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1.
Insect Sci ; 27(2): 239-255, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30328680

RESUMO

Larval Galleria mellonella (L.) hemocytes form microaggregates in response to stimulation by Gram-positive bacteria. Hemocyte adhesion to foreign materials is mediated by the cAMP/ protein kinase A pathway and the ß-subunit of cholera toxin using a cAMP-independent mechanism. Cholera toxin-induced microaggregation was inhibited by the integrin inhibitory RGDS peptide, implying integrins may be part of the mechanism. Based on the types of mammalian integrin-antibody reactive proteins affecting hemocyte adhesion and bacterial-induced responses α5 , αv , ß1 , and ß3 subunits occurred on both granular cell and plasmatocyte hemocyte subtypes. A fluorescent band representing the binding of rabbit α5 -integrin subunit antibodies occurred between adhering heterotypic hemocytes. The frequency of the bands was increased by cholera toxin. The α5 and ß1 rabbit integrin subunit antibodies inhibited removal of Bacillus subtilis (Cohn) from the hemolymph in vivo. A α5 ß1 -specific synthetic peptide blocker similarly diminished hemocyte function whereas the αv ß3 -specific inhibitory peptide and the corresponding integrin subunit antibodies did not influence nonself hemocyte activities. Western blots revealed several proteins reacting with a given integrin-antibody subtype. Thus integrin-antibody reactive proteins (which may include integrins) with possible α5 and ß1 epitopes modulate immediate hemocyte function. Confocal microscopy established plasmatocyte adhesion to and rosetting over substrata followed by granular cell microaggregate adhesion to plasmatocytes during early stage nodulation.


Assuntos
Hemócitos/imunologia , Integrinas/imunologia , Mariposas/imunologia , Animais , Larva/imunologia
2.
Ecotoxicol Environ Saf ; 188: 109889, 2020 Jan 30.
Artigo em Inglês | MEDLINE | ID: mdl-31704331

RESUMO

It is known that abamectin (ABM) inflicts oxidative damage on aquatic animals; however, knowledge about the immune response under pesticide-induced oxidative stress is incomplete. In the present study, several cellular and humoral immune parameters, including total haemocyte counts (THC), lysosomal membrane stability (LMS), activities of acid phosphatase (ACP), alkaline phosphatase (AKP) and lysozyme (LZM) were investigated to reveal the effects of ABM exposure on the immune defence mechanisms of the important freshwater crab, Erocheir sinensis. According to the results, a significant increase of THC was found in low concentration groups (0.03 and 0.06 mg/L), while dramatic decreases occurred in high concentration groups (0.12 and 0.24 mg/L) after 96 h of exposure. We also detected significant increases of reactive oxygen species (ROS) in haemocytes at 0.12 and 0.24 mg/L, and there was a dose- and time-dependent decrease of lysosomal membrane stability. These results suggest that the excessive generation of ROS induced by ABM may be leading the massive collapse of lysosomal membrane, which in turn may be causing the sharp drop of haemocyte counts in E. sinensis. The increase of hydrolytic enzymes ACP and AKP at low concentrations and the decrease at high concentrations also indicate an immune response associated with haemocytes status under stress. However, activities of LZM decreased significantly. After injection of Aeromonas hydrophil, mortalities increased under exposure to ABM and were positively related to ABM concentration. These results confirm that ABM exposure has the ability to impair immune defence and result in the host's susceptibility to pathogens.


Assuntos
Braquiúros/imunologia , Inseticidas/toxicidade , Ivermectina/análogos & derivados , Estresse Oxidativo/efeitos dos fármacos , Poluentes Químicos da Água/toxicidade , Animais , Braquiúros/efeitos dos fármacos , Braquiúros/metabolismo , Hemócitos/efeitos dos fármacos , Hemócitos/metabolismo , Hemócitos/patologia , Imunidade Inata/efeitos dos fármacos , Ivermectina/toxicidade , Lisossomos/efeitos dos fármacos , Lisossomos/metabolismo , Espécies Reativas de Oxigênio/metabolismo
3.
Chemosphere ; 239: 124668, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31494325

RESUMO

Nitrite is one of major environmental pollutants that can impact immunological parameters in aquatic organisms. In the present study, we investigated the effects of nitrite exposure on oxidative stress, DNA damage and apoptosis in mud crab (Scylla paramamosain). Mud crab were exposed to 0, 5, 10 and 15 mg L-1 nitrite for 72 h. These data showed that acid phosphatase (ACP) and alkaline phosphatase (ALP) activity significantly decreased in treatments with various concentrations of nitrite (5, 10 and 15 mg L-1) after 24 and 48 h, while the levels of nitric oxide (NO) significantly increased in these treatments. Nitrite exposure could suppress superoxide dismutase (SOD) and catalase (CAT) activity, and increase the formation of malondialdehyde (MDA) after 48 and 72 h of exposure. In addition, nitrite exposure decreased total haemocyte counts after 48 and 72 h of exposure. Cytological damage, DNA damage and apoptosis was observed obviously at 72 h after nitrite exposure. Moreover, nitrite exposure significantly induced the mRNA levels of phosphorylated Jun N-terminal kinases (JNK), and eventually activated p53 signaling and caspase-3. These results indicated that nitrite exposure could induce oxidative stress, which further caused DNA damage and apoptosis in mud crab. Our results will be helpful to understand the mechanism of nitrite toxicity on crustaceans.


