RESUMO
Meningitis caused by Gram-negative bacteria is a serious public health problem, causing morbidity and mortality in both children and adults. Here, we propose a novel experimental model using Nile tilapia (Oreochromis niloticus) to study neuroinflammation. The fish were infected with Aeromonas hydrophila, and the course of infection was monitored in the peripheral blood. Septicemia was obvious in the blood, while in the brain tissue, infection of the meninges was present. The histopathological examination showed suppurative meningitis, and the cellular immune response in the brain tissue during infection was mediated by microglia. These cells were morphologically characterized and phenotyped by MHC class II markers and CD68. The increased production of TNF-α, IL-1ß and iNOS supported the infiltration of these cells during the neuroinflammatory process. In the proteomic analysis of A. hydrophila isolated from brain tissue, we found chemotactic and transport proteins, proteolytic enzymes and enzymes associated with the dismutation of nitric oxide (NO), as well as motor proteins and those responsible for cell division. After characterizing the most abundant proteins during the course of infection, we investigated the druggability index of these proteins and identified promising peptide sequences as molecular targets that are similar among bacteria. Thus, these findings deepened the understanding of the pathophysiology of meningitis caused by A. hydrophila. Moreover, through the proteomics analysis, important mechanisms and pathways used by the pathogen to subvert the host response were revealed, providing insights for the development of novel antibiotics and vaccines.
Assuntos
Aeromonas hydrophila , Ciclídeos , Doenças dos Peixes , Infecções por Bactérias Gram-Negativas , Proteômica , Fatores de Virulência , Animais , Aeromonas hydrophila/fisiologia , Ciclídeos/imunologia , Doenças dos Peixes/imunologia , Doenças dos Peixes/microbiologia , Infecções por Bactérias Gram-Negativas/veterinária , Infecções por Bactérias Gram-Negativas/imunologia , Fatores de Virulência/imunologia , Meningites Bacterianas/veterinária , Meningites Bacterianas/imunologia , Proteínas de Peixes/imunologiaRESUMO
Atrazine (ATZ) is a herbicide that is frequently present in surface waters and may result in damage to the health of various organisms, including humans. However, most scientific literature reports injuries caused by ATZ at high concentrations, which are not found in the environment. Therefore, the scope of this study was to investigate the impacts of realistic concentrations of ATZ found in surface waters (1, 2, 5, 10, 15 and 20 µg/L) using the bioindicators Allium cepa, Daphnia magna and zebrafish (Danio rerio). ATZ elicited a genotoxic effect in A. cepa, manifested by the induction of chromosomal aberrations, and a mutagenic effect with increased incidence of micronuclei formation, promotion of cell death and reduction in nuclear size revealed by flow cytometry analysis. D. magna exposed to 10, 15 and 20 µg/L of ATZ showed significant reduction in body size after 21 days, delayed first-brood release, decreased egg production and total offspring, as well as swimming behavioral changes. ATZ exposure promoted physiological and developmental alterations in zebrafish embryos, including an increased spontaneous movement rate, which led to premature hatching at all concentrations investigated. Increase in total body length, decrease of the yolk sac area, pericardial edema and higher heart rate were also detected in ATZ-treated zebrafish. In summary, environmentally relevant concentrations of ATZ can induce substantial alterations in the three bioindicators investigated. This study evidences the deleterious effects of ATZ on three aquatic bioindicators employing established and current techniques, and may contribute to elucidate the risks caused by this widely used herbicide even at low concentrations and short-to-medium-term exposure.
Assuntos
Atrazina , Herbicidas , Poluentes Químicos da Água , Animais , Humanos , Atrazina/toxicidade , Peixe-Zebra , Biomarcadores Ambientais , Herbicidas/toxicidade , Poluentes Químicos da Água/toxicidadeRESUMO
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.
