RESUMO
The transcription factor Nkx2.3 regulates the vascular specification of Peyer patches in mice through determining endothelial addressin preference and may function as a susceptibility factor in inflammatory bowel diseases in humans. We wished to analyze the role of Nkx2.3 in colonic solitary intestinal lymphoid tissue composition and in colitis pathogenesis. We studied the colonic solitary intestinal lymphoid tissue of Nkx2.3-deficient mice with immunofluorescence and flow cytometry. Colitis was induced in mice using 2.5% dextran sodium sulfate, and severity was assessed with histology, flow cytometry, and quantitative PCR. We found that the lack of Nkx2.3 impairs maturation of isolated lymphoid follicles and attenuates dextran sodium sulfate-induced colitis independent of endothelial absence of mucosal addressin cell-adhesion molecule-1 (MAdCAM-1), which was also coupled with enhanced colonic epithelial regeneration. Although we observed increased numbers of group 3 innate lymphoid cells and Th17 cells and enhanced transcription of IL-22, Ab-mediated neutralization of IL-22 did not abolish the protection from colitis in Nkx2.3-deficient mice. Nkx2.3-/- hematopoietic cells could not rescue wild-type mice from colitis. Using LacZ-Nkx2.3 reporter mice, we found that Nkx2.3 expression was restricted to VAP-1+ myofibroblast-like pericryptal cells. These results hint at a previously unknown stromal role of Nkx2.3 as driver of colitis and indicate that Nkx2.3+ stromal cells play a role in epithelial cell homeostasis.
Assuntos
Colite/imunologia , Proteínas de Homeodomínio/imunologia , Nódulos Linfáticos Agregados/imunologia , Fatores de Transcrição/imunologia , Animais , Colite/metabolismo , Interleucinas/metabolismo , Linfócitos/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Nódulos Linfáticos Agregados/metabolismo , Células Estromais/imunologia , Fatores de Transcrição/deficiência , Interleucina 22RESUMO
Regeneration of body parts and their interaction with the immune response is a poorly understood aspect of earthworm biology. Consequently, we aimed to study the mechanisms of innate immunity during regeneration in Eisenia andrei earthworms. In the course of anterior and posterior regeneration, we documented the kinetical aspects of segment restoration by histochemistry. Cell proliferation peaked at two weeks and remitted by four weeks in regenerating earthworms. Apoptotic cells were present throughout the cell renewal period. Distinct immune cell (e.g., coelomocyte) subsets were accumulated in the newly-formed blastema in the close proximity of the apoptotic area. Regenerating earthworms have decreased pattern recognition receptors (PRRs) (e.g., TLR, except for scavenger receptor) and antimicrobial peptides (AMPs) (e.g., lysenin) mRNA patterns compared to intact earthworms. In contrast, at the protein level, mirroring regulation of lysenins became evident. Experimental coelomocyte depletion caused significantly impaired cell divisions and blastema formation during anterior and posterior regeneration. These obtained novel data allow us to gain insight into the intricate interactions of regeneration and invertebrate innate immunity.
Assuntos
Imunidade Inata , Oligoquetos/fisiologia , Regeneração , Ferimentos e Lesões , Animais , Apoptose , Proliferação de Células , Regulação da Expressão Gênica , Oligoquetos/genética , Oligoquetos/imunologia , Toxinas BiológicasRESUMO
BACKGROUND: Based on the phenotypic and functional characteristics unconventional T-lymphocytes such as invariant natural killer T (iNKT) cells and mucosal-associated invariant T (MAIT) cells link the innate and adaptive immune responses. Up to now data are scarce about their involvement in pulmonary disorders including chronic obstructive pulmonary disease (COPD). This study explores simultaneously the frequencies of iNKT and MAIT cells in the peripheral blood and sputum of stable and exacerbating COPD patients. METHODS: By means of multicolor flow cytometry frequencies of total iNKT and MAIT cells and their subsets were enumerated in peripheral blood and sputum samples of healthy controls, and COPD patients. In addition, gene expression of TCR for iNKT, MAIT cells, and CD1d, MR1 were assessed by qPCR in the study cohorts. RESULTS: Percentages of total iNKT and MAIT cells were dramatically dropped in blood, and reduced numbers of iNKT cells were observed in the sputum of COPD patients. Furthermore decreased DN and increased CD4+ iNKT subsets, while increased DN and decreased CD8+ MAIT subpopulations were measured in the blood of COPD patients. Reduced invariant TCR mRNA levels in COPD patients had confirmed these previous findings. The mRNA expression of CD1d and MR1 were increased in stable and exacerbating COPD patients; however both molecules were decreased upon antibiotic and systemic steroid treatments. CONCLUSIONS: Our results support the notion that both invariant T-cell populations are affected in COPD. Further detailed analysis of invariant T cells could shed more light into the complex interactions of these lymphocyte groups in COPD pathogenesis.