Assuntos
Apoptose/efeitos dos fármacos , Braquiúros/efeitos dos fármacos , Dano ao DNA/genética , Nitritos/toxicidade , Estresse Oxidativo/efeitos dos fármacos , Poluentes Químicos da Água/toxicidade , Fosfatase Ácida/metabolismo , Fosfatase Alcalina/metabolismo , Animais , Braquiúros/genética , Catalase/metabolismo , Hemócitos , Malondialdeído/metabolismo , Óxido Nítrico/metabolismo , Transdução de Sinais , Superóxido Dismutase/metabolismo
4.
Chemosphere ; 239: 124759, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31518920

RESUMO

Ammonia is an important environmental stress factor in aquaculture. Long-term ammonia stress could affect the normal growth, and also increase the risk for the occurrence of various diseases. In order to learn the mechanism that ammonia caused the outbreak of the shrimp disease, transcriptomics and metabolomics approaches were used to analyze the differential expressions of the genes in hemocytes and different metabolites in the serum of the Pacific white shrimp Litopenaeus vannamei under ammonia exposure. Transcriptional analysis showed that 17 cell apoptosis related genes, seven phagocytosis related genes, 10 immunity related genes and seven cell cycle and lipid metabolism related genes showed differential expressions after ammonia exposure. Metabolomics analysis on the serum showed that 25 differential metabolites were identified in positive and negative ion patterns. They are involved in purine metabolism, amino acids metabolism and lipid metabolism. Injection of two up-regulated metabolites triethanolamine and oxypurinol to normal shrimp could induce apoptosis in normal shrimp. The total hemocytes counts in shrimp showed a significant decrease and the apoptotic cell ratio increased significantly under ammonia exposure. These results suggested that ammonia exposure increased the apoptosis of hemocytes, which affected the immunity of shrimp, and thus caused susceptibility to pathogenic infection. These data will help us understand the mechanism of ammonia stress leading to the immunity decline of shrimp.


Assuntos
Amônia/toxicidade , Hemócitos/efeitos dos fármacos , Penaeidae/efeitos dos fármacos , Estresse Fisiológico/efeitos dos fármacos , Aminoácidos/genética , Aminoácidos/metabolismo , Animais , Apoptose/efeitos dos fármacos , Apoptose/genética , Aquicultura , Perfilação da Expressão Gênica , Hemócitos/metabolismo , Metabolismo dos Lipídeos/efeitos dos fármacos , Metabolômica , Penaeidae/fisiologia , Fagocitose/efeitos dos fármacos
5.
Bull Environ Contam Toxicol ; 103(6): 796-801, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31676939

RESUMO

The aim of this study was to determine the cytotoxic and genotoxic effects of copper on the bivalve Perumytilus purpuratus. The individuals were exposed to three copper concentrations: 1, 30 and 45 µg L-1 for 24, 48 and 96 h. Lysosomal membrane stability in hemocytes was determined through the neutral red retention time (NRRT) and micronucleus (MN) frequency tests in hemocytes and gills. The results show that the NRRT decreased significantly at 30 µg L-1 after 48 h of exposure. The frequency of MN was significantly greater in gills after 24 h in all concentrations tested. Copper is cytotoxic from 30 µg L-1 and genotoxic from 1 µg L-1. The use of these biomarkers of effects in P. purpuratus is proposed as an early warning tool for monitoring in environmental assessment of coastal ecosystems impacted by mining activities.


Assuntos
Cobre/toxicidade , Monitoramento Ambiental/métodos , Brânquias/efeitos dos fármacos , Membranas Intracelulares/efeitos dos fármacos , Micronúcleos com Defeito Cromossômico/induzido quimicamente , Mytilidae/efeitos dos fármacos , Poluentes Químicos da Água/toxicidade , Animais , Cobre/análise , Ecossistema , Biomarcadores Ambientais/efeitos dos fármacos , Brânquias/irrigação sanguínea , Hemócitos/citologia , Hemócitos/efeitos dos fármacos , Membranas Intracelulares/patologia , Lisossomos/efeitos dos fármacos , Lisossomos/ultraestrutura , Mytilidae/genética , Vermelho Neutro , Poluentes Químicos da Água/análise
6.
Pestic Biochem Physiol ; 160: 95-101, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31519262