Assuntos
COVID-19 , Animais , Humanos , Peixe-Zebra/metabolismo , SARS-CoV-2/metabolismo , Síndrome da Liberação de Citocina , Citocinas/metabolismo , RNA Mensageiro , Proteínas de Membrana , Proteínas MitocondriaisRESUMO
The effect of daily ingestion of polypropylene microplastic on the health of tilapia, Oreochromis niloticus, was evaluated. 60 fish (± 200 g) were placed in 6 aquariums (n = 10, 100 L each), constituting the following treatments: Control (without the addition of polymer), fed with 100 and 500 µg of polypropylene/kg of body weight (b.w.), respectively. After 30 days of feeding, the animals were submitted to blood collection for hemogram and biochemical study and later euthanized for gut microbiological analysis, somatic index of liver, spleen, heart, kidney, stomach, and intestine. In the serum biochemical study, an increase in cholesterol and serum Aspartate Aminotransferase (AST) activity levels was observed in animals treated with 500 µg of polypropylene. Tilapia-fed polypropylene in the diet showed an increase in thrombocyte and total leukocyte counts, marked by a significant increase in the number of circulating lymphocytes. The results of the somatic study revealed a significant increase in the stomach, liver, and heart of tilapia fed with the polymer. Increase in the number of Gram-negative microorganisms and decrease in mesophilic aerobic microorganisms were observed in the gut of fish exposed to the polymer, including a dose-response effect was observed for these analyses. Therefore, tilapias fed daily with diets containing polypropylene for 30 consecutive days showed deleterious effects, resulting in systemic inflammatory disturbs by altering liver functions, leukocyte profile, and organ morphometry, as well as changes in the intestinal microbiota. Such results demonstrate the impairment of fish health, highlighting the need for further studies that evaluate the impact of microplastics on aquatic organisms.
Assuntos
Ciclídeos , Tilápia , Animais , Ciclídeos/fisiologia , Microplásticos , Plásticos , Polipropilenos/toxicidade , Dieta , Ingestão de Alimentos , Ração Animal/análise , Suplementos Nutricionais/análiseRESUMO
Bacillus spp. supplementation as probiotics in cultured fish diets has a long history of safe and effective use. Specifically, B. velezensis show great promise in fine-tuning the European sea bass disease resistance against the pathogenicity caused by several members of the Vibrio family. However, the immunomodulatory mechanisms behind this response remain poorly understood. Here, to examine the inherent immune variations in sea bass, two equal groups were fed for 30 days with a steady diet, with one treatment supplemented with B. velezensis. The serum bactericidal capacity against live cells of Vibrio anguillarum strain 507 and the nitric oxide and lysozyme lytic activities were assayed. At the cellular level, the phagocytic response of peripheral blood leukocytes against inactivated Candida albicans was determined. Moreover, head-kidney (HK) total leukocytes were isolated from previously in vivo treated fish with LPS of V. anguillarum strain 507. Mechanistically, the expression of some essential proinflammatory genes (interleukin-1 (il1b), tumor necrosis factor-alpha (tnfa), and cyclooxygenase 2 (cox2) and the sea bass specific antimicrobial peptide (AMP) dicentracin (dic) expressions were assessed. Surprisingly, the probiotic supplementation significantly increased all humoral lytic and cellular activities assayed in the treated sea bass. In addition, time-dependent differences were observed between the control and probiotic treated groups for all the HK genes markers subjected to the sublethal LPS dose. Although the il1b was the fastest responding gene to a significant level at 48 h post-injection (hpi), all the other genes followed 72 h in the probiotic supplemented group. Finally, an in vivo bacteria challenge against live V. anguillarum was conducted. The probiotic fed fish observed a significantly higher survival. Overall, our results provide clear vertical evidence on the beneficial immune effects of B. velezensis and unveil some fundamental immune mechanisms behind its application as a probiotic agent in intensively cultured European sea bass.
Assuntos
Bacillus , Bass , Doenças dos Peixes , Vibrioses , Animais , Suplementos Nutricionais , Resistência à Doença , Lipopolissacarídeos , Vibrio , Vibrioses/veterináriaRESUMO
To identify activation pathways and effector mechanisms of innate immunity in fish has become relevant for the sanitary management of intensive fish farming. However, little is known about the blocking of cysteinyl leukotrienes receptors (CysLTRs) and their effects in teleost fish. Our study evaluated the anti-inflammatory effect of 250 and 500 µg zafirlukast (antagonist of CysLTRs)/kg b.w., administered orally in the diet, during acute inflammatory reaction induced by Aeromonas hydrophila bacterins in Oreochromis niloticus. 80 tilapia were distributed in 10 aquariums (100L of water each, n = 8) to constitute three treatments: Control (inoculated with A. hydrophila bacterin and untreated); Treated with 250 µg or 500 µg of zafirlukast/kg b.w. and inoculated. To be evaluated in three periods: 6, 24 and 48 h post-inoculation (HPI), totaling nine aquariums. A tenth group was sampled without any stimulus to constitute reference values (Physiological standards). Tilapia treated with zafirlukast demonstrated dose-response effect in the decrease of accumulated inflammatory cells, strongly influenced by granulocytes and macrophages. Zafirlukast treated-tilapia showed decrease in blood leukocyte counts (mainly neutrophils, and monocytes) and reactive oxygen species production. Treatment with zafirlukast resulted in down-regulation of ceruloplasmin, complement 3, alpha2-macroglobulin, transferrin and apolipoprotein A1, as well as up-regulation of haptoglobin. Our study provided convincing results in the pathophysiology of tilapia inflammatory reaction, considering that treatment with zafirlukast, antagonist of cysteinyl leukotriene receptors, resulted in a dose-response effect by suppressing the dynamics between leukocytes in the bloodstream and cell accumulation in the inflamed focus, as well as modulated the leukocyte oxidative burst and the acute phase protein response.
