Susceptibility of rat retina acyl-CoA: 1-acyl-sn-glycero-3-phosphocholine O-acyltransferase and CTP:phosphocholine cytidylyltransferase activity to lipid peroxidation and hydroperoxide treatment

AbstractTwo enzyme activities involved in phospholipid metabolism in the rat retina were determined after in vivo and in vitro peroxidation according to several model systems. The in vivo models were based on: (i) intravenous administration of a sonicated emulsion of phospholipid and linoleate photooxidized mixture to normal rat for a period of one week; (ii) acute injection of Fe2+ solution (20 mM) or (iii) 0.5 mg of hydroperoxylinoleate into the vitreous body, and collection of retinal tissue 4 h or 4 days later, respectively. Oleoyl CoA:lysophosphatidylcholine acyltransferase activity was unchanged or exhibited significant inhibition. On the contrary, CTP:phosphocholine cytidylyltransferase activity was stimulated. By incubating in vitro the retina with: (i) Fe2+-ascorbate; (ii) photooxidized phospholipid mixture (0.1–5 mM) or individual phospholipid classes; (iii) hydroperoxylinoleate (0.25–2 mM), with or without Fe2+, a significant inactivation of acyltransferase (six-fold maximum loss of initial activity) and a slight stimulation of cytidylyltransferase were seen. Altogether, the results suggest that in situ oxygen radical generation by a variety of agents irreversibly perturbs enzymes and/or membrane structures in which the enzymes are inserted; these events may be a causal factor in retinal degeneration accompanying some ocular diseases

A linguagem não verbal no plano de compliance: uma construção jurídica no contexto empresarial

Dissertação de mestrado em Direito dos Contratos e da EmpresaA presente pesquisa se propõe a realizar o cotejo entre duas grandes áreas, a primeira trata-se do compliance e a segunda da análise de linguagem não verbal. Sendo assim o debate proposto possui uma transdisciplinaridade, no entanto com foco no universo jurídico haja vista ser a especialidade desta pesquisa. O que se pretende, portanto, com o cotejo é uma análise da possibilidade de utilização do segundo campo dentro do primeiro, ou seja, se o compliance poderia se aproveitar das ferramentas de um outro campo do conhecimento para ter um melhoramento de suas atividades. Para tanto foi dissertado acerca do compliance, seu histórico e principalmente sua aplicação no contexto empresarial, sendo também tratado todas as questões que envolvem o tratamento de riscos, sendo nesse ponto que começa-se a tangenciar o campo de linguagem não verbal, uma vez que o principal risco tratado no presente trabalho é o risco humano, muito caro a toda gestão de risco. Sendo assim também se faz a análise acerca do que se trata a análise da linguagem não verbal, seus campos específicos, bem como a cientificidade dos métodos e sua eficácia para utilização, também sendo tratado a questão de detecção de mentiras. Por fim, o cotejo entre os dois campos se dá em uma análise aprofundada da utilidade prática de utilização da ferramenta da linguagem não verbal no plano de compliance em uma corporação, bem como toda legalidade que envolve essa aplicação sendo visto do ponto de vista ótico da proteção de dados, também sob as lentes principiológicas e o exercício hermenêutico de adequação em uma interpretação teleológica e sistêmica das normas que circundam o contexto.This research aims to carry out a comparison between two major areas, the first is compliance and the second is the analysis of non-verbal language. Thus, the proposed debate has a transdisciplinary context, however with a focus on the legal universe, as it is the specialty of this research. What is intended, therefore, with the collation is an analysis of the possibility of using the second field within the first, that is, whether compliance could take advantage of the tools of another field of knowledge to improve the activities. In this hand was made an analysis about compliance, the history and especially the application in the business context was discussed, as well as all issues involving the treatment of risks, and at this point that the field of non-verbal language begins to touch on, since the main risk addressed in this paper is human risk, which is very dear to all risk management. Thus, the analysis of what the analysis of non-verbal language is about, its specific fields, as well as the scientificity of the methods and their effectiveness for use is also made, also being treated the issue of detection of deception. Finally, the comparison between the two fields takes place in an in-depth analysis of the practical usefulness of using the non-verbal language tool in the compliance plan in a corporation, as well as all the legality that involves this application being seen from the optical point of view of data protection, also under the lens of principles and the hermeneutic exercise of adequacy in a teleological and systemic interpretation of the norms that surround the context

WNT Activation and TGFβ-Smad Inhibition Potentiate Stemness of Mammalian Auditory Neuroprogenitors for High-Throughput Generation of Functional Auditory Neurons In Vitro.

