10 results on '"Fernandes BHV"'
Search Results
2. Homozygous CDH2 variant may be associated with hypopituitarism without neurological disorders.
- Author
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Ferreira NGBP, Madeira JLO, Gergics P, Kertsz R, Marques JM, Trigueiro NSS, Benedetti AFF, Azevedo BV, Fernandes BHV, Bissegatto DD, Biscotto IP, Fang Q, Ma Q, Ozel AB, Li J, Camper SA, Jorge AAL, Mendonça BB, Arnhold IJP, and Carvalho LR
- Abstract
Context: Congenital hypopituitarism is a genetically heterogeneous condition. Whole exome sequencing (WES) is a promising approach for molecular diagnosis of patients with this condition., Objectives: The aim of this study is to conduct WES in a patient with congenital hypopituitarism born to consanguineous parents, CDH2 screening in a cohort of patients with congenital hypopituitarism, and functional testing of a novel CDH2 variant., Design: Genomic DNA from a proband and her consanguineous parents was analyzed by WES. Copy number variants were evaluated. The genetic variants were filtered for population frequency (ExAC, 1000 genomes, gnomAD, and ABraOM), in silico prediction of pathogenicity, and gene expression in the pituitary and/or hypothalamus. Genomic DNA from 145 patients was screened for CDH2 by Sanger sequencing., Results: One female patient with deficiencies in growth hormone, thyroid-stimulating hormone, adrenocorticotropic hormone, luteinizing hormone, and follicle-stimulating hormone and ectopic posterior pituitary gland contained a rare homozygous c.865G>A (p.Val289Ile) variant in CDH2. To determine whether the p.Val289Ile variant in CDH2 affects cell adhesion properties, we stably transfected L1 fibroblast lines, labeled the cells with lipophilic dyes, and quantified aggregation. Large aggregates formed in cells expressing wildtype CDH2, but aggregation was impaired in cells transfected with variant CDH2 or non-transfected., Conclusion: A homozygous CDH2 allelic variant was found in one hypopituitarism patient, and the variant impaired cell aggregation function in vitro. No disease-causing variants were found in 145 other patients screened for CDH2 variants. Thus, CDH2 is a candidate gene for hypopituitarism that needs to be tested in different populations., Significance Statement: A female patient with hypopituitarism was born from consanguineous parents and had a homozygous, likely pathogenic, CDH2 variant that impairs cell aggregation in vitro. No other likely pathogenic variants in CDH2 were identified in 145 hypopituitarism patients.
- Published
- 2023
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3. A novel insight on SARS-CoV-2 S-derived fragments in the control of the host immunity.
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Bastos TSB, de Paula AGP, Dos Santos Luz RB, Garnique AMB, Belo MAA, Eto SF, Fernandes DC, Ferraris FK, de Pontes LG, França TT, Barcellos LJG, Veras FP, Bermejo P, Guidelli G, Maneira C, da Silveira Bezerra de Mello F, Teixeira G, Pereira GAG, Fernandes BHV, Sanches PRS, Braz HLB, Jorge RJB, Malafaia G, Cilli EM, Olivier DDS, do Amaral MS, Medeiros RJ, Condino-Neto A, Carvalho LR, Machado-Santelli GM, Charlie-Silva I, Galindo-Villegas J, and Braga TT
- Subjects
- Animals, Humans, Zebrafish, Macrophages, Peptides, SARS-CoV-2, COVID-19
- Abstract
Despite all efforts to combat the pandemic of COVID-19, we are still living with high numbers of infected persons, an overburdened health care system, and the lack of an effective and definitive treatment. Understanding the pathophysiology of the disease is crucial for the development of new technologies and therapies for the best clinical management of patients. Since the manipulation of the whole virus requires a structure with an adequate level of biosafety, the development of alternative technologies, such as the synthesis of peptides from viral proteins, is a possible solution to circumvent this problem. In addition, the use and validation of animal models is of extreme importance to screen new drugs and to compress the organism's response to the disease. Peptides derived from recombinant S protein from SARS-CoV-2 were synthesized and validated by in silico, in vitro and in vivo