23 registros encontrados - material ARTIGO E unidade Centro Multidisciplinar para Investigação Biológica na Área da Ciência de Animais de Laboratório
ARTIGO - Inglês
Assunto: Nanotecnologia
One of the important components in tissue engineering is material structure, providing a model for fixing and the development of cells and tissues,...
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One of the important components in tissue engineering is material structure, providing a model for fixing and the development of cells and tissues, which allows for the transport of nutrients and regulatory molecules to and from cells. The community claims the need for new materials with better properties for use in the clinic. Poly (ε-caprolactone) (PCL) is a biodegradable polymer, semi crystalline, with superior mechanical properties and has attracted an increasing interest due to its usefulness in various biomedical applications. Herein, two different methods (electrospinning versus rotary jet spinning) with different concentrations of PCL produced ultra thin-fibers each with particular characteristics, verified and analyzed by morphology, wettability, thermal and cytotoxicity features and for bacteria colonization. Different PCL scaffold morphologies were found to be dependent on the fabrication method used. All PCL scaffolds showed greater mammalian cell interactions. Most impressively, rotary-jet spun fibers showed that a special rough surface decreased bacteria colonization, emphasizing that no nanoparticle or antibiotic was used; maybe this effect is related with physical (scaffold) and/or biological mechanisms. Thus, this study showed that rotary jet spun fibers possess a special topography compared to electrospun fibers to reduce bacteria colonization and present no cytotoxicity when in contact with mammalian cells
ARTIGO - Inglês
Assunto: Células matadoras naturais
Natural killer (NK) cells have long been considered short-lived effectors of innate immunity. However, recent animal models and human studies suggest...
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Natural killer (NK) cells have long been considered short-lived effectors of innate immunity. However, recent animal models and human studies suggest that subsets of NK cells have adaptive features. We investigate clonal relationships of various NK-cell subsets, including the adaptive population, by taking advantage of naturally occurring X-linked somatic PIGA mutations in hematopoietic stem and progenitor cells (HSPCs) from patients with paroxysmal nocturnal hemoglobinuria (PNH). The affected HSPCs and their progeny lack expression of glycosylphosphatidylinositol (GPI) anchors on their cell surface, allowing quantification of PIGA-mutant (GPI-negative) HSPC-derived peripheral blood cell populations. The fraction of GPI-negative cells within the CD56(dim) NK cells was markedly lower than that of neutrophils and the CD56(bright) NK-cell compartments. This discrepancy was most prominent within the adaptive CD56(dim) NK-cell population lacking PLZF expression. The functional properties of these adaptive NK cells were similar in PNH patients and healthy individuals. Our findings support the existence of a long-lived, adaptive NK-cell population maintained independently from GPI(pos)CD56(dim)
ARTIGO - Inglês
Assunto: Nanotubos de carbono
Biocompatibility tests were performed on vertically-aligned multi-walled carbon nanotube (MWCNT) films produced by microwave plasma chemical vapor...
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Biocompatibility tests were performed on vertically-aligned multi-walled carbon nanotube (MWCNT) films produced by microwave plasma chemical vapor deposition on titanium substrates with iron (Fe) and nickel (Ni) as catalysts. The cell adhesion and morphology of L-929 mouse fibroblast cells were studied by high resolution scanning electron microscopy, after up to 7 days incubation periods. Cell viability and proliferation were evaluated by two "in vitro" tests: (1) 2-(4,5-dimethyl-2-thiazolyl)-3,5-diphenyl-2H-tetrazolium bromide (MTT), and (2) lactate dehydrogenase (LDH) assays. Low level of bioavailable Fe and Ni was determined by inductively coupled plasma optical emission spectrometry. Neither functionalization nor purification of MWCNT films was necessary to obtain good response to the biocompatibility tests. Efficient cell growth and non-toxicity suggest the use MWCNTs in tissue regeneration. The MWCNT films stimulated the cell growth, showing a proliferation 20% higher than on Ti
ARTIGO - Inglês
Assunto: Adesão celular
Abstract: We analyzed GFP cells after 24 h cultivated on superhydrophilic vertically aligned carbon nanotube scaffolds. We produced two different...