Assuntos
Ciclídeos , Doenças dos Peixes , Infecções por Bactérias Gram-Negativas , alfa 2-Macroglobulinas Associadas à Gravidez , Tilápia , Aeromonas hydrophila/fisiologia , Animais , Anti-Inflamatórios , Apolipoproteína A-I , Vacinas Bacterianas , Ceruloplasmina , Complemento C3 , Feminino , Haptoglobinas , Indóis , Fenilcarbamatos , Gravidez , Espécies Reativas de Oxigênio , Receptores de Leucotrienos/genética , Sulfonamidas , Transferrinas , ÁguaRESUMO
BACKGROUND: Astaxanthin, classified as a xanthophyll, has antioxidant properties about 500 times greater than α-tocopherols and ten times greater than ß-carotenes. Based on the antioxidant activity of this carotenoid, this study aimed to evaluate the shelf-life of tilapia fillets (Oreochromis niloticus) fed with astaxanthin, by determining the microbiological quality (colimetry, counts of mesophilic and psychrotrophic microorganisms), physicochemical analyses (colorimetry, pH, thiobarbituric acid reactive substances (TBARS)) and sensory analysis. RESULTS: Tilapia supplemented with astaxanthin presented a reduction in the counts of microorganisms (mesophiles and psychrotrophics) and lower lipid oxidation index (TBARS), when compared to fillets of control fish. Colorimetric changes of fillet degradation were observed, associated with increased pH during storage, as well as loss of brightness and texture in addition to worsening of appearance and odor. These deteriorating changes were minimized using astaxanthin. CONCLUSION: Our results demonstrate the beneficial performance of astaxanthin in the shelf-life of tilapia fillets stored under refrigeration. Therefore, dietary supplementation with astaxanthin (100 and 200 mg kg-1 of feed) improves the microbiological and physicochemical quality of tilapia fillets during 50 days of shelf-life. © 2022 Society of Chemical Industry.
Assuntos
Tilápia , Animais , Conservação de Alimentos/métodos , Refrigeração , Substâncias Reativas com Ácido Tiobarbitúrico/análise , Xantofilas/análiseRESUMO
In order to understand events and mechanisms present in the pathophysiology of tilapia's chronic inflammation and based on the immunomodulatory activity attributed to cyclophosphamide which is widely used to suppress immune responses in human medicine, the present study investigated the effects of cyclophosphamide (CYP) treatment on the modulation of foreign body inflammatory reaction in Nile tilapia (Oreochromis niloticus) with round glass coverslip implanted in the subcutaneous tissue (9 mm of diameter). Forty tilapia (151 ± 10,2 g) were randomly distributed in 5 aquariums (n = 8) with a capacity of 250 L of water each, to compose two treatments (sampled 3 and 6 days post-implantation): implanted/untreated (control) and implanted/treated with 200 mg of CYP kg-1 of b.w., through i.p. route. A fifth group (n = 8) was sampled without any stimulus (naive) to obtain reference values. CYP-treated tilapia showed decrease in macrophage accumulation, giant cell formation and Langhans cells on the glass coverslip when compared to control fish. The treatment with CYP resulted in decrease of leukocyte and thrombocyte counts. Decrease in alpha-2-macroglobulin, ceruloplasmin, albumin and transferrin levels, as well as increase in haptoglobin, complement C3 and apolipoprotein A1 were observed in tilapias during foreign body inflammation. Blood levels of complement C3, alpha-2-macroglobulin, ceruloplasmin and transferrin were modulated by treatment with CYP. Therefore, the treatment with 200 mg of CYP kg-1 of b.w. in tilapia resulted in an anti-inflammatory effect by suppressing the dynamics between leukocytes in the bloodstream and macrophage accumulation with giant cell formation in the inflamed focus, as well as by modulating APPs during foreign body reaction.