RESUMO

Neonicotinoid insecticides are increasingly used in modern pest control and in conventional agriculture. Their residues are frequently found in our environment and in our food leading to chronic exposure of pollinating insects and humans. Indeed, evidence has become stronger that chronic exposure to neonicotinoids might have a direct impact on the immune response of invertebrates and vertebrates. Therefore, we compared the cellular immune response of human macrophages (THP-1) and Drosophila melanogaster hemocytes (Schneider 2 cells) after exposure to four different concentrations of the neonicotinoid imidacloprid. Cells were immune activated with LPS (lipopolysaccharide) of Escherichia coli to compare the phagocytic activity of immune activated and non-activated cells during pesticide exposure. Drosophila cells were more strongly affected by the insecticide than human macrophages. Even though imidacloprid showed an adverse effect on phagocytosis on both cells while immune activated, it decreased phagocytosis in Drosophila cells at shorter exposure time and without immune activation.


Assuntos
Drosophila melanogaster/metabolismo , Hemócitos/efeitos dos fármacos , Inseticidas/farmacologia , Macrófagos/efeitos dos fármacos , Neonicotinoides/farmacologia , Nitrocompostos/farmacologia , Fagocitose/efeitos dos fármacos , Animais , Linhagem Celular , Hemócitos/imunologia , Humanos , Macrófagos/imunologia
7.
Aquat Toxicol ; 215: 105284, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31479758

RESUMO

Metal pollution in the environment is a serious threat to the biological sustainability of coastal ecosystems. However, our current understanding of the biological effects of metals in these ecosystems is limited. Herein, we investigated the responses of the sea slug Onchidium reevesii to persistent sublethal Cd environmental stress. Dynamic expression was analyzed using various biomarkers. The full-length cDNA of O. reevesii metallothionein (MT) was cloned and consists of 1639 nucleotides encoding a 65 amino acid polypeptide. Phylogenetic analysis showed that Or-MT has conserved Cys residues typical of MTs, including a typical Cys-X-Cys motif, implying that it can function the same as the MT of other shellfish. Expression of Or-MT in response to Cd varied in different tissues, and was highest in gastropod tissues. Thus, regiotemporal expression of MT may be useful for assessing pollution in coastal areas. Cellular immunity (in the hemolymph) and enzyme activity (in the hepatopancreas) were investigated along with hemocyte viability, hemocyte phagocytosis, and superoxide dismutase (SOD) and aspartate aminotransferase (AST) activities. Hemocyte viability was elevated under continuous Cd exposure but hemocyte phagocytosis was decreased. SOD and AST activities in the hepatopancreas fluctuated considerably, and SOD activity was more sensitive. SOD activity was lowest at 4 h and highest at 12 h, while AST activity peaked at 2 h and was lowest at 48 h. Thus, changes in enzyme activity may reveal adaptation to stress. Furthermore, the response patterns of certain enzymes, cellular immunity, and MT expression in O. reevesii could serve as biomarkers of Cd pollution in aquatic environments.


Assuntos
Aspartato Aminotransferases/metabolismo , Biomarcadores/metabolismo , Cádmio/toxicidade , Gastrópodes/metabolismo , Hemócitos/metabolismo , Metalotioneína/metabolismo , Estresse Fisiológico/efeitos dos fármacos , Superóxido Dismutase/metabolismo , Sequência de Aminoácidos , Animais , Sequência de Bases , Sobrevivência Celular/efeitos dos fármacos , Exposição Ambiental , Gastrópodes/química , Gastrópodes/efeitos dos fármacos , Gastrópodes/genética , Hemócitos/efeitos dos fármacos , Fagocitose/efeitos dos fármacos , Filogenia , Homologia de Sequência de Aminoácidos , Poluentes Químicos da Água/toxicidade
8.
Aquat Toxicol ; 215: 105266, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31401474

RESUMO

The persistent pollutants polybrominated diphenyl ethers (PBDEs) have been demonstrated to produce several negative effects on marine organisms. Although Mytilus galloprovincialis was extensively studied as model system, the effects of PBDEs on the innate immune system of mussels remains unclear. In this study, except for the control treatment, specimens of M. galloprovincialis were fed with microalgae treated with increasing concentrations of PBDEs (maximum level 100 ng L-1 of BDE-47 per day). BDE-47 treatment was maintained for 15 days and then the animals were fed with the same control diet, without contaminants, for 15 days. Samples of haemolymph (HL) were obtained at T0, T15 and T30 days of the experiment to evaluate different parameters related to immunity, such as neutral red retention time, and peroxidase, protease, antiprotease, lysozyme and bactericidal activities. BDE-47 exposure for 15 days affected both the stability of haemocytes and humoral parameters. In addition, the obtained results indicated that, at 30 days, after 15 days of culture without contaminant, the immune parameters were still affected, as some of them did not return to the basal levels, and others remained stimulated. Overall the results indicate that BDE-47 exposures at environmentally realistic levels may affect various aspects of immune function in M. galloprovincialis, acting as stressor that can compromise the general welfare.