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Abstract: We analyzed GFP cells after 24 h cultivated on superhydrophilic vertically aligned carbon nanotube scaffolds. We produced two different densities of VACNT scaffolds on Ti using Ni or Fe catalysts. A simple and fast oxygen plasma treatment promoted the superhydrophilicity of them. We used five different substrates, such as: as-grown VACNT produced using Ni as catalyst (Ni), as-grown VACNT produced using Fe as catalyst (Fe), VACNT-O produced using Ni as catalyst (Nisingle bondO), VACNT-O produced using Fe as catalyst (Fesingle bondO) and Ti (control). The 4',6-diamidino-2-phenylindole reagent nuclei stained the adherent cells cultivated on five different analyzed scaffolds. We used fluorescence microscopy for image collect, ImageJ® to count adhered cell and GraphPad Prism 5® for statistical analysis. We demonstrated in crescent order: Fe, Ni, Nisingle bondO, Fesingle bondO and Ti scaffolds that had an improved cellular adhesion. Oxygen treatment associated to high VACNT density (group Fesingle bondO) presented significantly superior cell adhesion up to 24 h. However, they do not show significant differences compared with Ti substrates (control). We demonstrated that all the analyzed substrates were nontoxic. Also, we proposed that the density and hydrophilicity influenced the cell adhesion behavior
Bartonella henselae transmission by blood transfusion in mice
ARTIGO - Inglês
Assunto: Bartonella
Bartonella spp. are neglected fastidious Gram‐negative bacilli. We isolated Bartonella henselae from 1.2% of 500 studied blood donors and demonstrated...
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Bartonella spp. are neglected fastidious Gram‐negative bacilli. We isolated Bartonella henselae from 1.2% of 500 studied blood donors and demonstrated that the bacteria remain viable in red blood cell units after 35 days of experimental infection. Now, we aim to evaluate the possibility of B. henselae transmission by blood transfusion in a mouse model. STUDY DESIGN AND METHODS Eight BALB/c mice were intraperitoneal inoculated with a 30 µL of suspension with 104 CFU/mL of B. henselae and a second group of eight mice were inoculated with saline solution and used as control. After 96 hours of inoculation, the animals were euthanized. We collected blood and tissue samples from skin, liver, and spleen. Thirty microliters of blood from four Bartonella‐inoculated animals were transfused into a new group (n = 4). Another group received blood from the control animals. B. henselae infection was investigated by conventional and nested polymerase chain reaction (PCR). RESULTS Blood samples from all 24 mice were negative by molecular tests though half of the tissue samples were positive by nested PCR in the intraperitoneal Bartonella‐investigated animals. Tissues from two of the four mice that received blood transfusions from Bartonella‐inoculated animals were also nested PCR positives. CONCLUSIONS Transmission of B. henselae by transfusion is possible in mice even when donor animals have undetectable bloodstream infection. The impact of human Bartonella sp. transmission through blood transfusion recipients must be evaluated
Benchmarking the proteomic profile of animal models of mesial temporal epilepsy
ARTIGO - Inglês
Assunto: Artigo de pesquisa
Abstract: We compared the proteomic signatures of the hippocampal lesion induced in three different animal models of mesial temporal lobe epilepsy...
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Abstract: We compared the proteomic signatures of the hippocampal lesion induced in three different animal models of mesial temporal lobe epilepsy with hippocampal sclerosis (MTLE+HS): the systemic pilocarpine model (PILO), the intracerebroventricular kainic acid model (KA), and the perforant pathway stimulation model (PPS). We used shotgun proteomics to analyze the proteomes and find enriched biological pathways of the dorsal and ventral dentate gyrus (DG) isolated from the hippocampi of the three animal models. We also compared the proteomes obtained in the animal models to that from the DG of patients with pharmacoresistant MTLE+HS. We found that each animal model presents specific profiles of proteomic changes. The PILO model showed responses predominantly related to neuronal excitatory imbalance. The KA model revealed alterations mainly in synaptic activity. The PPS model displayed abnormalities in metabolism and oxidative stress. We also identified common biological pathways enriched in all three models, such as inflammation and immune response, which were also observed in tissue from patients. However, none of the models could recapitulate the profile of molecular changes observed in tissue from patients SIGNIFICANCE: Our results indicate that each model has its own set of biological responses leading to epilepsy. Thus, it seems that only using a combination of the three models may one replicate more closely the mechanisms underlying MTLE+HS as seen in patients
DOI: https://doi.org/10.1002/acn3.51533
Texto completo: https://onlinelibrary.wiley.com/doi/10.1002/acn3.51533
ARTIGO - Inglês
Assunto: Primatas
Abstract: Recent advances in gene editing technologies using CRISPR/Cas9 allow precise genome editing at a site of interest and have accelerated human...