Hearing loss affects over 460 million people worldwide and is a major socioeconomic burden. Both genetic and environmental factors (i.e., noise overexposure, ototoxic drug treatment and ageing), promote the irreversible degeneration of cochlear hair cells and associated auditory neurons, leading to sensorineural hearing loss. In contrast to birds, fish and amphibians, the mammalian inner ear is virtually unable to regenerate due to the limited stemness of auditory progenitors, and no causal treatment is able to prevent or reverse hearing loss. As of today, a main limitation for the development of otoprotective or otoregenerative therapies is the lack of efficient preclinical models compatible with high-throughput screening of drug candidates. Currently, the research field mainly relies on primary organotypic inner ear cultures, resulting in high variability, low throughput, high associated costs and ethical concerns. We previously identified and characterized the phoenix auditory neuroprogenitors (ANPGs) as highly proliferative progenitor cells isolated from the A/J mouse cochlea. In the present study, we aim at identifying the signaling pathways responsible for the intrinsic high stemness of phoenix ANPGs. A transcriptomic comparison of traditionally low-stemness ANPGs, isolated from C57Bl/6 and A/J mice at early passages, and high-stemness phoenix ANPGs was performed, allowing the identification of several differentially expressed pathways. Based on differentially regulated pathways, we developed a reprogramming protocol to induce high stemness in presenescent ANPGs (i.e., from C57Bl6 mouse). The pharmacological combination of the WNT agonist (CHIR99021) and TGFβ/Smad inhibitors (LDN193189 and SB431542) resulted in a dramatic increase in presenescent neurosphere growth, and the possibility to expand ANPGs is virtually limitless. As with the phoenix ANPGs, stemness-induced ANPGs could be frozen and thawed, enabling distribution to other laboratories. Importantly, even after 20 passages, stemness-induced ANPGs retained their ability to differentiate into electrophysiologically mature type I auditory neurons. Both stemness-induced and phoenix ANPGs resolve a main bottleneck in the field, allowing efficient, high-throughput, low-cost and 3R-compatible in vitro screening of otoprotective and otoregenerative drug candidates. This study may also add new perspectives to the field of inner ear regeneration

Tracking the immunopathological response to Pseudomonas aeruginosa during respiratory infections

Repeated cycles of infections, caused mainly by Pseudomonas aeruginosa, combined with a robust host immune response and tissue injury, determine the course and outcome of cystic fibrosis (CF) lung disease. As the disease progresses, P. aeruginosa adapts to the host modifying dramatically its phenotype; however, it remains unclear whether and how bacterial adaptive variants and their persistence influence the pathogenesis and disease development. Using in vitro and murine models of infection, we showed that P. aeruginosa CF-adaptive variants shaped the innate immune response favoring their persistence. Next, we refined a murine model of chronic pneumonia extending P. aeruginosa infection up to three months. In this model, including CFTR-deficient mice, we unveil that the P. aeruginosa persistence lead to CF hallmarks of airway remodelling and fibrosis, including epithelial hyperplasia and structure degeneration, goblet cell metaplasia, collagen deposition, elastin degradation and several additional markers of tissue damage. This murine model of P. aeruginosa chronic infection, reproducing CF lung pathology, will be instrumental to identify novel molecular targets and test newly tailored molecules inhibiting chronic inflammation and tissue damage processes in pre-clinical studies

Huntingtin’s neuroprotective activity occurs via inhibition of pro-caspase 9 processing

