour ROP18. Les analyses phylogénétiques ont montré que les isolats formaient un seul groupe. Nous avons évalué s'il existait des associations entre des échantillons fécaux positifs à T. gondii et des variables d'enquête telles que les soins de santé des chats et les caractéristiques socioéconomiques des propriétaires, mais aucune association statistiquement significative n'a été trouvée. La présence de T. gondii dans les excréments de chats est un facteur important contribuant à la forte prévalence dans la population humaine de cette ville.
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Doenças do Gato/epidemiologia , Gatos/parasitologia , DNA de Protozoário/genética , Toxoplasma/genética , Toxoplasmose Animal/epidemiologia , Animais , Doenças do Gato/parasitologia , Colômbia/epidemiologia , DNA de Protozoário/isolamento & purificação , Fezes/parasitologia , Feminino , Genótipo , Masculino , Oocistos/genética , Oocistos/fisiologia , Animais de Estimação/parasitologia , Filogenia , Prevalência , Proteínas Serina-Treonina Quinases/genética , Proteínas de Protozoários , Toxoplasma/isolamento & purificaçãoRESUMO
PROPOSE: To report two cases of severe acute multi-systemic failure with bilateral ocular toxoplasmosis in immunocompetent patients from urban settings in Colombia. OBSERVATIONS: We report two immunocompetent male patients aged 44- and 67-years-old who, despite not having visited the Amazonian region in Colombia, had severe bilateral posterior uveitis and extensive-bilateral macular lesions and multiple organ failure that required admission to an intensive care unit. Toxoplasma gondii was positive by PCR assay in vitreous humor samples. Patients were treated with intravitreal clindamycin and dexamethasone in addition to systemic treatment with trimethoprim-sulfamethoxazole. In both patients, infection by atypical strains was confirmed; in one case by serotyping and in another one by genotyping (ROP 18 virulent allele). After 2 and 4 months of treatment respectively, the patients showed improvement of the posterior uveitis and its systemic manifestations. However, there was no significant visual acuity improvement due to bilateral extensive macular involvement. CONCLUSIONS AND IMPORTANCE: Clinicians should be aware that toxoplasmosis originating from South America could be associated with severe acute multisystemic and intraocular bilateral involvement, even in patients with no history of exposure to jungle environments.
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Toxoplasma gondii engenders the common parasitic disease toxoplasmosis in almost all warm-blooded animals. Being a critical secretory protein, ROP18 is a major virulence factor of Toxoplasma. There are no reports about ROP18 detection in human serum samples with different clinical manifestations. New aptamers against ROP18 protein were developed through Systematic Evolution of Ligands by Exponential enrichment (SELEX). An Enzyme-Linked Aptamer Assay (ELAA) platform was developed using SELEX-derived aptamers, namely AP001 and AP002. The ELAA was used to evaluate total antigen from T. gondii RH strain (RH Ag) and recombinant protein of ROP18 (rROP18). The results showed that the ELAA presented higher affinity and specificity to RH Ag and rROP18, compared to negative controls. Detection limit of rROP18 protein in serum samples was measured by standard addition method, achieving a lower concentration of 1.56 µg/mL. Moreover, 62 seropositive samples with different clinical manifestations of toxoplasmosis and 20 seronegative samples were tested. A significant association between ELAA test positive for human serum samples and severe congenital toxoplasmosis was found (p = 0.006). Development and testing of aptamers-based assays opens a window for low-cost and rapid tests looking for biomarkers and improves our understanding about the role of ROP18 protein on the pathogenesis of human toxoplasmosis.