methodologies. Macrophages and neutrophils were challenged with the peptides and the production of inflammatory mediators and activation profile were evaluated. These peptides were also inoculated into the swim bladder of transgenic zebrafish larvae at 6 days post fertilization (dpf) to mimic the inflammatory process triggered by the virus, which was evaluated by confocal microscopy. In addition, toxicity and oxidative stress assays were also developed. In silico and molecular dynamics assays revealed that the peptides bind to the ACE2 receptor stably and interact with receptors and adhesion molecules, such as MHC and TCR, from humans and zebrafish. Macrophages stimulated with one of the peptides showed increased production of NO, TNF-α and CXCL2. Inoculation of the peptides in zebrafish larvae triggered an inflammatory process marked by macrophage recruitment and increased mortality, as well as histopathological changes, similarly to what is observed in individuals with COVID-19. The use of peptides is a valuable alternative for the study of host immune response in the context of COVID-19. The use of zebrafish as an animal model also proved to be appropriate and effective in evaluating the inflammatory process, comparable to humans., (© 2023. The Author(s).)
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- 2023
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4. Photobiomodulation Reduces the Cytokine Storm Syndrome Associated with COVID-19 in the Zebrafish Model.
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Rosa IF, Peçanha APB, Carvalho TRB, Alexandre LS, Ferreira VG, Doretto LB, Souza BM, Nakajima RT, da Silva P, Barbosa AP, Gomes-de-Pontes L, Bomfim CG, Machado-Santelli GM, Condino-Neto A, Guzzo CR, Peron JPS, Andrade-Silva M, Câmara NOS, Garnique AMB, Medeiros RJ, Ferraris FK, Barcellos LJG, Correia-Junior JD, Galindo-Villegas J, Machado MFR, Castoldi A, Oliveira SL, Costa CC, Belo MAA, Galdino G, Sgro GG, Bueno NF, Eto SF, Veras FP, Fernandes BHV, Sanches PRS, Cilli EM, Malafaia G, Nóbrega RH, Garcez AS, Carrilho E, and Charlie-Silva I
- Subjects
- Animals, Humans, Zebrafish metabolism, SARS-CoV-2 metabolism, Cytokine Release Syndrome, Cytokines metabolism, RNA, Messenger, Membrane Proteins, Mitochondrial Proteins, COVID-19
- Abstract
Although the exact mechanism of the pathogenesis of coronavirus SARS-CoV-2 (COVID-19) is not fully understood, oxidative stress and the release of pro-inflammatory cytokines have been highlighted as playing a vital role in the pathogenesis of the disease. In this sense, alternative treatments are needed to reduce the level of inflammation caused by COVID-19. Therefore, this study aimed to investigate the potential effect of red photobiomodulation (PBM) as an attractive therapy to downregulate the cytokine storm caused by COVID-19 in a zebrafish model. RT-qPCR analyses and protein-protein interaction prediction among SARS-CoV-2 and Danio rerio proteins showed that recombinant Spike protein (rSpike) was responsible for generating systemic inflammatory processes with significantly increased levels of pro-inflammatory ( il1b , il6 , tnfa , and nfkbiab ), oxidative stress ( romo1 ) and energy metabolism ( slc2a1a and coa1 ) mRNA markers, with a pattern similar to those observed in COVID-19 cases in humans. On the other hand, PBM treatment was able to decrease the mRNA levels of these pro-inflammatory and oxidative stress markers compared with rSpike in various tissues, promoting an anti-inflammatory response. Conversely, PBM promotes cellular and tissue repair of injured tissues and significantly increases the survival rate of rSpike-inoculated individuals. Additionally, metabolomics analysis showed that the most-impacted metabolic pathways between PBM and the rSpike treated groups were related to steroid metabolism, immune system, and lipid metabolism. Together, our findings suggest that the inflammatory process is an incisive feature of COVID-19 and red PBM can be used as a novel therapeutic agent for COVID-19 by regulating the inflammatory response. Nevertheless, the need for more clinical trials remains, and there is a significant gap to overcome before clinical trials can commence.