Assuntos
Células T Invariantes Associadas à Mucosa/metabolismo , Células T Matadoras Naturais/metabolismo , Doença Pulmonar Obstrutiva Crônica/sangue , Adulto , Feminino , Citometria de Fluxo/métodos , Humanos , Leucócitos Mononucleares/imunologia , Leucócitos Mononucleares/metabolismo , Masculino , Pessoa de Meia-Idade , Células T Invariantes Associadas à Mucosa/imunologia , Células T Matadoras Naturais/imunologia , Doença Pulmonar Obstrutiva Crônica/diagnóstico , Doença Pulmonar Obstrutiva Crônica/imunologia , Escarro/imunologia , Escarro/metabolismoRESUMO
Age-related neurocognitive disorders are common problems in developed societies. Aging not only affects memory processes, but may also disturb attention, vigilance, and other executive functions. In the present study, we aimed to investigate age-related cognitive deficits in rats and associated molecular alterations in the brain. We also aimed to test the effects of the alpha7 nicotinic acetylcholine receptor (nAChR) agonist PHA-543613 on memory as well as on the sustained attention and vigilance of aged rats. Short- and long-term spatial memories of the rats were tested using the Morris water maze (MWM) task. To measure attention and vigilance, we designed a rat version of the psychomotor vigilance task (PVT) that is frequently used in human clinical examinations. At the end of the behavioral experiments, mRNA and protein expression of alpha7 nAChRs, cytokines, and brain-derived neurotrophic factor (BDNF) were quantitatively measured in the hippocampus, frontal cortex, striatum, and cerebellum. Aged rats showed marked cognitive deficits in both the MWM and the PVT. The deficit was accompanied by increased IL-1beta and TNFalpha mRNA expression and decreased BDNF protein expression in the hippocampus. PHA-543613 significantly improved the reaction time of aged rats in the PVT, especially for unexpectedly appearing stimuli, while only slightly (non-significantly) alleviating spatial memory deficits in the MWM. These results indicate that targeting alpha7 nAChRs may be an effective strategy for the amelioration of attention and vigilance deficits in age-related neurocognitive disorders.
Assuntos
Fator Neurotrófico Derivado do Encéfalo , Receptor Nicotínico de Acetilcolina alfa7 , Humanos , Ratos , Animais , Receptor Nicotínico de Acetilcolina alfa7/genética , Receptor Nicotínico de Acetilcolina alfa7/metabolismo , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Citocinas/metabolismo , Encéfalo/metabolismo , RNA MensageiroRESUMO
In addition to their role, immunoglobulins can be used in animal and human diagnostic (immunoassay-based) measurements, prophylaxis and (immuno)therapy. For these purposes, today's "alternative" that is advantageous from an animal ethical point of view is the bird immunoglobulin Y isolated from egg yolk. Its development and production are cost-effective, the complexity is low, and due to its advantageous properties, it can be used in assays or even more so in medical therapies (primarily passive immunization). It is widely used (against pathogens or their toxins) in treatments of intestinal or metabolic diseases and inflammations. Its application in human diagnostics is limited, some markers are measured using immunoglobulin Y as assay component. In this study, a possible application, which is less common today, is presented. The problem of environmental impacts is becoming significant. Due to human activities, industrialization, environmental changes increase the appearance of natural environmental pollutants, including the effects of mycotoxins produced by molds locally and/or globally, which (mainly through nutrition) affect humans. Such agents often appear together, several mycotoxins affect the individual. As a result of their persistence, mycotoxins absorbed in the intestinal tract and accumulated in organs, can already reach levels that can cause physiological and/or behavioral effects. Although the examination of sources (contaminated foods) is regulated by law, the extent of accumulation has not been or cannot be examined and is often insufficiently taken into account. Due to the nature of the technique, the anti-mycotoxin avian immunoglobulin Y could be used both for detection of (deposited) mycotoxin(s) and/or even for immunotherapy (e.g., mycotoxin neutralization). Demonstrating the endocrine-disrupting mycotoxins using the example of zearalenone (with an explanation of its reproductive and immunological effects), we present generation of zearalenone (and mycotoxin-specific) avian immunoglobulin developments, advocate its use in human detection, urging the development of measurements that are suitable for detecting (multiple) accumulation. Orv Hetil. 2023; 164(39): 1527-1536.