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Ensaio de Imunoadsorção Enzimática , Proteínas Serina-Treonina Quinases/imunologia , Técnica de Seleção de Aptâmeros , Toxoplasma/imunologia , Toxoplasmose/diagnóstico , Toxoplasmose/parasitologia , Aptâmeros de Peptídeos , Ensaio de Imunoadsorção Enzimática/métodos , Ensaio de Imunoadsorção Enzimática/normas , Humanos , Proteínas Serina-Treonina Quinases/sangue , Proteínas de Protozoários , Proteínas Recombinantes , Sensibilidade e Especificidade , Toxoplasmose/imunologiaRESUMO
Toxoplasma gondii ROP16 and ROP18 proteins have been identified as important virulence factors for this parasite. Here, we describe the effect of ROP16 and ROP18 proteins on peripheral blood mononuclear cells (PBMCs) from individuals with different clinical status of infection. We evaluated IFN-γ, IL-10, and IL-1ß levels in supernatants from PBMCs cultures infected with tachyzoites of the T. gondii wild-type RH strain or with knock-out mutants of the rop16 and rop18 encoding genes (RHΔrop16 and RHΔrop18). Cytokine secretion was compared between PBMCs obtained from seronegative individuals (n = 10), with those with chronic asymptomatic (n = 8), or ocular infection (n = 12). We also evaluated if polymorphisms in the genes encoding for IFN-γ, IL-10, IL-1ß, Toll-like receptor 9 (TLR9), and purinoreceptor P2RX7 influenced the production of the encoded proteins after ex vivo stimulation. In individuals with chronic asymptomatic infection, only a moderate effect on IL-10 levels was observed when PBMCs were infected with RHΔrop16, whereas a significant difference in the levels of inflammatory cytokines IFN-γ and IL-1ß was observed in seronegative individuals, but this was also dependent on the host's cytokine gene polymorphisms. Infection with ROP16-deficient parasites had a significant effect on IFN-γ production in previously non-infected individuals, suggesting that ROP16 which is considered as a virulence factor plays a role during the primary infection in humans, but not in the secondary immune response.
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Leucócitos Mononucleares/imunologia , Proteínas Serina-Treonina Quinases/imunologia , Proteínas Tirosina Quinases/imunologia , Proteínas de Protozoários/imunologia , Toxoplasma/imunologia , Toxoplasmose/imunologia , Toxoplasmose/parasitologia , Citocinas/metabolismo , Fibroblastos , Interações Hospedeiro-Parasita/genética , Interações Hospedeiro-Parasita/imunologia , Humanos , Leucócitos Mononucleares/metabolismo , Polimorfismo Genético , Fator de Transcrição STAT3/metabolismo , Fator de Transcrição STAT6/metabolismo , Toxoplasma/patogenicidade , Toxoplasmose/genética , Virulência , Fatores de Virulência/imunologiaRESUMO
BACKGROUND: Retinopathy of prematurity (ROP), the primary cause of blindness in children, is a potential complication for 7.7% of live births in Mexico. Given that less than one-third of all neonatal intensive care units follow Mexican National ROP guidelines, there have been few reports regarding the incidences of types 1 and 2 ROP. METHODS: This was a retrospective study that investigated the incidence and onset of ROP in a representative sample of children in Mexico. We analyzed the results obtained by the ROP Detection and Treatment Program, compliant with the Mexican National ROP guidelines, over a 1-year period. This study included 132 children who were born prematurely, were initially screened between October 2, 2017 and October 1, 2018, and underwent follow-up based on their risk group (in accordance with the Mexican National ROP guidelines). RESULTS: The mean gestational age (GA) at birth was 32 weeks and 3 days (32w3d) (95% CI, ± 3 days), and the mean birth weight (BW) was 1594 g (95% CI, ± 96 g). The clinical features were as follows: 36.4% had immature retina without ROP, 22.0% had mild ROP, 5.3% had type 2 ROP, 27.3% had type 1 ROP, and 1.5% had advanced disease. Premature children with ROP requiring treatment (i.e., type 1 ROP + advanced ROP) were born at an MGA of 30w4d (95% CI, ± 5d; range, 26-35 weeks); their MBW was 1316 g (95% CI, ± 110 g; range, 830-2220 g). Diagnosis of ROP requiring treatment was made at a mean postmenstrual age (PMA) of 37w3d (95% CI, ± 5d; range, 31w1d to 42w1d). CONCLUSION: In Mexico, screening and close ophthalmological follow-up of children who present with risk factors of birth weight < 1750 g and gestational age ≤ 34 weeks, both of which are observed more frequently in children with type 1 ROP, appears essential for implementing timely treatments (within 72 h). This is particularly important for children with PMA between 36 and 38 weeks, which is considered to be the peak age for disease stages that require timely intervention.