- Published
- 2023
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5. Virulomic Analysis of Multidrug-Resistant Klebsiella pneumoniae Isolates and Experimental Virulence Model Using Danio rerio (Zebrafish).
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Duarte ELT, Rizek CF, Espinoza ES, Marchi AP, Noguera SV, Côrtes MF, Fernandes BHV, Guimarães T, de Maio Carrilho CMD, Neto LVP, Trindade PA, and Costa SF
- Abstract
This study evaluates a possible correlation between multidrug-resistant Klebsiella pneumoniae strains and virulence markers in a Danio rerio (zebrafish) model. Whole-genome sequencing (WGS) was performed on 46 strains from three Brazilian hospitals. All of the isolates were colistin-resistant and harbored bla
KPC-2 . Ten different sequence types (STs) were found; 63% belonged to CC258, 22% to ST340, and 11% to ST16. The virulence factors most frequently found were type 3 fimbriae, siderophores, capsule regulators, and RND efflux-pumps. Six strains were selected for a time-kill experiment in zebrafish embryos: infection by ST16 was associated with a significantly higher mortality rate when compared to non-ST16 strains (52% vs. 29%, p = 0.002). Among the STs, the distribution of virulence factors did not differ significantly except for ST23, which harbored a greater variety of factors than other STs but was not related to a higher mortality rate in zebrafish. Although several virulence factors are described in K. pneumoniae , our study found ST16 to be the only significant predictor of a virulent phenotype in an animal model. Further research is needed to fully understand the correlation between virulence and sequence types.- Published
- 2022
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6. Modular Label-Free Electrochemical Biosensor Loading Nature-Inspired Peptide toward the Widespread Use of COVID-19 Antibody Tests.
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Castro ACH, Bezerra ÍRS, Pascon AM, da Silva GH, Philot EA, de Oliveira VL, Mancini RSN, Schleder GR, Castro CE, de Carvalho LRS, Fernandes BHV, Cilli EM, Sanches PRS, Santhiago M, Charlie-Silva I, Martinez DST, Scott AL, Alves WA, and Lima RS
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- COVID-19 Testing, Carbon chemistry, Electrochemical Techniques, Electrodes, Gold chemistry, Humans, Molecular Docking Simulation, Peptides chemistry, Biosensing Techniques methods, COVID-19 diagnosis, Metal Nanoparticles chemistry
- Abstract
Limitations of the recognition elements in terms of synthesis, cost, availability, and stability have impaired the translation of biosensors into practical use. Inspired by nature to mimic the molecular recognition of the anti-SARS-CoV-2 S protein antibody (Ab
S ) by the S protein binding site, we synthesized the peptide sequence of Asn-Asn-Ala-Thr-Asn-COOH (abbreviated as PEP2003) to create COVID-19 screening label-free (LF) biosensors based on a carbon electrode, gold nanoparticles (AuNPs), and electrochemical impedance spectroscopy. The PEP2003 is easily obtained by chemical synthesis, and it can be adsorbed on electrodes while maintaining its ability for AbS recognition, further leading to a sensitivity 3.4-fold higher than the full-length S protein, which is in agreement with the increase in the target-to-receptor size ratio. Peptide-loaded LF devices based on noncovalent immobilization were developed by affording fast and simple analyses, along with a modular functionalization. From studies by molecular docking, the peptide-AbS binding was found to be driven by hydrogen bonds and hydrophobic interactions. Moreover, the peptide is not amenable to denaturation, thus addressing the trade-off between scalability, cost, and robustness. The biosensor preserves 95.1% of the initial signal for 20 days when stored dry at 4 °C. With the aid of two simple equations fitted by machine learning (ML), the method was able to make the COVID-19 screening of 39 biological samples into healthy and infected groups with 100.0% accuracy. By taking advantage of peptide-related merits combined with advances in surface chemistry and ML-aided accuracy, this platform is promising to bring COVID-19 biosensors into mainstream use toward straightforward, fast, and accurate analyses at the point of care, with social and economic impacts being achieved.- Published
- 2022
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7. Immunization with SARS-CoV-2 Nucleocapsid protein triggers a pulmonary immune response in rats.