Assuntos
Micotoxinas , Zearalenona , Animais , Humanos , Micotoxinas/análise , Aves Domésticas , Contaminação de Alimentos/análise , Contaminação de Alimentos/prevenção & controle , ImunoglobulinasRESUMO
Mycotoxins are bioaccumulative contaminants impacting animals and humans. The simultaneous detection of frequent active exposures and accumulated mycotoxin level (s) in exposed organisms would be the most ideal to enable appropriate actions. However, few methods are available for the purpose, and there is a demand for dedicated, sensitive, reliable, and practical assays. To demonstrate the issue, mice were exposed to a relevant agent Ochratoxin A (OTA), and accumulated OTA was measured by fine-tuned commercial assays. Quantitative high-performance liquid chromatography with fluorescence detection, enzyme-linked immunosorbent assay, and flow cytometry assays have been developed/modified using reagents available as commercial products when appropriate. Assays were performed on excised samples, and results were compared. Accumulated OTA could be detected and quantified; positive correlations (between applied doses of exposure and accumulated OTA levels and the results from assays) were found. Dedicated assays could be developed, which provided comparable results. The presence and accumulation of OTA following even a short exposure could be quantitatively detected. The assays performed similarly, but HPLC had the greatest sensitivity. Blood contained higher levels of OTA than liver and kidney. We demonstrate that specific but flexible and practical assays should be used for specific/local purposes, to measure the exposure itself and accumulation in blood or organs.
Assuntos
Líquidos Corporais , Micotoxinas , Ocratoxinas , Animais , Líquidos Corporais/química , Cromatografia Líquida de Alta Pressão/métodos , Contaminação de Alimentos/análise , Humanos , Camundongos , Micotoxinas/análise , Ocratoxinas/análiseRESUMO
Porcine sapeloviruses, teschoviruses of family Picornaviridae and type 3 porcine astroviruses of family Astroviridae are (re-)emerging enteric pathogens that could be associated with severe, disseminated infections in swine, affecting multiple organs including the central nervous system (CNS). Furthermore, small-scale pioneer studies indicate the presence of these viruses in porcine nasal samples to various extents. The laboratory diagnostics are predominantly based on the detection of the viral RNA from faecal and tissue samples using different nucleic-acid-based techniques such as RT-qPCR. In this study, a novel highly sensitive one-step triplex RT-qPCR assay was introduced which can detect all known types of neurotropic sapelo-, tescho- and type 3 astroviruses in multiple types of samples of swine. The assay was evaluated using in vitro synthesized RNA standards and a total of 142 archived RNA samples including known sapelo-, tescho- and type 3 astrovirus positive and negative CNS, enteric and nasal specimens. The results of a large-scale epidemiological investigation of these viruses on n = 473 nasal swab samples from n = 28 industrial-type swine farms in Hungary indicate that all three neurotropic viruses, especially type 3 astroviruses, are widespread and endemically present on most of the investigated farms.