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Triagem Neonatal/métodos , Retinopatia da Prematuridade/epidemiologia , Feminino , Seguimentos , Idade Gestacional , Humanos , Incidência , Recém-Nascido , Masculino , México/epidemiologia , Estudos Retrospectivos , Fatores de RiscoRESUMO
The superfamily of small monomeric GTPases originated in a common ancestor of eukaryotic multicellular organisms and, since then, it has evolved independently in each lineage to cope with the environmental challenges imposed by their different life styles. Members of the small GTPase family function in the control of vesicle trafficking, cytoskeleton rearrangements and signaling during crucial biological processes, such as cell growth and responses to environmental cues. In this review, we discuss the emerging roles of these small GTPases in the pathogenic and symbiotic interactions established by plants with microorganisms present in their nearest environment, in which membrane trafficking is crucial along the different steps of the interaction, from recognition and signal transduction to nutrient exchange.
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Membrana Celular/enzimologia , GTP Fosfo-Hidrolases/metabolismo , Proteínas de Plantas/metabolismo , Plantas/enzimologia , Transdução de Sinais/fisiologia , Transporte Biológico Ativo/fisiologiaRESUMO
Recent studies carried out in Brazil have shown that strains from the same Toxoplasma gondii genotype can infect humans, domestic animals (dogs and cats) and animals slaughtered for human consumption (pigs, sheep, goats, and chickens), suggesting a common infection route. However, little is known about the importance of free-living wild birds within this epidemiological context. The objective of this work was to isolate, genotype, and evaluate the virulence for mice of new isolates of T. gondii obtained from free-living wild birds from the state of Minas Gerais, Southeastern Brazil. From August 2016 to June 2017, T. gondii was isolated from the hearts and brains collected from 6 out of 45 free-living wild birds, namely, a roadside hawk (Rupornis magnirostris), a campo flicker (Colaptes campestris), a southern caracara (Caracara plancus) and a tropical screech-owl (Megascops choliba), all rescued in Belo Horizonte. One isolate was obtained from a toco toucan (Ramphastos toco), rescued in Cristiano Otoni, and another was obtained from southern caracara, rescued in Santa Luzia. Five different genotypes were identified by PCR-RFLP. A unique genotype was shared in two different isolates obtained from a southern caracara and a toco toucan. This genotype has never been previously described in any other host or place. Three isolates were classified as of intermediary virulence and three isolates as avirulent for mice. The combined analysis of alleles ROP18/ROP5 (a serine/threonine kinase, and a polymorphic pseudokinase, respectively) was effective in determining the virulence of five of all the isolates with the exception of that from R. magnirostris. Atypical isolates of T. gondii obtained from free-living wild birds rescued in the state of Minas Gerais share the same genotypes of strains that infect humans, domestic animals, and animals slaughtered for human consumption.
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Toxoplasma gondii (T. gondii) is an obligate intracellular protozoan parasite that can infect almost all warm-blooded species and induce a chronic infection in human hosts. The aim of this work was to investigate Th1, Th2, Th17 and Treg polarization, induced by four important T. gondii antigens (SAG1, ROP1, GRA8 and MAG1) in acutely and chronically infected patients. For this purpose, SAG1, ROP1, GRA8 and MAG1 were expressed as recombinant proteins, purified, and used to evaluate the proinflammatory and regulatory immune response profiles in seropositive and seronegative individuals. Our results show that SAG1 and ROP1 elicited a proinflammatory profile (INF-γ, IL-12 and IL-17) in individuals in the acute phase, whereas MAG1 and GRA8 induced a regulatory pattern (Treg and TGF-ß) in chronically infected patients. These results reveal fundamental differences in T-cell polarization induced by T. gondii antigens, which could have important implications in the immunopathogenesis of the disease and in future proposals of therapeutic strategies.