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Abstract: Recent advances in gene editing technologies using CRISPR/Cas9 allow precise genome editing at a site of interest and have accelerated human disease modeling and the development of corrective gene therapies for various genetic disorders.1,2 We adapted CRISPR/Cas9 editing of rhesus macaque (RM) hematopoietic stem and progenitor cells (HSPCs) to create the first engineered large animal model of a hematologic disease based on close phylogenetic/functional similarity of RM to human HSPCs.3 Paroxysmal nocturnal hemoglobinuria (PNH) is a clonal hematopoietic disorder characterized by intravascular hemolysis, thrombosis, and often bone marrow failure, linked to acquired somatic loss-of-function mutations in HSPCs inactivating the X-linked PIG-A gene that encodes an enzyme necessary for cell surface expression of glycosylphophatidylinositol-anchored proteins (GPI-APs), such as CD55 and CD59.4,5 Mutant HSPCs in PNH clonally expand, often dominating erythroid, myeloid, and B-cell lineages.6 Several hypotheses have been proposed to explain PNH HSPC clonal expansion, including intrinsic resistance to apoptosis,7 additional events such as a second mutation,8 and relative protection of PIG-A–deficient HSPCs from immune attack.4,9,10 However, definitive mechanisms remain unclear. There have been attempts to model PNH by using conditional knockout strategies in mice,11,12 but these do not recapitulate the hemolytic phenotype or clonal dominance. To provide further insights into PNH, we used CRISPR/Cas9 gene editing of RM HSPCs to create loss-of-function insertions/deletions (indels) in the PIG-A gene, followed by autologous transplantation and long-term follow-up (see supplemental Methods, available on the Blood Web site). We designed 3 guide RNA (gRNA) sequences targeting sites within the PIG-A exon 2 locus on the X chromosome, the most common area for mutations in PNH patients (Figure 1A). All efficiently knocked out GPI-anchored CD55 and CD59 expression in vitro in RM FRhK-4 cells and in CD34+ HSPCs. Peripheral blood HSPCs were mobilized and collected from 1 male (ZL19-M) and 1 female (ZI35-F) RM as described.13,14 Purified CD34+ cells were electroporated with Cas9 protein–gRNA complexes15 and reinfused into the autologous macaque after myeloablative total body irradiation. Each animal received a fraction of CD34+ cells electroporated with the gRNAs targeting PIG-A and a fraction with gRNAs targeting the AAVS1 locus as an internal control for editing at a safe harbor biologically inert site16 (Figure 1A). We confirmed the presence of indels in the infusion product in AAVS1 at similar levels in both macaques, whereas the PIG-A indel frequency was higher in ZL19-M than in ZI35-F, with overall levels lower than at the AAVS1 locus (Figure 1B). Figure 1C summarizes transplantation parameters. After engraftment, the PNH phenotype was assessed via staining with fluorescein-labeled proaerolysin (FLAER), a fluorescent compound that binds to all GPI anchors, and by staining for lineage-specific markers with a focus on granulocytes, which best reflect ongoing HSPC activity and have been shown to correlate with clinical PNH outcomes. Deep sequencing was used to quantitate indels at the targeted sites
ARTIGO - Inglês
Assunto: Citotoxicidade
A cytotoxicity study is performed on a poly(methyl methacrylate) polymer (PMMA) to be used for the fabrication of bone tissue by Rapid Prototyping...
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A cytotoxicity study is performed on a poly(methyl methacrylate) polymer (PMMA) to be used for the fabrication of bone tissue by Rapid Prototyping (RP). The solution polymerization is conducted in a pilot plant reactor using more appropriated reagents in consideration of the medical application. Moreover, the polymer is efficiently handled to avoid the side effect of the monomer, reducing the concentration of this specie to 287,731 mu g MMA/kg PMMA. The cytotoxicity of the polymer is determined through growth monitoring, adherence and morphology of L-929 cells. Additionally, MTT and LIVE/DEAD tests are performed. The results showed continuous and progressive growth of the cells on the surface of the specimens. Moreover, the material did not influence on the viability of mesenchymal cells and inverted fluorescence microscopy images showed a polyanionic dye calcein well retained in the cells in contact with the PMMA as well as the negative control after 72 hours. Thus, the polymer was efficiently synthesized and handled for the expected demands
ARTIGO - Inglês
Assunto: Nanotubos de carbono
Poly(D,L-lactide acid, PDLLA) has been researched for scaffolds in bone regeneration. However, its hydrophobocity and smooth surface impedes its...