Assuntos
Infecções por Astroviridae , Astroviridae , Picornaviridae , Doenças dos Suínos , Teschovirus , Animais , Astroviridae/genética , Infecções por Astroviridae/diagnóstico , Infecções por Astroviridae/veterinária , Fezes , Mamastrovirus , Filogenia , Picornaviridae/genética , RNA Viral/genética , Suínos , Doenças dos Suínos/diagnóstico , Teschovirus/genéticaRESUMO
Invertebrates, including earthworms, are applied to study the evolutionarily conserved cellular immune processes. Earthworm immunocytes (so-called coelomocytes) are functionally similar to vertebrate myeloid cells and form the first line of defense against invading pathogens. Hereby, we compared the engulfment mechanisms of THP-1 human monocytic cells, differentiated THP-1 (macrophage-like) cells, and Eisenia andrei coelomocytes towards Escherichia coli and Staphylococcus aureus bacteria applying various endocytosis inhibitors [amantadine, 5-(N-ethyl-N-isopropyl) amiloride, colchicine, cytochalasin B, cytochalasin D, methyl-ß-cyclodextrin, and nystatin]. Subsequently, we investigated the messenger RNA (mRNA) expressions of immune receptor-related molecules (TLR, MyD88, BPI) and the colocalization of lysosomes with engulfed bacteria following uptake inhibition in every cell type. Actin depolymerization by cytochalasin B and D has strongly inhibited the endocytosis of both bacterial strains in the studied cell types, suggesting the conserved role of actin-dependent phagocytosis. Decreased numbers of colocalized lysosomes/bacteria supported these findings. In THP-1 cells TLR expression was increased upon cytochalasin D pretreatment, while this inhibitor caused a dropped LBP/BPI expression in differentiated THP-1 cells and coelomocytes. The obtained data reveal further insights into the evolution of phagocytes in eukaryotes. Earthworm and human phagocytes possess analogous mechanisms for bacterial internalization.
Assuntos
Infecções por Escherichia coli/imunologia , Escherichia coli/fisiologia , Macrófagos/fisiologia , Infecções Estafilocócicas/imunologia , Staphylococcus aureus/fisiologia , Amantadina/farmacologia , Animais , Evolução Biológica , Diferenciação Celular , Endocitose , Humanos , Imunidade Celular , Imunidade Inata , Oligoquetos , Células THP-1RESUMO
PURPOSE: Decreased expression of TLR homolog CD180 in peripheral blood B cells and its potential role in antibody production have been described in autoimmune diseases. Effectiveness of anti-CD20 therapy in neuromyelitis optica spectrum disorder (NMOSD) and multiple sclerosis (MS) strengthens the role of B cells in the pathogenesis. Therefore, we aimed to investigate the CD180 expression of peripheral blood B cell subsets in NMOSD and MS patients and analyze the levels of natural anti-citrate synthase (CS) IgG autoantibodies and IgG antibodies induced by bacterial infections reported to play a role in the pathogenesis of NMOSD or MS. METHODS: We analyzed the distribution and CD180 expression of peripheral blood B cell subsets, defined by CD19/CD27/IgD staining, and measured anti-CS IgM/G natural autoantibody and antibacterial IgG serum levels in NMOSD, RRMS, and healthy controls (HC). RESULTS: We found decreased naïve and increased memory B cells in NMOSD compared to MS. Among the investigated four B cell subsets, CD180 expression was exclusively decreased in CD19+CD27+IgD+ nonswitched (NS) memory B cells in both NMOSD and MS compared to HC. Furthermore, the anti-CS IgM natural autoantibody serum level was lower in both NMOSD and MS. In addition, we found a tendency of higher anti-CS IgG natural autoantibody levels only in anti-Chlamydia IgG antibody-positive NMOSD and MS patients. CONCLUSIONS: Our results suggest that reduced CD180 expression of NS B cells could contribute to the deficient natural IgM autoantibody production in NMOSD and MS, whereas natural IgG autoantibody levels show an association with antibacterial antibodies.