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Anticorpos Antiprotozoários/imunologia , Antígenos de Protozoários/imunologia , Linfócitos T Reguladores/imunologia , Células Th1/imunologia , Células Th17/imunologia , Células Th2/imunologia , Toxoplasma/imunologia , Toxoplasmose/imunologia , Adulto , Animais , Feminino , Humanos , Interferon gama/biossíntese , Subunidade p35 da Interleucina-12/biossíntese , Interleucina-17/biossíntese , Masculino , Proteínas de Membrana/imunologia , Camundongos , Proteínas de Protozoários/imunologia , Toxoplasmose/parasitologia , Fator de Crescimento Transformador beta1/biossínteseRESUMO
RESUMEN Objetivo: describir los desenlaces luego de la aplicación de terapia antiangiogénica intravítrea con Ranibizumab en pacientes con retinopatía de la prematuridad (ROP), tratados inicialmente con termoterapia transpupilar láser sin regresión de la enfermedad. Métodos: cohorte histórica en Clínica Universitaria Bolivariana (CUB) durante 2008-2015. Evaluación de pacientes prematuros con retinopatía agresiva posterior y enfermedad umbral que recibieron termoterapia transpupilar láser como tratamiento inicial. Debido a que continuaba el proceso de neovascularización, se indicó terapia antiangiogénica intravítrea con ranibizumb. El desenlace primario fue la resolución de la neovascularización (NV) y la ausencia de recurrencia. Los desenlaces secundarios fueron agudeza visual y complicaciones oculares y sistémicas. Resultados: ocho ojos de 4 pacientes fueron incluidos. Se encontró resolución de la neovascularización en 6 de los 8 ojos, densidad de incidencia 48 x 100 personas año, IC 95 % = 17,6 - 100 con una mediana de seguimiento de 23.5 meses, rango 13-90 meses. No se encontraron complicaciones oculares ni sistémicas. Conclusión: esta pequeña cohorte sugiere que la inyección intravitrea de ranibizumab para ROP, en casos que inicialmente se trataron con termoterapia transpupilar con láser y no se resolvió la retinopatía, ayudaría como complemento para una evolución favorable.
SUMMARY Purpose: To describe ocular outcomes after intravitreal ranibizumab injections in patients with retinopathy of prematurity (ROP), who had transpupillary laser thermotherapy without regression of the disease. Methods: Historical cohort in Clínica Universitaria Bolivariana (CUB) during 2008-2015. Evaluation of patients with aggressive posterior ROP, threshold disease, who received intravitreal ranibizumab secondary to treatment with transpupillary thermotherapy. The primary outcome was treatment success defined as the resolution of neovascularization (NV) and no recurrence. The secondary endpoints were visual acuity and ocular and systemic complications. Results: Eight eyes of four patients were included in the study. Six eyes had resolution of retinopathy, Incidence density 48 x 100 person years, 95 % CI = 17.6 - 100 with a median follow-up of 23.5 months, range 13-90 months. No ocular or systemic complications associated with the procedure was found. Conclusion: This cohort suggests that intravitreal injection of ranibizumab for ROP in refractory laser photocoagulation cases, results in apparent ocular preservation a long-term the outcome.
Assuntos
Humanos , Lactente , Retinopatia da Prematuridade , Inibidores da Angiogênese , Ranibizumab , Anormalidades do OlhoRESUMO
BACKGROUND: Small monomeric GTPases act as molecular switches in several processes that involve polar cell growth, participating mainly in vesicle trafficking and cytoskeleton rearrangements. This gene superfamily has largely expanded in plants through evolution as compared with other Kingdoms, leading to the suggestion that members of each subfamily might have acquired new functions associated to plant-specific processes. Legume plants engage in a nitrogen-fixing symbiotic interaction with rhizobia in a process that involves polar growth processes associated with the infection throughout the root hair. To get insight into the evolution of small GTPases associated with this process, we use a comparative genomic approach to establish differences in the Ras GTPase superfamily between legume and non-legume plants. RESULTS: Phylogenetic analyses did not show clear differences in the organization of the different subfamilies of small GTPases between plants that engage or not in nodule symbiosis. Protein alignments revealed a strong conservation at the sequence level of small GTPases previously linked to nodulation by functional genetics. Interestingly, one Rab and three Rop proteins showed conserved amino acid substitutions in legumes, but these changes do not alter the predicted conformational structure of these proteins. Although the steady-state levels of most small GTPases do not change in response to rhizobia, we identified a subset of Rab, Rop and Arf genes whose transcript levels are modulated during the symbiotic interaction, including their spatial distribution along the indeterminate nodule. CONCLUSIONS: This study provides a comprehensive study of the small GTPase superfamily in several plant species. The genetic program associated to root nodule symbiosis includes small GTPases to fulfill specific functions during infection and formation of the symbiosomes. These GTPases seems to have been recruited from members that were already present in common ancestors with plants as distant as monocots since we failed to detect asymmetric evolution in any of the subfamily trees. Expression analyses identified a number of legume members that can have undergone neo- or sub-functionalization associated to the spatio-temporal transcriptional control during the onset of the symbiotic interaction.