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Silva EKVB, Bomfim CG, Barbosa AP, Noda P, Noronha IL, Fernandes BHV, Machado RRG, Durigon EL, Catanozi S, Rodrigues LG, Pieroni F, Lima SG, Teodoro WR, Queiroz ZAJ, Silveira LKR, Charlie-Silva I, Capelozzi VL, Guzzo CR, and Fanelli C
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- Animals, Antibodies, Viral, COVID-19 Vaccines, Humans, Immunity, Immunization, Lung, Nucleocapsid Proteins, Rats, Rats, Inbred Lew, Rats, Wistar, Recombinant Proteins, Spike Glycoprotein, Coronavirus, Vaccination, COVID-19 prevention & control, SARS-CoV-2
- Abstract
The SARS-CoV-2 pandemic have been affecting millions of people worldwide, since the beginning of 2020. COVID-19 can cause a wide range of clinical symptoms, which varies from asymptomatic presentation to severe respiratory insufficiency, exacerbation of immune response, disseminated microthrombosis and multiple organ failure, which may lead to dead. Due to the rapid spread of SARS-CoV-2, the development of vaccines to minimize COVID-19 severity in the world population is imperious. One of the employed techniques to produce vaccines against emerging viruses is the synthesis of recombinant proteins, which can be used as immunizing agents. Based on the exposed, the aim of the present study was to verify the systemic and immunological effects of IM administration of recombinant Nucleocapsid protein (NP), derived from SARS-CoV-2 and produced by this research group, in 2 different strains of rats (Rattus norvegicus); Wistar and Lewis. For this purpose, experimental animals received 4 injections of NP, once a week, and were submitted to biochemical and histological analysis. Our results showed that NP inoculations were safe for the animals, which presented no clinical symptoms of worrying side effects, nor laboratorial alterations in the main biochemical and histological parameters, suggesting the absence of toxicity induced by NP. Moreover, NP injections successfully triggered the production of specific anti-SARS-CoV-2 IgG antibodies by both Wistar and Lewis rats, showing the sensitization to have been well sufficient for the immunization of these strains of rats. Additionally, we observed the local lung activation of the Bronchus-Associated Lymphoid Tissue (BALT) of rats in the NP groups, suggesting that NP elicits specific lung immune response. Although pre-clinical and clinical studies are still required, our data support the recombinant NP produced by this research group as a potential immunizing agent for massive vaccination, and may represent advantages upon other recombinant proteins, since it seems to induce specific pulmonary protection., Competing Interests: The authors declare that no competing interests exist.
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- 2022
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8. Toxicological insights of Spike fragments SARS-CoV-2 by exposure environment: A threat to aquatic health?