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Poly(D,L-lactide acid, PDLLA) has been researched for scaffolds in bone regeneration. However, its hydrophobocity and smooth surface impedes its interaction with biological fluid and cell adhesion. To alter the surface characteristics, different surface Modification techniques have been developed to facilitate biological application. The present study compared two different routes to produce PDLLA/superhydrophilic vertically aligned carbon nanotubes:nanohydroxyapatite (PDLLA/VACNT-O:nHAp) scaffolds. For this, we used electrodeposition and immersion in simulated body fluid (SBF). Characterization by goniometry, scanning electron microscopy, X-ray diffraction, and infrared spectroscopy confirmed the polymer modifications, the in vitro bioactivity, and biomineralization. Differential sunning calorimetry and thermal gravimetric analyses showed that the inclusion of VACNT-O:nHA probably acts as a nucleating agent increasing the crystallization rate in the neat PDLLA without structural alteration. Our results showed the formation of a dense nHAp layer on all scaffolds after 14 days of immersion in SBF solution; the most intense carbonated nHAp peaks observed in the PDLLA/VACNT-O:nHAp samples suggest higher calcium precipitation compared to the PDLLA control. Both cell viability and alkaline phosphatase assays showed favorable results, because no cytotoxic effects were present and all produced scaffolds were able to induce detectable mineralization. Bone defects were used to evaluate the bone regeneration; the confocal Raman and histological results confirmed high potential for bone applications: In vivo study showed that the PDLLA/VACNT-O:nHAp scaffolds mimicked the immature bone and induced bone remodeling: These findings indicate surface improvement and the applicability of this new nanobiomaterial for hone regenerative medicine
ARTIGO - Inglês
Assunto: Nanotubos de carbono
Poly(D,L-lactide acid, PDLLA) has been researched for scaffolds in bone regeneration. However, its hydrophobocity and smooth surface impedes its...
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Resumo
Poly(D,L-lactide acid, PDLLA) has been researched for scaffolds in bone regeneration. However, its hydrophobocity and smooth surface impedes its interaction with biological fluid and cell adhesion. To alter the surface characteristics, different surface Modification techniques have been developed to facilitate biological application. The present study compared two different routes to produce PDLLA/superhydrophilic vertically aligned carbon nanotubes:nanohydroxyapatite (PDLLA/VACNT-O:nHAp) scaffolds. For this, we used electrodeposition and immersion in simulated body fluid (SBF). Characterization by goniometry, scanning electron microscopy, X-ray diffraction, and infrared spectroscopy confirmed the polymer modifications, the in vitro bioactivity, and biomineralization. Differential sunning calorimetry and thermal gravimetric analyses showed that the inclusion of VACNT-O:nHA probably acts as a nucleating agent increasing the crystallization rate in the neat PDLLA without structural alteration. Our results showed the formation of a dense nHAp layer on all scaffolds after 14 days of immersion in SBF solution; the most intense carbonated nHAp peaks observed in the PDLLA/VACNT-O:nHAp samples suggest higher calcium precipitation compared to the PDLLA control. Both cell viability and alkaline phosphatase assays showed favorable results, because no cytotoxic effects were present and all produced scaffolds were able to induce detectable mineralization. Bone defects were used to evaluate the bone regeneration; the confocal Raman and histological results confirmed high potential for bone applications: In vivo study showed that the PDLLA/VACNT-O:nHAp scaffolds mimicked the immature bone and induced bone remodeling: These findings indicate surface improvement and the applicability of this new nanobiomaterial for hone regenerative medicine
DOI: https://doi.org/10.1021/acsami.5b01066
Texto completo: https://pubs.acs.org/doi/10.1021/acsami.5b01066
ARTIGO - Inglês
Assunto: Óxido nítrico
Priapism is defined as prolonged and persistent penile erection, unassociated with sexual interest or stimulation, and is one of the many serious...