Assuntos
Fabaceae/metabolismo , Proteínas Monoméricas de Ligação ao GTP/metabolismo , Fabaceae/genética , Genômica , Proteínas Monoméricas de Ligação ao GTP/classificação , Proteínas Monoméricas de Ligação ao GTP/genética , Filogenia , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Simbiose/genética , Simbiose/fisiologiaRESUMO
Recent studies indicate that proteins GRA15, ROP5, ROP16, ROP17, and ROP18 of Toxoplasma gondii are involved in the process of interaction, cellular invasion, and immune response of the host. Among these proteins, alleles of the polymorphic ROP18 and ROP5 seem to be directly associated with T. gondii virulence in mice. The purpose of this work was to isolate and genotype T. gondii from pig, goat, and sheep slaughtered for human consumption in the state of Piauí, Northeastern Brazil and relate the variability of genes that express virulence proteins of the parasite to virulence in mice. T. gondii was isolated from 16 pigs and 9 goats. The parasite was not isolated from sheep samples. Eleven different genotypes were identified using PCR-RFLP. A unique genotype not yet described in any other host and or anywhere else was common to three pig isolates. Eighteen isolates (72%) were characterized as avirulent, four (16%) as intermediate virulence and three (12%) as virulent to mice. The combined analysis of ROP18 and ROP5 in the isolates studied in Piauí, showed four different allele associations: 4/3 (virulent strains), 3/3, 3/1, and 2/3 (avirulent strains). The association 2/3 was not previously described in the literature. Our results indicated that GRA15, ROP16, and ROP17 alleles were not associated with T. gondii virulence in mice. Pigs and goats raised and slaughtered for human consumption in the state of Piauí are infected with isolates of T. gondii presenting different genotypes. We concluded that the virulence protein ROP18, analyzed alone or in combination with ROP5, was effective in determining virulence for mice for the new isolates of T. gondii.
Assuntos
Doenças das Cabras/parasitologia , Proteínas Serina-Treonina Quinases/metabolismo , Doenças dos Suínos/parasitologia , Toxoplasma/genética , Toxoplasmose Animal/parasitologia , Fatores de Virulência/metabolismo , Alelos , Animais , Biomarcadores , Feminino , Genótipo , Técnicas de Genotipagem/veterinária , Cabras , Camundongos , Camundongos Endogâmicos BALB C , Filogenia , Proteínas Serina-Treonina Quinases/genética , Proteínas de Protozoários/genética , Proteínas de Protozoários/metabolismo , Análise de Sequência de DNA/veterinária , Suínos , Toxoplasma/isolamento & purificação , Toxoplasma/metabolismo , Toxoplasma/patogenicidade , Virulência , Fatores de Virulência/genéticaRESUMO
A series of lithium complexes ([Ph2P(o-C6H4-CH2Li·TMEDA)] (1-Li), [PhP(o-C6H4-CH3)(o-C6H4-CH2Li·TMEDA)] (2-Li), [PhP(o-C6H4-CH2Li·TMEDA)2] (2-Li2) and [P(o-C6H4-CH2Li·TMEDA)3] (3-Li3)) was prepared from mono-, di- and tri-benzylphosphines and varying amounts of nBuLi and was characterized extensively by IR and ¹H, 7Li, 13C and 31P NMR spectroscopy. The molecular structures of complexes 1-Li and 2-Li were determined by single-crystal X-ray diffraction studies. The two complexes have monomeric structures in the solid state comprising seesaw lithium atoms. In each case, the ligand exhibits an asymmetric C-C η²-coordination mode and an intramolecular P-Li bond interaction. Theoretical calculations at Density functional theory (DFT) level M06/6111+G(2d,p) show that indeed a P-Li bond is established which can be explained as the P lone pair (sp1.26) being partially delocalized on an available sp² orbital on Li (sp2.04) and additional bonding contribution of the phosphorous atom to Li stems from further delocalization of a σ P-C orbital into the sp² orbital on Li. The observed short contact distances between an aromatic ipso carbon and Li in the crystal structures of 1-Li and 2-Li are explained as due to the interaction of a σ C-Li orbital into the π* orbital of a C-C aromatic bond. Preliminary tests show compounds 1-Li, 2-Li, 2-Li2 and 3-Li3 are active catalysts in the solvent free ring-opening polymerization (ROP) of ε-caprolactone (ε-CL) and rac-lactide (rac-LA). High conversions to polycaprolactones were obtained in short periods of time: 1-6 min at 25 °C. Additionally, all four lithium complexes behave as moderately good initiators for the ROP of rac-LA showing high conversions to polylactides at 140 °C in one hour.