Assuntos
Exposição Ambiental , Éteres Difenil Halogenados/toxicidade , Mytilus/imunologia , Animais , Antibacterianos/farmacologia , Comportamento Alimentar/efeitos dos fármacos , Hemócitos/efeitos dos fármacos , Hemócitos/metabolismo , Hemolinfa/efeitos dos fármacos , Hemolinfa/metabolismo , Hemolinfa/microbiologia , Membranas Intracelulares/efeitos dos fármacos , Membranas Intracelulares/metabolismo , Lisossomos/efeitos dos fármacos , Lisossomos/metabolismo , Microalgas/fisiologia , Mytilus/efeitos dos fármacos , Mytilus/microbiologia , Peptídeo Hidrolases/metabolismo , Análise de Sobrevida , Poluentes Químicos da Água/toxicidade
9.
J Chem Ecol ; 45(8): 715-724, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31385154

RESUMO

Plants emit a specific blend of volatiles in response to herbivory and these volatiles, which often attract predators and parasitoids function as an indirect plant defense. The impact of plant volatiles in shaping herbivore defenses is unclear. Here, we report that specific plant volatiles induce immune responses in the polyphagous herbivore, Spodoptera litura. We characterized the hemocyte profile and established their functional significance with respect to ontogeny and exposure to specific plant volatiles. Fifth instar larvae showed the highest number and hemocytes diversity. We characterized seven different types of hemocytes, of which granulocytes performed phagocytosis, oenocytoids showed melanization activity, and plasmatocytes along with granulocytes and oenocytoids were found to be involved in encapsulation. Among the six volatiles tested, exposure to (E)-ß-ocimene caused the highest increase in total hemocytes number (THC) followed by linalool and (Z)-3-hexenyl acetate exposure. Although THC did not differ between these three volatile treatments, circulating hemocytes diversity varied significantly. (E)-ß-ocimene exposure showed higher number of plasmatocytes and phenol oxidase activity. The interaction of the parasitic wasp Bracon brevicornis with (E)-ß-ocimene exposed larvae was poor in terms of delayed paralysis and lower egg deposition. In choice assays, the wasp showed clear preference towards control larvae indicating (E)-ß-ocimene treatment renders the host unattractive. Hemocyte profiles post-parasitoid exposure and (E)-ß-ocimene treatment were similar indicating cue-based priming. When challenged with Bacillus thuringiensis, linalool exposure resulted in the highest survival as compared to other volatiles. Our results show that specific HIPVs can modulate cellular immunity of S. litura, revealing a new role for HIPVs in tri-trophic interactions.


Assuntos
Compostos Orgânicos Voláteis/farmacologia , Vespas/fisiologia , Alcenos/farmacologia , Animais , Comportamento Animal/efeitos dos fármacos , Hemócitos/citologia , Hemócitos/efeitos dos fármacos , Hemócitos/metabolismo , Herbivoria , Interações Hospedeiro-Parasita , Sistema Imunitário/efeitos dos fármacos , Sistema Imunitário/metabolismo , Larva/fisiologia , Monofenol Mono-Oxigenase/metabolismo , Monoterpenos/farmacologia , Oviposição/efeitos dos fármacos , Fagocitose/efeitos dos fármacos , Plantas/química , Plantas/metabolismo , Plantas/parasitologia , Compostos Orgânicos Voláteis/química , Vespas/imunologia
10.
J Therm Biol ; 84: 136-145, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31466746

RESUMO

Temperature is one of the important environmental elements affecting ecological fitness of insects through alterations in physiological systems. In the current study, a comparison was made on the cellular and humoral immune responses of the Chilo suppressalis larvae exposed to thermal stress (34 °C) and optimal rearing temperature (24 °C). Although total hemocyte count increased in the injected larvae by Beauveria bassiana, elevation of hemocyte numbers was significantly different in the larvae exposed to 34 °C for a short-time period compared to long-term exposure and control. A similar trend was observed in plasmatocyte and granulocyte counts as well as phenoloxidase activity. Gene expression of some antimicrobial peptides, including attacin1, attacin2, cecropin1, cecropin2, defensin, gallerimycin, lysozyme and prophenoloxidase-activating proteinase-3, was compared in the larvae exposed to thermal regimes and injection challenges. In all cases, expression of the target genes was relatively higher in the larvae injected by B. bassiana and short-term exposure at 34 °C. The present results confirmed that C. suppressalis could modulate the immune system in response to different thermal stress conditions mainly over a short period.