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Priapism is defined as prolonged and persistent penile erection, unassociated with sexual interest or stimulation, and is one of the many serious complications associated with sickle cell disease (SCD). Aim. The aim of this study was to evaluate the role of the NO-cGMP signaling pathway in priapism in Berkeley murine model of SCD (SS). Methods. SS mice and C57BL/6 mice (control) penile tissues were removed and the erectile tissue within the corpus cavernosum (CC) was surgically dissected free. The strips were mounted in 10 mL organ baths containing Krebs solution at 37 degrees C (95% O(2), 5% CO(2), pH 7.4), and vertically suspended between two metal hooks. Main Outcome Measures. Cumulative concentration-response curves were constructed for acetylcholine (ACh; endothelium-dependent responses), sodium nitroprusside (SNP; endothelium-independent relaxations) and BAY 41-2272 (a potent activator of NO-independent site of soluble guanylate cyclase) in CC precontracted with phenylephrine. Cavernosal responses induced by frequency-dependent electrical field stimulation (EFS) were also carried out to evaluate the nitrergic cavernosal relaxations. Results. In SS mice, ACh-induced cavernosal relaxations were leftward shifted by 2.6-fold (P < 0.01) that was accompanied by increases in the maximal responses (78 +/- 5% and 60 +/- 3% in SS and C57B6/6J mice, respectively). Similarly, SNP- and BAY 41-2272-induced CC relaxations were leftward shifted by approximately 3.3- and 2.2-fold (P < 0.01) in SS mice, respectively. A significant increase in maximal responses to SNP and BAY 41-2272 in SS mice was also observed (113 +/- 6% and 124 +/- 5%, respectively) compared with C57B6/6J mice (83 +/- 4% and 99 +/- 2%, respectively). The EFS-induced cavernosal relaxations were also significantly higher SS mice. Conclusion. These results showed that SS mice exhibit amplified corpus carvenosum relaxation response mediated by NO-cGMP signaling pathway. Intervention in this signaling pathway may be a potential therapeutic target to treat SCD priapism. Claudino MA, Franco-Penteado CF, Corat MAF, Gimenes AP, Passos LAC, Antunes E, and Costa FF. Increased cavernosal relaxations in sickle cell mice priapism are associated with alterations in the NO-cGMP signaling pathway
ARTIGO - Inglês
Assunto: Artigo original
Abstract: Mesial temporal lobe epilepsy (MTLE) is a chronic neurological disorder affecting almost 40% of adult patients with epilepsy. Hippocampal...
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Abstract: Mesial temporal lobe epilepsy (MTLE) is a chronic neurological disorder affecting almost 40% of adult patients with epilepsy. Hippocampal sclerosis (HS) is a common histopathological abnormality found in patients with MTLE. HS is characterised by extensive neuronal loss in different hippocampus sub-regions. In this study, we used laser microdissection-based microproteomics to determine the protein abundances in different regions and layers of the hippocampus dentate gyrus (DG) in an electric stimulation rodent model which displays classical HS damage similar to that found in patients with MTLE. Our results indicate that there are differences in the proteomic profiles of different layers (granule cell and molecular), as well as different regions, of the DG (ventral and dorsal). We have identified new signalling pathways and proteins present in specific layers and regions of the DG, such as PARK7, RACK1, and connexin 31/gap junction. We also found two major signalling pathways that are common to all layers and regions: inflammation and energy metabolism. Finally, our results highlight the utility of high-throughput microproteomics and spatial-limited isolation of tissues in the study of complex disorders to fully appreciate the large biological heterogeneity present in different cell populations within the central nervous system
ARTIGO - Inglês
Assunto: Ilhotas pancreáticas
Protein restriction in the early stages of life can result in several changes in pancreatic function. These alterations include documented reductions...
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Protein restriction in the early stages of life can result in several changes in pancreatic function. These alterations include documented reductions in insulin secretion and in cytoplasmic calcium concentration [Ca2+](i). However, the mechanisms underlying these changes have not been completely elucidated and may result, in part, from alterations in signaling pathways that potentiate insulin secretion in the presence of glucose. Our findings suggest that protein restriction disrupts the insulin secretory synergism between Cyclic adenosine monophosphate (cAMP)-dependent protein kinase (PKA) and Ca2+ -dependent protein kinase C (PKC) in isolated islets. Western blot analysis demonstrated reduced levels of both phospho-cAMP response element-binding protein (phospho-CREB) at Ser-133 and substrates phosphorylated by PKCs (Phospho-(Ser) PKC substrate), suggesting that PICA and PKC activity was impaired in islets from rats fed a low-protein diet (LP). cAMP levels and global Ca2+ entry were also reduced in LP islets. In summary, our findings showed that protein restriction altered the crosstalk between PICA and PKC signaling pathways, resulting in the alteration of secretory synergism in isolated islets
ARTIGO - Inglês
Assunto: Artigo original
Abstract: The pilocarpine-induced (PILO) model has helped elucidate the electrophysiological and molecular aspects related to mesial temporal lobe...