Assuntos
Compostos de Benzil/química , Complexos de Coordenação/química , Lítio/química , Fosfinas/química , Poliésteres/síntese química , Caproatos/química , Catálise , Dioxanos/química , Lactonas/química , Ligantes , Modelos Moleculares , Polimerização , Teoria QuânticaRESUMO
PURPOSE: The aim of the study was to assess the outcomes of severe retinopathy of prematurity (ROP) in zone I or posterior zone II treated with intravitreal ranibizumab (IVR) as monotherapy or combined treatment with laser photocoagulation. METHODS: This is a retrospective study analyzing clinical records of the included patients. Patients were divided into two groups: group 1 included patients who received only IVR treatment; and group 2 was subdivided into group 2A - including patients with IVR as initial treatment and complementary laser photocoagulation if retinal neovascularization or plus disease did not regress, and group 2B - including patients with initial laser photocoagulation and IVR as rescue therapy. Favorable outcomes were regression of the retinal neovascularization and plus disease, meaning control of the disease. Unfavorable outcomes were progression to stages 4 and 5 of ROP. RESULTS: Fifty-seven eyes were included in the study. Mean birth weight and gestational age were 1,281±254 g and 29.5±2.1 weeks, respectively. Group 1 comprised of 16 eyes, with favorable outcomes in 14 eyes (87.5%). Group 2 comprised of 41 eyes, with favorable outcomes in 29 eyes (70.7%), in a mean follow-up period of 12.8 months. CONCLUSION: IVR was effective to treat severe cases of ROP as a primary or a combined treatment. Forty-three of the 57 treated eyes (75.4%) achieved regression of ROP and favorable outcomes.
RESUMO
BACKGROUND: ROP16 is a protein kinase of Toxoplasma gondii identified in the mouse model as a virulent marker, but it is unknown whether this finding is relevant in human toxoplasmosis. METHODS: We obtained the Toxoplasma ROP16 locus DNA sequence in samples from 12 patients with ocular toxoplasmosis, 1 sample from a patient with congenital toxoplasmosis, 22 samples from soldiers operating in the jungle, 2 samples from urban soldiers, and 10 samples from meat for human consumption. An enzyme-linked immunosorbent assay specific for antibodies against the ROP16 mouse-virulent peptide was performed in 46 serum specimens from patients with ocular toxoplasmosis and in 28 serum specimens from patients with chronic asymptomatic infection, of whom 19 had congenital infection and 11 had toxoplasmic lymphadenitis. RESULTS: We found a striking divergence of the ROP16 nucleotide sequences. Ten of 12 sequences (83.3%) from patients with ocular toxoplasmosis clustered with those of mouse-virulent strains, whereas 7 of 7 ROP16 sequences (100%) from meat were clustered with those of mouse-avirulent strains. Only 11 of 104 serum specimens (10.5%) had specific antibodies against the mouse-virulent peptide, and there was no association between clinical forms and positive results of serological assays. CONCLUSIONS: The majority of ROP16 nucleotide sequences from Colombian patients with ocular toxoplasmosis belonged to the group of mouse-virulent strains.