Assuntos
Ciclídeos/imunologia , Ciclofosfamida/farmacologia , Doenças dos Peixes/imunologia , Reação a Corpo Estranho/veterinária , Imunidade Inata , Imunossupressores/farmacologia , Animais , Reação a Corpo Estranho/imunologiaRESUMO
In order to understand the pathophysiology of melanomcrophage centers (MMCs) formation during the tilapia defense response to bacterial infections, the present study evaluated the response, in terms of area, number and pigment constitution, of splenic MMCs of Oreochromis niloticus subjected to intraperitoneal (i.p.) infection with Aeromonas hydrophila and Mycobacterium marinum. Eighty-four fish (396.9⯱â¯21.0â¯g) were randomly distributed into twelve plastic tanks (300â¯L), to constitute three treatments with 28 animals each: control group (inoculated with PBS); Infected with A. hydrophila (1â¯×â¯107 UFC mL-1); Infected with M. marinum (1â¯×â¯106 UFC mL-1). The spleen was collected in seven fish per treatment on the 3rd, 7th, 14th and 21st day post-infection (DPI). The results revealed the participation of MMCs in the defense response of tilapia during bacterial infection by A. hydrophila and M. marinum, since there was an increase in the number and size of these cell aggregates. Variation of pigment accumulation with significant increase of hemosiderin, in infected tilapias by A. hydrophila, bacteria responsible for causing hemolytic anemia in fish was also found. On the other hand, M. marinum-infected tilapia had high amount of melanin in MMCs. In general, mycobacterial infections are notoriously difficult to treat, being characterized as a chronic disease. These findings demonstrate different strategies of fish response during the evolution of these bacterial diseases.
Assuntos
Ciclídeos/imunologia , Doenças dos Peixes/imunologia , Infecções por Bactérias Gram-Negativas/imunologia , Macrófagos/imunologia , Infecções por Mycobacterium não Tuberculosas/veterinária , Baço/imunologia , Aeromonas hydrophila/imunologia , Animais , Ciclídeos/microbiologia , Feminino , Doenças dos Peixes/microbiologia , Hemossiderina/análise , Masculino , Melaninas/análise , Infecções por Mycobacterium não Tuberculosas/imunologiaRESUMO
Recently, chitosan-based nanoparticles with mucoadhesive properties emerged as a strategy for mucosal drug release. This study aimed to characterize the interaction of mucoadhesive system chitosancoated PLGA nanoparticles (NPMA) with fish external mucus. NP suspensions with fluorescent probe were prepared and characterized by size, polydispersity, zeta potential and pH measures. In post-exposure fish were observed an increase in fluorescence imaging over time and it was significantly influenced by NPMA concentration. We also observed the main predominance the fluorescence in the spleen, followed by liver, gill and other tissues. The use of mucoadhesive nanocarriers becomes an alternative for administration of drugs and immunomodulators in immersion systems since the nanosystem can adhere to the mucosal surface of the fish with little residual effect in the water.