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Abstract: The pilocarpine-induced (PILO) model has helped elucidate the electrophysiological and molecular aspects related to mesial temporal lobe epilepsy. It has been suggested that the extensive cell death and edema observed in the brains of these animals could be induced by increased inflammatory responses, such as the rapid release of the inflammatory cytokine interleukin 1 beta (Il1b). In this study, we investigate the role of endogenous Il1b in the acute phase of the PILO model. Our aim is twofold. First, we want to determine whether it is feasible to silence Il1b in the central nervous system using a non-invasive procedure. Second, we aim to investigate the effect of silencing endogenous Il1b and its antagonist, Il1rn.We used RNA interference applied non-invasively to knockdown Il1b and its endogenous antagonist Il1rn. We found that knocking down Il1b prior to pilocarpine injection increased the mortality rate of treated animals. Furthermore, we observed that, when exposing the animals to more Il1b by silencing its endogenous antagonist Il1rn, there was a better response to status epilepticus with decreased animal mortality in the acute phase of the PILO model. Thus, we show the feasibility of using a novel, less invasive approach to study genes involved in the inflammatory response in the central nervous system. Furthermore, our results provide suggestive evidence that modulating endogenous Il1b improves animal survival in the acute phase of the PILO model and may have effects that extend into the chronic phase
ARTIGO - Inglês
Assunto: Artigo original
Abstract: Mesial temporal lobe epilepsy (MTLE) is a chronic neurological disorder characterized by the occurrence of seizures, and histopathological...
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Abstract: Mesial temporal lobe epilepsy (MTLE) is a chronic neurological disorder characterized by the occurrence of seizures, and histopathological abnormalities in the mesial temporal lobe structures, mainly hippocampal sclerosis (HS). We used a multi-omics approach to determine the profile of transcript and protein expression in the dorsal and ventral hippocampal dentate gyrus (DG) and Cornu Ammonis 3 (CA3) in an animal model of MTLE induced by pilocarpine. We performed label-free proteomics and RNAseq from laser-microdissected tissue isolated from pilocarpine-induced Wistar rats. We divided the DG and CA3 into dorsal and ventral areas and analyzed them separately. We performed a data integration analysis and evaluated enriched signaling pathways, as well as the integrated networks generated based on the gene ontology processes. Our results indicate differences in the transcriptomic and proteomic profiles among the DG and the CA3 subfields of the hippocampus. Moreover, our data suggest that epileptogenesis is enhanced in the CA3 region when compared to the DG, with most abnormalities in transcript and protein levels occurring in the CA3. Furthermore, our results show that the epileptogenesis in the pilocarpine model involves predominantly abnormal regulation of excitatory neuronal mechanisms mediated by N-methyl D-aspartate (NMDA) receptors, changes in the serotonin signaling, and neuronal activity controlled by calcium/calmodulin-dependent protein kinase (CaMK) regulation and leucine-rich repeat kinase 2 (LRRK2)/WNT signaling pathways
DOI: https://doi.org/10.1002/hipo.23268
Texto completo: https://onlinelibrary.wiley.com/doi/10.1002/hipo.23268
Murine norovirus infection in Brazilian animal facilities
ARTIGO - Inglês
Assunto: Rato como animal de laboratório
Murine norovirus (MNV) is a single-stranded positive-sense RNA virus of the Caliciviridae family. MNV has been reported to infect laboratory mice with...
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Murine norovirus (MNV) is a single-stranded positive-sense RNA virus of the Caliciviridae family. MNV has been reported to infect laboratory mice with the ability to cause lethal infections in strains lacking components of the innate immune response. Currently, MNV is considered the most prevalent infectious agent detected in laboratory mouse facilities. In this study, mice in 22 laboratory animal facilities within Brazil were analyzed for MNV infection. Using primers targeting a conserved region of the viral capsid, MNV was detected by RT-PCR in 137 of 359 mice from all 22 facilities. Nucleotide sequencing and phylogenetic analysis of the capsid region from the viral genome showed identity ranging from 87% to 99% when compared to reported MNV sequences. In addition, RAW264.7 cells inoculated with a mouse fecal suspension displayed cytopathic effect after the fifth passage. This study represents the first report of MNV in mouse colonies in Brazilian laboratory animal facilities, emphasizing the relevance of a health surveillance program in such environments
ARTIGO - Inglês
Assunto: Nanopartículas
The increase of Zika virus (ZIKV) infections in Brazil in the last two years leaves a prophylactic measures on alert for this new and emerging...