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Charlie-Silva I, Araújo APC, Guimarães ATB, Veras FP, Braz HLB, de Pontes LG, Jorge RJB, Belo MAA, Fernandes BHV, Nóbrega RH, Galdino G, Condino-Neto A, Galindo-Villegas J, Machado-Santelli GM, Sanches PRS, Rezende RM, Cilli EM, and Malafaia G
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- Animals, Anura, Humans, Larva, Spike Glycoprotein, Coronavirus, COVID-19, SARS-CoV-2
- Abstract
The Spike protein (S protein) is a critical component in the infection of the new coronavirus (SARS-CoV-2). The objective of this work was to evaluate whether peptides from S protein could cause negative impact in the aquatic animals. The aquatic toxicity of SARS-CoV-2 Spike protein peptides derivatives has been evaluated in tadpoles (n = 50 tadpoles/5 replicates of 10 animals) from species Physalaemus cuvieri (Leptodactylidae). After synthesis, purification, and characterization of peptides (PSDP2001, PSDP2002, PSDP2003) an aquatic contamination has been simulated with these peptides during 24 h of exposure in two concentrations (100 and 500 ng/mL). The control group ("C") was composed of tadpoles kept in polyethylene containers containing de-chlorinated water. Oxidative stress, antioxidant biomarkers and AChE activity were assessed. In both concentrations, PSPD2002 and PSPD2003 increased catalase and superoxide dismutase antioxidants enzymes activities, as well as oxidative stress (nitrite levels, hydrogen peroxide and reactive oxygen species). All three peptides also increased acetylcholinesterase activity in the highest concentration. These peptides showed molecular interactions in silico with acetylcholinesterase and antioxidant enzymes. Aquatic particle contamination of SARS-CoV-2 has cholinesterasic effect in P. cuvieri tadpoles. These findings indicate that the COVID-19 can constitute environmental impact or biological damage potential., (Copyright © 2021 Elsevier B.V. All rights reserved.)
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- 2021
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9. Zebrafish embryo sensitivity test as in vivo platform to anti-Shiga toxin compound screening.
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de Sousa Melo B, Fernandes BHV, Lopes-Ferreira MVA, Henrique C, Piazza RMF, and Luz D
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- Animals, Drug Evaluation, Preclinical, Embryo, Nonmammalian, Lethal Dose 50, Shiga Toxin toxicity, Shiga-Toxigenic Escherichia coli chemistry, Zebrafish, Antitoxins pharmacology, Shiga Toxin antagonists & inhibitors
- Abstract
Shiga toxin-producing Escherichia coli (STEC) pathotype secretes two types of AB
5 cytotoxins (Stx1 and Stx2), responsible for complications such as hemorrhagic colitis (HC) and hemolytic uremic syndrome (HUS) in infected patients, which could lead to sequels and death. Currently, there is no effective treatment against the cytotoxic effect of these toxins. However, in order to approve any therapy molecule, an animal experiment is required in order to evaluate the efficacy and safety of therapeutic approaches. The use of alternative small host models is growing among human infectious disease studies, particularly the vertebrate zebrafish model, since relevant results have been described for pathogen-host interaction. In this sense, the present work aimed to analyze the toxic effect of Shiga toxins in zebrafish embryo model in order to standardize this method in the future to be used as a fast, simple, and efficient methodology for the screening of therapeutic molecules. Herein, we demonstrated that the embryos were sensitive in a dose-dependent manner to both Stx toxins, with LD50 of 22 μg/mL for Stx1 and 33 μg/mL for Stx2, and the use of anti-Stx polyclonal antibody abolished the toxic effect. Therefore, this methodology can be a rapid alternative method for selecting promising compounds against Stx toxins, such as recombinant antibodies.- Published
- 2020
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10. An Easy Method for Cryopreservation of Zebrafish ( Danio rerio ) Sperm.
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Caetano Da Silva C, Kollár T, Csenki-Bakos Z, Fernandes BHV, Horváth Á, and Carvalho LR
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- Animals, Male, Reproducibility of Results, Cryopreservation methods, Semen Preservation methods, Spermatozoa physiology, Zebrafish
- Abstract
We developed an easy, efficient, and cheap protocol for zebrafish sperm cryopreservation carried out on dry ice (20 min) using simple composition solution (200 mM glucose, 40 mM KCl, 30 mM Tris, pH = 8.0). The average efficiency of the present cryopreserve method was between 10% and 20% (expressed as fertilization rate). The experiments were conducted and repeated at two different locations, in different countries, yielding very similar results, showing the reproducibility and applicability of the method.
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- 2019
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