Assuntos
Beauveria , Resposta ao Choque Térmico , Interações Hospedeiro-Patógeno/imunologia , Larva/imunologia , Lepidópteros/imunologia , Animais , Contagem de Células Sanguíneas , Granulócitos/imunologia , Hemócitos/imunologia , Proteínas de Insetos/imunologia , Irã (Geográfico) , Larva/microbiologia , Lepidópteros/microbiologia , Monofenol Mono-Oxigenase/imunologia
11.
Fish Shellfish Immunol ; 93: 997-1006, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31412281

RESUMO

The viral responsive protein 15 from the black tiger shrimp Penaeus monodon (PmVRP15) is a highly responsive gene upon white spot syndrome virus (WSSV) challenge. It is identified from hemocyte and important for WSSV trafficking and assembly. However, the knowledge of PmVRP15 gene regulation is limited. In the present study, the genome organization and 5'upstream promoter sequences of PmVRP15 gene were investigated. The PmVRP15 gene was found to contain 4 exons interrupted by 3 introns and the start codon was located in the exon 2. The transcription start site and TATA box were also determined from the 5' upstream sequence. By using the narrow down experiment, the 5' upstream promoter active region was determined to be at the nucleotide positions -525 to +612. Mutagenesis of the putative transcription factor (TF) binding sites revealed that the binding site of interferon regulatory factor (IRF) (-495/-479) was a repressor-binding site whereas those of the octamer transcription factor 1 (Oct-1) (-275/-268) and the nuclear factor of activated T-cells transcription factor (NFAT) (-228/-223) were activator-binding sites. This is the first report on the transcription factors that might play essential roles in modulating the PmVRP15 gene expression. Nevertheless, the underlying regulation mechanism of PmVRP15 gene expression needs further investigation.


Assuntos
Proteínas de Artrópodes/genética , Proteínas de Artrópodes/imunologia , Regulação da Expressão Gênica/imunologia , Genoma , Imunidade Inata/genética , Penaeidae/genética , Penaeidae/imunologia , Sequência de Aminoácidos , Animais , Proteínas de Artrópodes/química , Sequência de Bases , Perfilação da Expressão Gênica , Hemócitos/metabolismo , Interações Hospedeiro-Patógeno , Vírus da Síndrome da Mancha Branca 1/fisiologia
12.
Fish Shellfish Immunol ; 93: 416-427, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31374314

RESUMO

Phagocytosis is one of the fundamental cellular immune defense parameter that helps in the elimination of the invading pathogens in both vertebrates and invertebrates, which require plenty of energy for functioning. In the present study, we identified the critical energy regulator AMP-activated protein kinase (AMPK) in Crassostrea hongkongensis which is composed of three subunits, named ChAMPK-α, ChAMPK-ß, and ChAMPK-γ, and then analyzed the function of AMPK in regulating hemocyte phagocytosis. All the three ChAMPK subunits mRNA were detected to be expressed at various embryological stages, and also constitutively expressed in multiple tissues with high expression in gill and mantle. The phylogenetic tree showed that the three subunits of AMPK were correspondingly clustered with its orthologue branches. Furthermore Western Blot analysis revealed that the AMPK pharmacological inhibitors Compound C could effectively down-regulate the Thr172 phosphorylation level of AMPK-α, and the hemocyte phagocytosis was inhibited by Compound C (CC), which indicate its existence in the oyster. Our results showed that treatment of AMPK inhibitors significantly attenuated the capacity of hemocytes phagocytosis. Moreover, Compound C could also change the organization of actin cytoskeleton in the oyster hemocytes, demonstrating the crucial role of AMPK signaling in control of phagocytosis.


Assuntos
Proteínas Quinases Ativadas por AMP/genética , Proteínas Quinases Ativadas por AMP/imunologia , Crassostrea/genética , Crassostrea/imunologia , Regulação da Expressão Gênica/imunologia , Imunidade Inata/genética , Proteínas Quinases Ativadas por AMP/química , Sequência de Aminoácidos , Animais , Sequência de Bases , Perfilação da Expressão Gênica , Hemócitos , Fagocitose , Alinhamento de Sequência , Transdução de Sinais
13.
Parasitol Res ; 118(9): 2509-2521, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31377908

RESUMO

In this study, we characterize the HIP-55 protein in the mosquito Aedes aegypti for the first time. HIP-55 is a 55-kDa HPK1-interacting protein that is also called SH3P7. HIP-55 constitutively binds HPK1 'via' an HPK1 proline-rich motif 2(PR2) through its C-terminal SH3 domain. HIP-55 critically interacts with ZAP-70, and this interaction was induced by TCR signalling. ZAP-70 phosphorylated HIP-55 at Tyr-334 and Tyr-344 in vitro and in vivo. In our previous findings, AaZAP gene expression strongly proved that AaZAP-70 was involved in immunity-like functions in mosquito. Northern blot analysis of HIP-55 mRNA expression confirmed that it is only expressed in the abdomen and haemocyte tissues; this prediction correlates 100% and a polyclonal antibody also confirmed its localization in haemocytes and the abdomen. We prepared extracts to show the cytoplasmic expression (CE) of this protein. Previous results had proven that this protein is secreted from the cytoplasm; thus, we confirmed here that the protein is a cytoplasmic adaptor protein in mosquitoes and mammalian systems. Furthermore, our polyclonal antibody against HIP-55 also demonstrated that this protein is found in haemocytes and abdomen tissues, which assumes that the protein may be involved in phagocytic-like functions. RNAi (siRNA) silencing studies were used to degrade mosquito HIP-55; however, silencing only slightly affected the HIP-55 sequence and the gene transcriptional level. To characterize this protein, we cloned 609 bp from the 1.6-kb full-length cDNA using a pET28 vector for polyclonal antibody production. Graphical abstract.