Assuntos
Antígenos CD/metabolismo , Autoanticorpos/imunologia , Subpopulações de Linfócitos B/imunologia , Citrato (si)-Sintase/imunologia , Células B de Memória/imunologia , Esclerose Múltipla/imunologia , Neuromielite Óptica/imunologia , Adulto , Idoso , Antígenos CD/genética , Células Cultivadas , Regulação para Baixo , Feminino , Humanos , Imunoglobulina M/metabolismo , Masculino , Pessoa de Meia-Idade , Receptores Toll-Like/genética , Adulto JovemRESUMO
Serological testing is a tool to predict protection against later infection. This potential heavily relies on antibody levels showing acceptable agreement with gold standard virus neutralization tests. The aim of our study was to investigate diagnostic value of the available serological tests in terms of predicting virus neutralizing activity of serum samples drawn 5-7 weeks after onset of symptoms from 101 donors with a history of COVID-19. Immune responses against Receptor Binding Domain (RBD), Spike1 and 2 proteins and Nucleocapsid antigens were measured by various ELISA tests. Neutralizing antibody activity in serum samples was assessed by a cell-based virus neutralization test. Spearman correlation coefficients between serological and neutralization results ranged from 0.41 to 0.91 indicating moderate to strong correlation between ELISA test results and virus neutralization. The sensitivity and specificity of ELISA tests in the prediction of neutralization were 35-100% and 35-90% respectively. No clear cut off levels can be established that would reliably indicate neutralization activity. For some tests, however, a value below which the sample is not expected to neutralize can be established. Our data suggests that several of the ELISA kits tested may be suitable for epidemiological surveys 1-2 months after the infection, estimating whether a person may have recently exposed to the virus. Sensitivities considerably superseding specificity at the cut-off values proposed by the manufacturers suggest greater potential in the identification of insufficient antibody responses than in confirming protection. Nevertheless, the former might be important in assessing response to vaccination and characterizing therapeutic plasma preparations.
RESUMO
Earthworms and leeches are sentinel animals that represent the annelid phylum within terrestrial and freshwater ecosystems, respectively. One early stress signal in these organisms is related to innate immunity, but how nanomaterials affect it is poorly characterized. In this survey, we compare the latest literature on earthworm and leeches with examples of their molecular/cellular responses to inorganic (silver nanoparticles) and organic (carbon nanotubes) nanomaterials. A special focus is placed on the role of annelid immunocytes in the evolutionarily conserved antioxidant and immune mechanisms and protein corona formation and probable endocytosis pathways involved in nanomaterial uptake. Our summary helps to realize why these environmental sentinels are beneficial to study the potential detrimental effects of nanomaterials.
RESUMO
Lumbricin and its orthologue antimicrobial peptides were typically isolated from annelids. In this report, mRNA for lumbricin and -serendipitously- a novel lumbricin-related mRNA sequence were identified in Eisenia andrei earthworms. The determined mRNA sequences of E. andrei lumbricin and lumbricin-related peptide consist of 477 and 575 nucleotides. The precursors of proline-rich E. andrei lumbricin and the related peptide contain 63 and 59 amino acids, respectively. Phylogenetic analysis indicated close relationship with other annelid lumbricins. Highest expression of both mRNAs appeared in the proximal part of the intestine (pharynx, gizzard), while other tested organs had moderate (body wall, midgut, ovary, metanephridium, seminal vesicles, ventral nerve cord) or low (coelomocytes) levels. During ontogenesis their expression revealed continuous increase in embryos. Following 48â¯h of in vivo Gram-positive bacteria challenge both mRNAs were significantly elevated in coelomocytes, while Gram-negative bacteria or zymosan stimulation had no detectable effects.
Assuntos
Infecções por Bactérias Gram-Negativas/imunologia , Infecções por Bactérias Gram-Positivas/imunologia , Proteínas de Helminto/genética , Intestinos/fisiologia , Oligoquetos/imunologia , Peptídeos/genética , Animais , Anti-Infecciosos/metabolismo , Clonagem Molecular , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Proteínas de Helminto/metabolismo , Imunidade Inata , Oligoquetos/genética , Oligoquetos/microbiologia , Peptídeos/metabolismo , Filogenia , TranscriptomaRESUMO
OBJECTIVE: Autoantibody detection is crucial for the early diagnosis of autoimmune encephalitis (AIE) since prompt therapy can determine the disease outcome. Here, we report a single-center 6-year retrospective study of autoantibody testing in AIE in the Hungarian population. METHODS: Serum and/or cerebrospinal fluid (CSF) autoantibody tests were performed using cell-based indirect immunofluorescence assay for AIE diagnosis. Samples were provided by neurology clinics as part of a nationwide program. Test results were analyzed for samples received during the period from 2012 to 2018. RESULTS: We tested 1,247 samples from 1,034 patients with suspected AIE. Autoantibodies were present in 60 patients (5.8% of total). The distribution of patients with different autoantibodies by age and sex was as follows: NMDAR (70%), mostly in young females, LGI1 (15%) in middle-aged males, GABAB R (12%) in elderly males, and Caspr2 (7%) in males. Long-term follow-up was conducted in 30 patients with repeated test requests, of which 17 remained positive, and 13 switched to negative. CONCLUSION: We report the most comprehensive clinical laboratory study of autoantibody testing in AIE in the Hungarian population. Our results show that the frequency of different autoantibody types in AIE corresponds to the data described in the literature.