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The increase of Zika virus (ZIKV) infections in Brazil in the last two years leaves a prophylactic measures on alert for this new and emerging pathogen. Concerning of our positive experience, we developed a new prototype using Neisseria meningitidis outer membrane vesicles (OMV) on ZIKV cell growth in a fusion of OMV in the envelope of virus particles. The fusion of nanoparticles resulting from outer membrane vesicles of N. meningitidis with infected C6/36 cells line were analyzed by Nano tracking analysis (NTA), zeta potential, differential light scattering (DLS), scan and scanning transmission eletronic microscopy (SEM and STEM) and high resolution mass spectometry (HRMS) for nanostructure characterization. Also, the vaccination effects were viewed by immune response in mice protocols immunization (ELISA and inflammatory chemokines) confirmed by Zika virus soroneutralization test. The results of immunizations in mice showed that antibody production had a titer greater than 1:160 as compared to unvaccinated mice. The immune response of the adjuvant and non-adjuvant formulation activated the cellular immune response TH1 and TH2. In addition, the serum neutralization was able to prevent infection of virus particles in the glial tumor cell model (M059J). This research shows efficient strategies without recombinant technology or DNA vaccines
ARTIGO - Inglês
Assunto: Hipertensão
This Study evaluated the effects of BAY 41-2272 (BAY), a specific activator of sGC NO-independent action on changes of mean arterial blood pressure,...
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This Study evaluated the effects of BAY 41-2272 (BAY), a specific activator of sGC NO-independent action on changes of mean arterial blood pressure, heart and left ventricle weight indexes, cardiomyocyte hypertrophy (Vv) and fibrosis area induced by chronic N-nitro-L-arginine methyl ester (L-NAME) treatment in rats. The animals were divided into (a) control group, (b) L-NAME group, (c) L-NAME+BAY group, and (d) BAY group. Eight weeks of L-NAME treatment caused a significant increase in mean arterial blood pressure when compared with untreated rats (173 +/- 11.1 and 109 +/- 5.0 mm Hg, respectively; P < 0.01). L-NAME+BAY cotreatment abolished the L-NAME-induced hypertension (112 +/- 5.1 mm Hg; P < 0.01). Significant increases in heart and left ventricle weight indexes and in Vv were observed in the L-NAME-treated animals compared with control group, and concomitant treatment with BAY significantly attenuated this hypertrophic effect. Treatment with L-NAME presented several areas of repairing fibrosis in left ventricles, and this effect was also abolished by BAY cotreatment. Our results demonstrate that BAY 41-2272 inhibits hypertension and prevents heart abnormalities (cardiac hypertrophy and increased fibrosis areas) induced by NO synthase inhibition
DOI: https://doi.org/fjc.0000210071.54828.9b
Texto completo: https://insights.ovid.com/pubmed?pmid=16633081
ARTIGO - Inglês
Assunto: Células-tronco embrionárias
Nonhuman primate (NHP) induced pluripotent stem cells (iPSCs) offer the opportunity to investigate the safety, feasibility, and efficacy of proposed...
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Nonhuman primate (NHP) induced pluripotent stem cells (iPSCs) offer the opportunity to investigate the safety, feasibility, and efficacy of proposed iPSC-derived cellular delivery in clinically relevant in vivo models. However, there is need for stable, robust, and safe labeling methods for NHP iPSCs and their differentiated lineages to study survival, proliferation, tissue integration, and biodistribution following transplantation. Here we investigate the utility of the adenoassociated virus integration site 1 (AAVS1) as a safe harbor for the addition of transgenes in our rhesus macaque iPSC (RhiPSC) model. A clinically relevant marker gene, human truncated CD19 (h Delta CD19), or GFP was inserted into the AAVS1 site in RhiPSCs using the CRISPR/Cas9 system. Genetically modified RhiPSCs maintained normal karyotype and pluripotency, and these clones were able to further differentiate into all three germ layers in vitro and in vivo. In contrast to transgene delivery using randomly integrating viral vectors, AAVS1 targeting allowed stable transgene expression following differentiation. Off -target mutations were observed in some edited clones, highlighting the importance of careful characterization of these cells prior to downstream applications. Genetically marked RhiPSCs will be useful to further advance clinically relevant models for iPSC-based cell therapies
ARTIGO - Inglês
Assunto: Artigo original
Abstract: Clinically, bone tissue replacements and/or bone repair are challenging. Strategies based on well-defined combinations of osteoconductive...