Assuntos
Aedes/imunologia , Proteínas dos Microfilamentos/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Domínios de Homologia de src/fisiologia , Animais , Hemócitos/metabolismo , Proteínas dos Microfilamentos/genética , Fosforilação , Ligação Proteica , Interferência de RNA , RNA Mensageiro/genética , RNA Interferente Pequeno/genética , Transdução de Sinais , Proteína-Tirosina Quinase ZAP-70/metabolismo , Domínios de Homologia de src/genética
14.
Aquat Toxicol ; 214: 105262, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31351400

RESUMO

To understand the toxic effects of nitrite exposure on crayfish, expression of genes involved in the immune system, the antioxidant defense, and the heat shock protein 70 (HSP70) was measured after 12, 24, and 48 h of different nitrite concentrations exposure in the hepatopancreas and hemocytes of Procambarus clarkii. Nitrite exposure up-regulated mRNA levels of cytoplasmic Mn superoxide dismutase (cMn-SOD), catalase (CAT), glutathione peroxidase (GPx), and glutathione-S-transferase (GST), after 24 h nitrite exposure. At 48 h, nitrite exposure decreased the mRNA levels of mitochondrial MnSOD (mMn-SOD), CAT, and GPx. High concentrations of nitrite at 48 h of exposure decreased expression of ß-1,3-glucan-bingding protein in the hepatopancreas, and lysozyme expression in hemocytes. Nitrite exposure caused little effect on the heat shock protein 70 (HSP70) in hemocytes. Through overall clustering analysis, we found that 24 h of nitrite exposure caused stronger transcriptional responses. Our study indicated that the response of P. clarkii to acute nitrite exposure was exposure time-dependent. These results will help to understand the dynamic response pattern of crustaceans to nitrite pollution, and improve our understanding of the toxicological mechanisms of nitrite in crustaceans.


Assuntos
Antioxidantes/metabolismo , Astacoidea/genética , Astacoidea/imunologia , Exposição Ambiental , Nitritos/toxicidade , Transcrição Genética , Animais , Astacoidea/efeitos dos fármacos , Catalase/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Glutationa Peroxidase/metabolismo , Glutationa Transferase/metabolismo , Proteínas de Choque Térmico HSP70/metabolismo , Hemócitos/efeitos dos fármacos , Hemócitos/enzimologia , Hepatopâncreas/efeitos dos fármacos , Hepatopâncreas/enzimologia , Superóxido Dismutase/metabolismo , Fatores de Tempo , Transcrição Genética/efeitos dos fármacos , Poluentes Químicos da Água/toxicidade
15.
Arch Insect Biochem Physiol ; 102(4): e21607, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31338878

RESUMO

Prostaglandin E2 (PGE2 ) mediates cellular immune responses in insects by stimulating hemocyte-spreading behavior that is driven by actin remodeling to form filopodial or lamellipodial cytoplasmic extensions. In Spodoptera exigua (Lepidoptera: Noctuidae), Cdc42, a small G protein, played a crucial role in mediating PGE2 signal on hemocyte-spreading behavior. Hemocyte-spreading behavior requires actin cytoskeletal rearrangement. A plethora of actin-related proteins have been predicted to have functional links with Cdc42. Here, we selected four actin-associated genes (Actin-related protein 2 [Arp2], Profilin, Cofilin, and Fascin) and evaluated their influences on cytoskeletal rearrangement in S. exigua. Bioinformatic analysis confirmed their gene identities. Transcript analysis using reverse-transcription polymerase chain reaction indicated that all four actin-associated genes were expressed in most developmental stages, showing high expression levels in larval hemocytes. RNA interference (RNAi) against these genes was performed by injecting double-stranded RNA (dsRNA) to hemocoel. Under RNAi condition, the hemocyte-spreading behavior was significantly impaired except for dsRNA treatment against Cofilin, an actin-depolymerizing factor. Alteration of cytoskeletal rearrangement appeared to vary after different RNAi treatments. RNAi against Arp2 markedly suppressed lamellipodial extension while RNAi against Profilin or Fascin adversely influenced filopodial extension. RNAi of these actin-associated factors prevented cellular immune responses measured by nodule formation against bacterial challenge. Under RNAi conditions, addition of PGE2 did not well induce hemocyte-spreading behavior, suggesting that these actin-associated factors might act downstream of the hormone signaling pathway. These results suggest that PGE2 can mediate hemocyte-spreading behavior via Cdc42 to activate downstream actin polymerization/branching/bundling factors, thus inducing actin cytoskeletal rearrangement.