Huntington's Disease is an inherited neurodegenerative disease that affects the medium spiny neurons in the striatum. The disease is caused by the expansion of a polyglutamine sequence in the N terminus of Huntingtin (Htt), a widely expressed protein. Recently, we have found that Htt is an antiapoptotic protein in striatal cells and acts by preventing caspase-3 activity. Here we report that Htt overexpression in other CNS-derived cells can protect them from more than 20 days exposure to fatal stimuli. In particular, we found that cytochrome c continues to be released from mitochondria into the cytosol of cells that overexpress normal Htt. However, procaspase-9 is not processed, indicating that wild-type Htt (wtHtt) acts downstream of cytochrome c release. These data show that Htt inhibits neuronal cell death by interfering with the activity of the apoptosome complex

Inhibitory Kcnip2 neurons of the spinal dorsal horn control behavioral sensitivity to environmental cold

Proper sensing of ambient temperature is of utmost importance for the survival of euthermic animals, including humans. While considerable progress has been made in our understanding of temperature sensors and transduction mechanisms, the higher-order neural circuits processing such information are still only incompletely understood. Using intersectional genetics in combination with circuit tracing and functional neuron manipulation, we identified Kcnip2-expressing inhibitory (Kcnip2GlyT2) interneurons of the mouse spinal dorsal horn as critical elements of a neural circuit that tunes sensitivity to cold. Diphtheria toxin-mediated ablation of these neurons increased cold sensitivity without affecting responses to other somatosensory modalities, while their chemogenetic activation reduced cold and also heat sensitivity. We also show that Kcnip2GlyT2 neurons become activated preferentially upon exposure to cold temperatures and subsequently inhibit spinal nociceptive output neurons that project to the lateral parabrachial nucleus. Our results thus identify a hitherto unknown spinal circuit that tunes cold sensitivity. Keywords: circuit; cold; cold allodynia; cold analgesia; cooling; dre recombinase; interneuron; intersectional gene targeting; kcnip2; pai

IL-17A impairs host tolerance during airway chronic infection by Pseudomonas aeruginosa

Resistance and tolerance mechanisms participate to the interplay between host and pathogens. IL-17-mediated response has been shown to be crucial for host resistance to respiratory infections, whereas its role in host tolerance during chronic airway colonization is still unclear. Here, we investigated whether IL-17-mediated response modulates mechanisms of host tolerance during airways chronic infection by P. aeruginosa. First, we found that IL-17A levels were sustained in mice at both early and advanced stages of P. aeruginosa chronic infection and confirmed these observations in human respiratory samples from cystic fibrosis patients infected by P. aeruginosa. Using IL-17a(-/-) or IL-17ra(-/-) mice, we found that the deficiency of IL-17A/IL-17RA axis was associated with: i) increased incidence of chronic infection and bacterial burden, indicating its role in the host resistance to P. aeruginosa; ii) reduced cytokine levels (KC), tissue innate immune cells and markers of tissue damage (pro-MMP-9, elastin degradation, TGF-β1), proving alteration of host tolerance. Blockade of IL-17A activity by a monoclonal antibody, started when chronic infection is established, did not alter host resistance but increased tolerance. In conclusion, this study identifies IL-17-mediated response as a negative regulator of host tolerance during P. aeruginosa chronic airway infection

N6-Furfuryladenine is protective in Huntington’s disease models by signaling huntingtin phosphorylation

© 2018 National Academy of Sciences. All Rights Reserved. The huntingtin N17 domain is a modulator of mutant huntingtin toxicity and is hypophosphorylated in Huntington’s disease (HD). We conducted high-content analysis to find compounds that could restore N17 phosphorylation. One lead compound from this screen was N6-furfuryladenine (N6FFA). N6FFA was protective in HD model neurons, and N6FFA treatment of an HD mouse model corrects HD phenotypes and eliminates cortical mutant huntingtin inclusions. We show that N6FFA restores N17 phosphorylation levels by being salvaged to a triphosphate form by adenine phosphoribosyltransferase (APRT) and used as a phosphate donor by casein kinase 2 (CK2). N6FFA is a naturally occurring product of oxidative DNA damage. Phosphorylated huntingtin functionally redistributes and colocalizes with CK2, APRT, and N6FFA DNA ad-ducts at sites of induced DNA damage. We present a model in which this natural product compound is salvaged to provide a triphosphate substrate to signal huntingtin phosphorylation via CK2 during low-ATP stress under conditions of DNA damage, with protective effects in HD model systems