Assuntos
Autoanticorpos/sangue , Encefalite/diagnóstico , Doença de Hashimoto/diagnóstico , Adulto , Idoso , Encefalite/sangue , Encefalite/imunologia , Feminino , Doença de Hashimoto/sangue , Doença de Hashimoto/imunologia , Humanos , Hungria , Masculino , Pessoa de Meia-Idade , Receptores de N-Metil-D-Aspartato , Estudos RetrospectivosRESUMO
Because of measles outbreaks there is a need for continuous monitoring of immunological protection against infection at population level. For such monitoring to be feasible, a cost-effective, reliable and high-throughput assay is necessary. Herein we describe an ELISA protocol for assessment of anti-measles antibody levels in human serum samples that fulfills the above criteria and is easily adaptable by various laboratories. A serum bank of anonymous patient sera was established (Nâ¯>â¯3000 samples). Sera were grouped based on measles immunization schedules and/or changes in vaccine components since the introduction of the first measles vaccine in Hungary in 1969. Newly designed ELISA was performed by using Siemens BEP 2000 Advance System and data were confirmed using commercially available kits. Our indirect ELISA was compared to indirect immunfluoresence and to anti-measles nucleocapsid (N) monoclonal antibody-based sandwich ELISA. The results obtained are in high agreement with the confirmatory methods, and reflect measles vaccination history in Hungary ranging from pre-vaccination era, through the initial period of measles vaccination, to present. Based on measurement of 1985 sera, the highest ratio of low/questionable antibody level samples was detected in cluster '1978-1987' (~25.4%), followed by cluster '1969-1977' (~15.4%).Our assay is suitable for assessment of anti-measles immunity in a large cohort of subjects. The assay is cost-effective, allows high-throughput screening and has superior signal-to-noise ratio. This assay can serve as a first step in assessment of the effectiveness of all three components of the MMR vaccine.
Assuntos
Anticorpos Antivirais/sangue , Ensaio de Imunoadsorção Enzimática/métodos , Imunogenicidade da Vacina , Imunoglobulina G/sangue , Vírus do Sarampo/imunologia , Vacina contra Sarampo-Caxumba-Rubéola/imunologia , Sarampo/prevenção & controle , Biomarcadores/sangue , Técnica Indireta de Fluorescência para Anticorpo , Humanos , Hungria , Imunidade Coletiva , Limite de Detecção , Sarampo/sangue , Sarampo/imunologia , Sarampo/virologia , Vacina contra Sarampo-Caxumba-Rubéola/administração & dosagem , Valor Preditivo dos Testes , VacinaçãoRESUMO
Traumatic brain injury (TBI) induces cerebrovascular oxidative stress, which is associated with neurovascular uncoupling, autoregulatory dysfunction, and persisting cognitive decline in both pre-clinical models and patients. However, single mild TBI (mTBI), the most frequent form of brain trauma, increases cerebral generation of reactive oxygen species (ROS) only transiently. We hypothesized that comorbid conditions might exacerbate long-term ROS generation in cerebral arteries after mTBI. Because hypertension is the most important cerebrovascular risk factor in populations prone to mild brain trauma, we induced mTBI in normotensive and spontaneously hypertensive rats (SHR) and assessed changes in cytoplasmic and mitochondrial superoxide (O2-) production by confocal microscopy in isolated middle cerebral arteries (MCA) 2 weeks after mTBI using dihydroethidine (DHE) and the mitochondria-targeted redox-sensitive fluorescent indicator dye MitoSox. We found that mTBI induced a significant increase in long-term cytoplasmic and mitochondrial O2- production in MCAs of SHRs and increased expression of the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase subunit Nox4, which were reversed to the normal level by treating the animals with the cell-permeable, mitochondria-targeted antioxidant peptide SS-31 (5.7 mg kg-1 day-1, i.p.). Persistent mTBI-induced oxidative stress in MCAs of SHRs was significantly decreased by inhibiting vascular NADPH oxidase (apocyinin). We propose that hypertension- and mTBI-induced cerebrovascular oxidative stress likely lead to persistent dysregulation of cerebral blood flow (CBF) and cognitive dysfunction, which might be reversed by SS-31 treatment.