Assuntos
Actinas/metabolismo , Dinoprostona/metabolismo , Hemócitos/citologia , Proteínas de Insetos/metabolismo , Spodoptera/citologia , Actinas/genética , Animais , Escherichia coli , Imunidade Celular , Proteínas de Insetos/genética , Larva/metabolismo , Interferência de RNA , Spodoptera/genética , Spodoptera/metabolismo
16.
Chemosphere ; 236: 124349, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31326753

RESUMO

Heavy metal Pb is a common toxic pollutant present in our environment adversely affecting health of the living organisms. Recent studies suggest positive correlation between heavy metal exposure and immune dysfunction and present work utilizes Drosophila to address this issue in relation to Pb exposure. In-vivo Pb toxicity was established by dietary intake where essential parameters like development and life span were found to be hampered and augmented upon metallothionein B (mtnB) downregulation hinting towards potential role of mtnB in Pb detoxification. Further response of Drosophila to B. subtilis bacterial infection was monitored by carrying out oral infections. Pb fed flies showed increased susceptibility to infection as compared to their controls. Since Drosophila hemocytes play dual role as immune cells, we checked for the total hemocyte count and found significant decrease in hemocyte numbers in Pb fed larvae. Both crystal cells and plasmatocytes, the two major hemocytes in third instar larval hemolymph were reduced. However we did not find any visible morphological changes in Giemsa stained hemocytes. Crystal cells are crucial for synthesis and release of phenoloxidase (PO), an enzyme required for melanin clot synthesis and deposition. PO activity assessed from total hemolymph protein isolates was found to be substantially decreased in Pb raised animals. Results were also confirmed by spot test and native gel activity assay of PO. Overall our results suggest immunotoxic effect of Pb through decrease in hemocyte count including crystal cell which in turn leads to decreased PO activity and increased susceptibility to B. subtilis.


Assuntos
Drosophila melanogaster/imunologia , Poluentes Ambientais/toxicidade , Infecções por Bactérias Gram-Positivas/induzido quimicamente , Hemócitos/citologia , Chumbo/toxicidade , Animais , Bacillus subtilis , Drosophila melanogaster/microbiologia , Infecções por Bactérias Gram-Positivas/microbiologia , Hemócitos/metabolismo , Hemolinfa/metabolismo , Larva/efeitos dos fármacos , Metalotioneína/metabolismo , Monofenol Mono-Oxigenase/metabolismo
17.
Environ Pollut ; 252(Pt B): 1764-1771, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31295695

RESUMO

Phagocytosis suppression induced by nanoparticles (NPs) exposure is increasingly reported in marine species. However, the mechanisms underlying this impact remain poorly understood. In order to improve our present understanding of the immunotoxicity of NPs, acute (96 h) TiO2 NP exposure and rescue trials via exogenous supply of Ca2+ were performed in the blood clam, Tegillarca granosa. The results show that the phagocytosis rate, cell viability, and intracellular Ca2+ concentration of haemocytes were significantly suppressed, whereas the intracellular ROS concentration of haemocytes significantly increased upon nTiO2 exposure. Exposure to nTiO2 also led to the significant downregulation of Caspase-3, Caspase-6, apoptosis regulator Bcl-2, Bcl-2-associated X, calmodulin kinase II, and calmodulin kinase kinase II. Furthermore, the toxic impacts of nTiO2 were partially mitigated by the addition of exogenous Ca2+, as indicated by the recovery tendency in almost all the measured parameters. The present study indicates that Ca2+ signaling could be one of the key pathways through which nTiO2 attacks phagocytosis.


Assuntos
Apoptose/efeitos dos fármacos , Arcidae/efeitos dos fármacos , Cálcio/farmacologia , Hemócitos/efeitos dos fármacos , Nanopartículas/toxicidade , Fagocitose/efeitos dos fármacos , Titânio/toxicidade , Poluentes Químicos da Água/toxicidade , Animais , Apoptose/genética , Arcidae/fisiologia , Cálcio/metabolismo , Sinalização do Cálcio/efeitos dos fármacos , Sinalização do Cálcio/genética , Sobrevivência Celular/efeitos dos fármacos , Regulação para Baixo , Hemócitos/patologia
18.
Fish Shellfish Immunol ; 93: 559-566, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31330256