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Abstract: Clinically, bone tissue replacements and/or bone repair are challenging. Strategies based on well-defined combinations of osteoconductive materials and osteogenic cells are promising to improve bone regeneration but still require improvement. Herein, we combined polycaprolactone (PCL) fibers, carbon nanotubes (CNT), and hydroxyapatite (nHap) nanoparticles to develop the next generation of bone regeneration material. Fibers formed by rotary jet spinning (RJS) instead of traditional electrospinning (ES) with embedded bone marrow mesenchymal stem cells (BMMSCs) showed the best outcomes to repair rat calvarial defects after 6 weeks. To understand this, it was observed that different morphologies were formed depending on the manufacturing method used. RJS fibers presented a particular topography with rough fibers, which allowed for better cellular growth and cell spreading in vitro around and into a three-dimensional (3D) mesh, while fibers made by ES were more smooth and cellular growth was only measured on the 3D mesh surface. The fibers with incorporated nHap/CNT nanoparticles enhanced in vitro cell performance as indicated by more cellular proliferation, alkaline phosphatase activity, proliferation, and deposition of calcium. Greater bone neoformation occurred by combining three characteristics: the presence of nHap and CNT nanoparticles, the topography of the RJS fibers, and the addition of BMMSCs. RJS fibers with nanoparticles and seeded with BMMSCs showed 10?136 mm3 of bone neoformation, meaning a 10-fold increase compared to using RJS only and BMMSCs (0.853 mm3) and a 5-fold increase from using ES only (2054 mm3) after 6 weeks of implantation. Conversely, none of these approaches used individually showed any significant difference for in vivo bone neoformation, suggesting that their combination is essential for optimizing bone formation. In summary, our work generated a potential material composed of well-defined combinations of suitable scaffolds seeded with BMMSCs for enhancing numerous orthopedic tissue engineering applications
DOI: https://doi.org/10.1021/acsabm.1c00365
Texto completo: https://pubs.acs.org/doi/10.1021/acsabm.1c00365
ARTIGO - Inglês
Assunto: Artigo original
Abstract: Three of the main requirements that remain major challenges in tissue engineering of the knee meniscus are to engineer scaffolds with...
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Abstract: Three of the main requirements that remain major challenges in tissue engineering of the knee meniscus are to engineer scaffolds with compatible anatomical shape, good mechanical properties, and microstructure able to mimic the architecture of the extracellular matrix (ECM). In this context, we presented a new biofabrication strategy to develop a three-dimensional (3D) meniscus-regenerative scaffold with custom-made macroscopic size and microarchitecture bioinspired by the organization of structural fibers of native tissue ECM Methods: The concept was based on the combination of bioprinted cell-laden hydrogel (type 1 collagen) reinforced by multilayers of biomimetically aligned electrospun nanofibrous mats (polycaprolactone/carbon nanotubes, PCL/CNT), using a patient-specific 3D digital meniscus model reconstructed from MRI data by free and open-source software. Results: The results showed that the incorporation of aligned nanofibers sheets between the hydrogel layers enhanced the scaffold's structural integrity and shape fidelity compared to the nanofiber-free collagen hydrogel. Furthermore, mechanical compression tests demonstrated that the presence of nanofiber layers significantly improved the mechanical properties of the bioprinted construct. Importantly, the introduction of PCL/CNT nanofibrous mats between the layers of the bioprinted collagen hydrogel did not negatively affect cell viability, in which mesenchymal stem cells remained viable even after 7 days of culture within the scaffold. Conclusion: Overall, these findings evidence that this bioengineering approach offers a promising strategy for fabricating biomimetic meniscus scaffolds for tissue engineering
ARTIGO - Inglês
Assunto: Trypanosoma cruzi
In recent decades some outbreaks of food-borne acute Chagas disease (ACD) in humans were identified by clinical and epidemiological characterization...
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In recent decades some outbreaks of food-borne acute Chagas disease (ACD) in humans were identified by clinical and epidemiological characterization after association through the ingestion of açaí pulp probably contaminated with Trypanosoma cruzi. Whereas Belém and Abaetetuba stood out as important risk regions for disease transmission, the importance of Rhodnius pictipes, and Philander opossum for the biological cycle of T. cruzi, and data from agribusiness market of açaí to study T. cruzi from vector and reservoir of the Brazilian Amazon region is critical for this context. Thus, the purpose of this study was to verify the infective capacity and the virulence of T. cruzi in açaí pulp from vector and reservoir at Pará State experimentally. Methods: 10 5 T. cruzi I in in natura açaí pulp from Belém at Pará State, at room temperature, after forced sieving, by intraperitoneal, gavage or oral route of inoculation in B6.129S7Rag1 −/−tmMom /J Unib allowed food-borne ACD analysis using common light microscopy. Principal findings: T. cruzi in in natura açaí pulp from R. pictipes (Val-De-Cans Forest, Belém, and Ajuaí River, Abaetetuba, Pará), and P. opossum (Combu Island, Belém, Pará) caused ACD and death between 17 and 52 days after experimental infections in murine immunodeficient hosts. Conclusions: T. cruzi from different sources and locations at Pará State in in natura açaí pulp retained its infective capacity and virulence, and can cause new outbreaks of ACD by oral transmission. Additionally, quality basic education will facilitate efficient hygiene practices throughout the açaí productive chain can eradicate food-borne ACD in the coming decades