Assuntos
Quitosana/administração & dosagem , Portadores de Fármacos/administração & dosagem , Nanopartículas/administração & dosagem , Ácido Poliglicólico/administração & dosagem , Adesividade , Animais , Quitosana/química , Portadores de Fármacos/química , Corantes Fluorescentes/administração & dosagem , Brânquias/metabolismo , Imunomodulação , Fígado/metabolismo , Mucosa/química , Nanopartículas/química , Ácido Poliglicólico/química , Baço/metabolismo , Peixe-ZebraRESUMO
Layered double hydroxides (LDHs) are stimuli-responsive anionic nanoclays. The vast possibilities of using LDHs can lead to their existence in the ecosystem, raising a question of potential ecological concern. However, little is known about the effect of these nanomaterials on freshwater organisms. The present study aimed to assess the ecotoxicological effects of Zinc-Aluminium LDH-nitrate (ZnAl LDH-NO3) in zebrafish (Danio rerio) early life stages. The endpoints measured were mortality, malformations and hatching rate after exposure of D. rerio embryos and larvae to ZnAl LDH-NO3 following the OECD 236 guideline. The behavioral, biochemical (markers of oxidative stress and neurotoxicity), and molecular (at DNA level) alterations were also assessed using sub-lethal concentrations. No observable acute effects were detected up to 415.2 mg LDH/L while the 96 h-LC50 was estimated as 559.9 mg/L. Tested LDH caused malformations in D. rerio embryos, such as pericardial edema, incomplete yolk sac absorption and tail deformities (96 h-EC50 = 172.4 mg/L). During the dark periods, the locomotor behavior in zebrafish larvae was affected upon ZnAl LDH-NO3 exposure. However, no significant biochemical and molecular changes were recorded. The present findings suggest that ZnAl LDH-NO3 can be regarded as a non-toxic nanomaterial towards D. rerio (E/LC50 > > 100 mg/L) although impairment of the locomotion behavior on zebrafish embryos can be expected at concentrations below 100 mg/L.
Assuntos
Embrião não Mamífero , Peixe-Zebra , Animais , Peixe-Zebra/anormalidades , Embrião não Mamífero/anormalidades , Ecossistema , Locomoção , Zinco/farmacologiaRESUMO
While the multifaceted social, economic, and public health impacts associated with the COVID-19 pandemic are known, little is known about its effects on non-target aquatic ecosystems and organisms. Thus, we aimed to evaluate the potential ecotoxicity of SARS-CoV-2 lysate protein (SARS.CoV2/SP02.2020.HIAE.Br) in adult zebrafish (Danio rerio) at predicted environmentally relevant concentrations (0.742 and 2.226 pg/L), by 30 days. Although our data did not show locomotor alterations or anxiety-like or/and anxiolytic-like behavior, we noticed that exposure to SARS-CoV-2 negatively affected habituation memory and social aggregation of animals in response to a potential aquatic predator (Geophagus brasiliensis). An increased frequency of erythrocyte nuclear abnormalities was also observed in animals exposed to SARS-CoV-2. Furthermore, our data suggest that such changes were associated with a redox imbalance [↑ROS (reactive oxygen species), ↑H2O2 (hydrogen peroxide), ↓SOD (superoxide dismutase), and ↓CAT (catalase)], cholinesterasic effect [↑AChE (acetylcholinesterase) activity], as well as the induction of an inflammatory immune response [↑NO (nitric oxide), ↑IFN-γ (interferon-gamma), and ↓IL-10 (interleukin-10)]. For some biomarkers, we noticed that the response of the animals to the treatments was not concentration-dependent. However, principal component analysis (PCA) and the "Integrated Biomarker Response" index (IBRv2) indicated a more prominent ecotoxicity of SARS-CoV-2 at 2.226 pg/L. Therefore, our study advances knowledge about the ecotoxicological potential of SARS-CoV-2 and reinforces the presumption that the COVID-19 pandemic has negative implications beyond its economic, social, and public health impacts.
Assuntos
COVID-19 , Poluentes Químicos da Água , Animais , Humanos , Peixe-Zebra/fisiologia , SARS-CoV-2 , Estresse Oxidativo , Acetilcolinesterase/metabolismo , Ecossistema , Pandemias , Água Doce , Poluentes Químicos da Água/análiseRESUMO
The COVID-19 pandemic has caused unprecedented negative impacts in the modern era, including economic, social, and public health losses. On the other hand, the potential effects that the input of SARS-CoV-2 in the aquatic environment from sewage may represent on non-target organisms are not well known. In addition, it is not yet known whether the association of SARS-CoV-2 with other pollutants, such as microplastics (MPs), may further impact the aquatic biota. Thus, we aimed to evaluate the possible ecotoxicological effects of exposure of male adults Poecilia reticulata, for 15 days, to inactivated SARS-CoV-2 (0.742 pg/L; isolated SARS.CoV2/SP02.2020.HIAE.Br) and polyethylene MP (PE MPs) (7.1 × 104 particles/L), alone and in combination, from multiple biomarkers. Our data suggest that exposure to SARS-CoV-2 induced behavioral changes (in the open field test), nephrotoxic effect (inferred by the increase in creatinine), hepatotoxic effect (inferred by the increase in bilirubin production), imbalance in the homeostasis of Fe, Ca, and Mg, as well as an anticholinesterase effect in the animals [marked by the reduction of acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) activity]. On the other hand, exposure to PE MPs induced a genotoxic effect (assessed by the comet assay), as well as an increase in enzyme activity alpha-amylase, alkaline phosphatase, and carboxylesterases. However, we did not show synergistic, antagonistic, or additive effects caused by the combined exposure of P. reticulata to SARS-CoV-2 and PE MPs. Principal component analysis (PCA) and values from the "Integrated Biomarker Response" index indicate that exposure to SARS-CoV-2 was determinant for a more prominent effect in the evaluated animals. Therefore, our study sheds light on the ecotoxicity of the new coronavirus in non-target organisms and ratifies the need for more attention to the impacts of COVID-19 on aquatic biota.