Assuntos
Concussão Encefálica/metabolismo , Sistemas de Liberação de Medicamentos/métodos , Hipertensão/metabolismo , Mitocôndrias/metabolismo , Oligopeptídeos/administração & dosagem , Estresse Oxidativo/fisiologia , Animais , Antioxidantes/administração & dosagem , Concussão Encefálica/complicações , Concussão Encefálica/tratamento farmacológico , Hipertensão/complicações , Hipertensão/tratamento farmacológico , Masculino , Mitocôndrias/efeitos dos fármacos , Fármacos Neuroprotetores/administração & dosagem , Estresse Oxidativo/efeitos dos fármacos , Ratos , Ratos Endogâmicos SHR , Ratos WistarRESUMO
Selenium (Se) is an essential element for humans, animals, and certain lower plants, but can be toxic at high concentration. Even though Se is potentially toxic, little information is available about the effects of Se on soil animals. The aim of this study was to assess the impact of different concentrations of two Se forms, selenate and selenite, on earthworm Eisenia andrei. In order to obtain comprehensive overview on the Se effects, different parameters were measured. Namely, acute toxicity, apoptosis, efflux pump activity, different enzymatic and non-enzymatic biomarkers (acetylcholinesterase, carboxylesterase, glutathione S-transferase, catalase, glutathione reductase and superoxide dismutase activities, lipid peroxidation level and GSH/GSSG ratio) and expression of genes involved in oxidative and immune response have been investigated. Additionally, measurement of metallothioneins concentration and concentration of Se in exposed earthworms has been also performed. The assessment of acute toxicity showed a greater sensitivity of E. andrei to selenite exposure, whereas Se concentration measurements in earthworms showed higher accumulation of selenate form. Both Se forms caused inhibition of the efflux pump activity. Decrease in superoxide dismutase activity and increase in lipid peroxidation and glutathione reductase activity indicate that Se has a significant impact on the oxidative status of earthworms. Selenate exposure caused an apoptotic-like cell death in the coelomocytes of exposed earthworms, whereas decreased mRNA levels of stress-related genes and antimicrobial factors were observed upon the exposure to selenite. The obtained data give insight into the effects of two most common forms of Se in soil on the earthworm E. andrei.
Assuntos
Oligoquetos/efeitos dos fármacos , Ácido Selênico/toxicidade , Ácido Selenioso/toxicidade , Poluentes do Solo/toxicidade , Animais , Peroxidação de Lipídeos/efeitos dos fármacos , Oligoquetos/enzimologia , Oligoquetos/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Solo/químicaRESUMO
Flow cytometry is a common approach to study invertebrate immune cells including earthworm coelomocytes. However, the link between light-scatter- and microscopy-based phenotyping remains obscured. Here we show, by means of light scatter-based cell sorting, both subpopulations (amoebocytes and eleocytes) can be physically isolated with good sort efficiency and purity confirmed by downstream morphological and cytochemical applications. Immunocytochemical analysis using anti-EFCC monoclonal antibodies combined with phalloidin staining has revealed antigenically distinct, sorted subsets. Screening of lectin binding capacity indicated wheat germ agglutinin (WGA) as the strongest reactor to amoebocytes. This is further evidenced by WGA inhibition assays that suggest high abundance of N-acetyl-d-glucosamine in amoebocytes. Post-sort phagocytosis assays confirmed the functional differences between amoebocytes and eleocytes, with the former being in favor of bacterial engulfment. This study has proved successful in linking flow cytometry and microscopy analysis and provides further experimental evidence of phenotypic and functional heterogeneity in earthworm coelomocyte subsets.