RESUMO

Shrimps like other arthropods rely on innate immune system, and may have some form of adaptive immunity in defending against pathogens. Phagocytosis is one of the oldest cellular processes, serving as a development process, a feeding mechanism and especially as a key defense reaction in innate immunity of all multicellular organisms. It is confirmed that crustacean hyperglycemic hormone (CHH) is one of the most important neuropeptides produced by Neuro-endocrine Immune (NEI) regulatory network, which undertakes important roles in various biological processes, especially in immune function and stress response. In this study, the recombinant Litopenaeus vannamei CHH (rLvCHH) was obtained from a bacterial expression system and the intracellular signaling pathways involved in the mechanism of phagocytosis after rLvCHH injection was investigated. The results showed that the contents of adenylyl cyclase (AC), phospholipase C (PLC) and calmodulin (CaM) in hemocytes were increased significantly after rLvCHH injection. Furthermore, the mRNA expression levels of NF-kB family members (relish and dorsal) and phagocytosis-related proteins in hemocytes were basically overexpressed after rLvCHH stimulation, while the expression level of NF-kB repressing factor (NKRF) gene was down-regulated significantly. Eventually, the total hemocyte count and phagocytic activity of hemocyte were dramatically enhanced within 3 h. Collectively, these results indicate that shrimps L. vannamei could carry out a simple but 'smart' NEI regulation through the action of neuroendocrine factors, which could couple with their receptors and trigger the downstream signaling pathways during the phagocytic responses of hemocytes.


Assuntos
Proteínas de Artrópodes/imunologia , Hemócitos/imunologia , Imunidade Inata/genética , Hormônios de Invertebrado/imunologia , Proteínas do Tecido Nervoso/imunologia , Penaeidae/genética , Penaeidae/imunologia , Fagocitose/imunologia , Animais , Proteínas de Artrópodes/genética , Relação Dose-Resposta a Droga , Hormônios de Invertebrado/genética , Proteínas do Tecido Nervoso/genética , Distribuição Aleatória , Transdução de Sinais/imunologia
19.
Ecotoxicology ; 28(7): 801-808, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31317358

RESUMO

Zinc oxide nanoparticles (ZnO NPs) are now commonly used in many consumer products (detergents, antibacterial products, protective creams). The aim of the study is to determine the ecotoxicological effects of ZnO NPs on the survival and in the total hemocyte counts of Galleria mellonella L. 1758 (Lepidoptera: Pyralidae) with two different methods (automated cell counter and hemocytometer). A toxicity test was performed to determine the lethal concentrations of ZnO NPs on larvae by force feeding method. After 24 h the treatment, LC50 was 6.03 µg/10 µl and LC99 was 12.86 µg/10 µl for force fed larvae according to probit assay. The NP that induced changes in the total hemocytes counts were counted by optical microscopy (larvae exposed to four different doses of ZnO NPs) and by automated cell counter. Analyses of total hemocyte counts of the insect were performed with four doses (0.5, 1, 2.5, 5 µg/10 µl)

Assuntos
Hemócitos/efeitos dos fármacos , Nanopartículas Metálicas/toxicidade , Mariposas/efeitos dos fármacos , Testes de Toxicidade Aguda/métodos , Óxido de Zinco/toxicidade , Animais , Larva/efeitos dos fármacos , Larva/crescimento & desenvolvimento , Dose Letal Mediana , Mariposas/crescimento & desenvolvimento
20.
Sci Total Environ ; 689: 381-389, 2019 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-31277005

RESUMO

There are several reports on the damage smoking causes to human health available in the literature, but little is known about the environmental and biological consequences from inappropriate cigarette butt (CB) disposal in urban and natural environments. The immunotoxic, morphotoxic and mutagenic potential of leachates from cigarette butts (LCB) diluted at environmentally relevant rates (LCB1x: 1.375%; LCB10x: 13.75%) was evaluated in adult representatives of the bivalve species Anodontites trapesialis, which was adopted as model organism. Type II hyalinocytes and granulocytes (phagocytic cells) frequency increased in the hemolymph of subjects exposed to the pollutant for 14 days. Based on this outcome, LCB chemical constituents did not induce immunotoxic effects. The treatments also did not seem to have any impact on the subjects' hemocitary morphometry parameters: diameter, area, perimeter, circularity and nucleus - cytoplasm ratio. However, subjects in groups LCB1x and LCB10x recorded a larger number of hyalinocytes with some nuclear abnormality such as micronucleus, blebbed nucleus, asymmetric constriction nucleus, and nuclear multilobulation and binucleation. The association between these abnormalities and the treatments was confirmed by the Cr, Ni, Pb, Zn, Mn and Na bioaccumulation in tissue samples of the bivalve models exposed to LCB. To the best of our knowledge, this is the first report on LCB mutagenicity in representatives of a freshwater bivalve group. Given the chemical complexity of the addressed pollutants, it is imperative to develop further investigations about the topic.


Assuntos
Bivalves/efeitos dos fármacos , Hemócitos/efeitos dos fármacos , Produtos do Tabaco , Poluentes Químicos da Água/toxicidade , Animais , Relação Dose-Resposta a Droga , Monitoramento Ambiental
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