Assuntos
COVID-19 , Poluentes Químicos da Água , Animais , Masculino , Humanos , Microplásticos/toxicidade , Polietileno/toxicidade , Plásticos/toxicidade , SARS-CoV-2 , Acetilcolinesterase , Pandemias , Butirilcolinesterase , Peixes , Biomarcadores , Poluentes Químicos da Água/toxicidade , Poluentes Químicos da Água/análiseRESUMO
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.
Assuntos
COVID-19 , SARS-CoV-2 , Animais , Humanos , Peixe-Zebra , Macrófagos , PeptídeosRESUMO
Advances in vaccine development depend on animal models to test innovative therapies. Recent studies have reported the successful introduction of teleost fish as a new vertebrate model in scientific research, with emphasis on the species Danio rerio (zebrafish). This chapter aims to give an overview of important aspects related to the immune system of fish, as well as the current progress of the successful use of these animals in studies for the development of vaccines, assisting in the determination of efficacy and clinical safety. Among the advantages of using fish for the development of vaccines and immunomodulatory drugs, it is worth highlighting the reproductive capacity of these animals resulting in a high number of individuals belonging to the same spawning, transparent embryos, low cost of breeding and high genetic similarity that favor translational responses to vertebrate organisms like humans.
Assuntos
Vacinas , Peixe-Zebra , Animais , Agentes de Imunomodulação , Modelos Teóricos , Desenvolvimento de Vacinas , Peixe-Zebra/imunologiaRESUMO
The use of Poly (lactic acid) (PLA) as a slow-release vehicle for vaccines has attracted the attention of researchers, since its insertion improves the uptake of them, and reduces side effects or by stimulating recruited defense cells, assisting immunity without the need for booster vaccine doses. Seeking to develop new strategies for the administration of drugs and vaccines in aquaculture, we evaluated the biocompatibility and biodegradation of polymeric PLA devices and PLA plus vitamin E devices, implanted through subcutaneous (SC) and intraperitoneal (IP) routes in Nile tilapia. To carry out this study, 84 male tilapia (initial 243.82 ± 56.74 g; final 400.71 ± 100.54 g) were randomly distributed in 3 tanks (n = 28 fish per treatment/tank). The devices were prepared in two formulations: neat PLA (containing 100% PLA) and PLAVE (PLA plus vitamin E) implanted using a commercial AnimalTag® applicator, and non-implanted fish (control). Fish were sampled 15, 30, 60, and 120 days post-implantation (DPI). Blood analysis was used to access blood cells and blood smear for differential leucocytes count. Serum biochemistry to evaluated changes in serum proteins and glycemia. Histopathological investigation using hematoxylin-eosin (H&E) was used to assess polymer-tissue interaction. Histochemistry and immunohistochemistry was used to detection immune cells and phagocytes in capsule, and analyses of melanomacrophage centers (MMCs) to morphometric evaluation and percentage amount of melanin, hemosiderin and lipofucsin pigments. Histopathological study revealed an increase of capsular formation and inflammatory cell infiltration in PLAVE-implanted tilapia through SC route (15 DPI). Tilapia implanted with PLAVE and PLA (SC) presented mast cells and eosinophilic granular cells during 15, 30, and 60 DPI, with a decrease in these cells in the fibrous capsule around the polymer at 120 DPI. PLAVE implanted tilapia SC at 60 DPI showed significantly phagocytosis points than other groups. Phagocytic cells (F4/80+) were observed near to biopolymers in phagocytosis sites. Lipofuscin at 120 DPI in spleen melanomacrophage centers were significantly high in PLAVE implanted tilapias when compared to fish with PLA implants and control. The serum biochemical study of tilapia did not reveal changes in cytotoxicity and liver function in implanted fish. The absence of side effects in hematological and biochemical findings, including the absence of mortality after device implantation, proves its clinical safety. PLA implants in tilapia have demonstrated biocompatibility, biodegradation, clinical safety, and excellent evolution of foreign body inflammatory responses.
RESUMO
Despite advances in research on the vaccine and therapeutic strategies of COVID-19, little attention has been paid to the possible (eco)toxicological impacts of the dispersion of SARS-CoV-2 particles in natural environments. Thus, in this study, we aimed to evaluate the behavioral and biochemical consequences of the short exposure of outbred and inbred mice (male Swiss and C57Bl/6 J mice, respectively) to PSPD-2002 (peptide fragments of the Spike protein of SARS-CoV-2) synthesized in the laboratory. Our data demonstrated that after 24 h of intraperitoneal administration of PSPD-2002 (at 580 µg/kg) the animals did not present alterations in their locomotor, anxiolytic-like, or anxiety-like behavior (in the open field test), nor antidepressant-like or depressive behavior in the forced swimming test. However, the C57Bl/6 J mice exposed to PSPD-2002 showed memory deficit in the novel object recognition task, which was associated with higher production of thiobarbituric acid reactive substances, as well as the increased suppression of acetylcholinesterase brain activity, compared to Swiss mice also exposed to peptide fragments. In Swiss mice the reduction in the activity of superoxide dismutase and catalase in the brain was not associated with increased oxidative stress biomarkers (hydrogen peroxide), suggesting that other antioxidant mechanisms may have been activated by exposure to PSPD-2002 to maintain the animals' brain redox homeostasis. Finally, the results of all biomarkers evaluated were applied into the "Integrated Biomarker Response Index" (IBRv2) and the principal component analysis (PCA), and greater sensitivity of C57Bl/6 J mice to PSPD-2002 was revealed. Therefore, our study provides pioneering evidence of mammalian exposure-induced toxicity (non-target SARS-CoV-2 infection) to PSPD-2002, as well as "sheds light" on the influence of genetic profile on susceptibility/resistance to the effects of viral peptide fragments.
Assuntos
COVID-19 , SARS-CoV-2 , Acetilcolinesterase , Animais , Biomarcadores , Masculino , Mamíferos , Camundongos , Camundongos Endogâmicos C57BL , Fragmentos de Peptídeos , PeptídeosRESUMO
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.
Assuntos
COVID-19 , SARS-CoV-2 , Animais , Anticorpos Antivirais , COVID-19/prevenção & controle , Vacinas contra COVID-19 , Humanos , Imunidade , Imunização , Pulmão , Proteínas do Nucleocapsídeo , Ratos , Ratos Endogâmicos Lew , Ratos Wistar , Proteínas Recombinantes , Glicoproteína da Espícula de Coronavírus , VacinaçãoRESUMO
There have been significant impacts of the current COVID-19 pandemic on society including high health and economic costs. However, little is known about the potential ecological risks of this virus despite its presence in freshwater systems. In this study, we aimed to evaluate the exposure of Poecilia reticulata juveniles to two peptides derived from Spike protein of SARS-CoV-2, which was synthesized in the laboratory (named PSPD-2002 and PSPD-2003). For this, the animals were exposed for 35 days to the peptides at a concentration of 40 µg/L and different toxicity biomarkers were assessed. Our data indicated that the peptides were able to induce anxiety-like behavior in the open field test and increased acetylcholinesterase (AChE) activity. The biometric evaluation also revealed that the animals exposed to the peptides displayed alterations in the pattern of growth/development. Furthermore, the increased activity of superoxide dismutase (SOD) and catalase (CAT) enzymes were accompanied by increased levels of malondialdehyde (MDA), reactive oxygen species (ROS) and hydrogen peroxide (H2O2), which suggests a redox imbalance induced by SARS-CoV-2 spike protein peptides. Moreover, molecular docking analysis suggested a strong interaction of the peptides with the enzymes AChE, SOD and CAT, allowing us to infer that the observed effects are related to the direct action of the peptides on the functionality of these enzymes. Consequently, our study provided evidence that the presence of SARS-CoV-2 viral particles in the freshwater ecosystems offer a health risk to fish and other aquatic organisms.