RESUMO
Porcine epidemic diarrhea virus (PEDV) belongs to the Alphacoronavirus genus within the Coronavirus family, causing severe watery diarrhea in piglets and resulting in significant economic losses. Medium-chain acyl-CoA dehydrogenase (ACADM) is an enzyme participating in lipid metabolism associated with metabolic diseases and pathogen infections. Nonetheless, the precise role of ACADM in regulating PEDV replication remains uncertain. In this study, we identified ACADM as the host binding partner of NSP4 via immunoprecipitation-mass spectrometry analysis. The interaction between ACADM and NSP4 was subsequently corroborated through coimmunoprecipitation and laser confocal microscopy. Following this, a notable upsurge in ACADM expression was observed during PEDV infection. ACADM overexpression effectively inhibited virus replication, whereas ACADM knockdown facilitated virus replication, suggesting ACADM has negative regulation effect on PEDV infection. Furthermore, we demonstrated fatty acid ß-oxidation affected PEDV replication for the first time, inhibition of fatty acid ß-oxidation reduced PEDV replication. ACADM decreased PEDV-induced ß-oxidation to suppress PEDV replication. Mechanistically, ACADM reduced cellular free fatty acid levels and subsequent ß-oxidation by hindering AMPK-mediated lipophagy. In summary, our results reveal that ACADM plays a negative regulatory role in PEDV replication by regulating lipid metabolism. The present study introduces a novel approach for the prevention and control of PEDV infection.
RESUMO
Fumonisin B1 (FB1), a water-soluble mycotoxin released by Fusarium moniliforme Sheld, is widely present in corn and its derivative products, and seriously endangers human life and health. Recent studies have reported that FB1 can lead to pyroptosis, however, the mechanisms by which FB1-induced pyroptosis remain indistinct. In the present study, we aim to investigate the mechanisms of pyroptosis in intestinal porcine epithelial cells (IPEC-J2) and the relationship between FB1-induced endoplasmic reticulum stress (ERS) and pyroptosis. Our experimental results showed that the pyroptosis protein indicators in IPEC-J2 were significantly increased after exposure to FB1. The ERS markers, including glucose-regulated Protein 78 (GRP78), PKR-like ER kinase protein (PERK), and preprotein translocation factor (Sec62) were also significantly increased. Using small interfering RNA silencing of PERK or Sec62, the results demonstrated that upregulation of Sec62 activates the PERK pathway, and activation of the PERK signaling pathway is upstream of FB1-induced pyroptosis. After using the ERS inhibitor 4-PBA reduced the FB1-triggered intestinal injury by the Sec62-PERK pathway. In conclusion, we found that FB1 induced pyroptosis by upregulating Sec62 to activate the PERK pathway, and mild ERS alleviates FB1-triggered damage. It all boils down to one fact, the study provides a new perspective for further, and improving the toxicological mechanism of FB1.
Assuntos
Chaperona BiP do Retículo Endoplasmático , Estresse do Retículo Endoplasmático , Piroptose , Transdução de Sinais , eIF-2 Quinase , Piroptose/efeitos dos fármacos , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Animais , eIF-2 Quinase/metabolismo , eIF-2 Quinase/genética , Suínos , Transdução de Sinais/efeitos dos fármacos , Chaperona BiP do Retículo Endoplasmático/metabolismo , Linhagem Celular , Intestinos/efeitos dos fármacos , Intestinos/patologia , Células Epiteliais/metabolismo , Células Epiteliais/efeitos dos fármacos , Mucosa Intestinal/metabolismo , Mucosa Intestinal/efeitos dos fármacos , FumonisinasRESUMO
Sustaining DNA damage response (DDR) signalling via retention of DDR factors at damaged sites is important for transmitting damage-sensing and repair signals. Herein, we found that DNA damage provoked the association of ribosomes with IRES region in lncRNA CTBP1-DT, which overcame the negative effect of upstream open reading frames (uORFs), and elicited the novel microprotein DNA damage-upregulated protein (DDUP) translation via a cap-independent translation mechanism. Activated ATR kinase-mediated phosphorylation of DDUP induced a drastic 'dense-to-loose' conformational change, which sustained the RAD18/RAD51C and RAD18/PCNA complex at damaged sites and initiated RAD51C-mediated homologous recombination and PCNA-mediated post-replication repair mechanisms. Importantly, treatment with ATR inhibitor abolished the effect of DDUP on chromatin retention of RAD51C and PCNA, thereby leading to hypersensitivity of cancer cells to DNA-damaging chemotherapeutics. Taken together, our results uncover a plausible mechanism underlying the DDR sustaining and might represent an attractive therapeutic strategy in improvement of DNA damage-based anticancer therapies.
Assuntos
Dano ao DNA , Reparo do DNA , RNA Longo não Codificante , Cromatina , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Recombinação Homóloga , Neoplasias/tratamento farmacológico , Neoplasias/genética , Antígeno Nuclear de Célula em Proliferação/genética , Antígeno Nuclear de Célula em Proliferação/metabolismo , Biossíntese de Proteínas , RNA Longo não Codificante/genéticaRESUMO
Chronic pain is frequently reported in clinical practice. Therefore, it is important to identify effective therapy to relieve pain. In this work, we selected Forsythoside B (FB), a phenylethanoid glycoside isolated from Forsythia suspensa (Thunb.) Vahl, to evaluate its effect in modulating inflammatory pain induced by complete Freund's adjuvant (CFA) and the involved mechanisms. We discovered that FB could attenuate inflammatory pain triggered by CFA injection and exert anti-anxiety effects. In detail, proinflammatory cytokines, consisting of IL-6 and TNF-α, were decreased after FB administration in the CFA-injected mice. Furthermore, the FB application ameliorated the activation of ionized calcium-binding adaptor molecule 1 (Iba-1) and glial fibrillary acidic protein (GFAP), the microglia and astrocytes markers respectively. Therefore, our findings indicate that FB could be a promising treatment for chronic inflammatory pain.
Assuntos
Dor Crônica , Inflamação , Camundongos , Animais , Adjuvante de Freund , Inflamação/induzido quimicamente , Inflamação/tratamento farmacológico , Inflamação/metabolismo , Anti-Inflamatórios/uso terapêutico , Anti-Inflamatórios/farmacologia , Dor Crônica/induzido quimicamente , Dor Crônica/tratamento farmacológico , Glucosídeos/farmacologia , Glucosídeos/uso terapêutico , Analgésicos/farmacologia , Analgésicos/uso terapêutico , Hiperalgesia/metabolismoRESUMO
Porcine epidemic diarrhoea (PED) caused by porcine epidemic diarrhoea virus (PEDV) has led to significant economic losses in the swine industry worldwide. Histone Cluster 2, H2BE (HIST2H2BE), the main protein component in chromatin, has been proposed to play a key role in apoptosis. However, the relationship between H2BE and PEDV remains unclear. In this study, H2BE was shown to bind and interact with PEDV nonstructural protein 9 (Nsp9) via immunoprecipitation-mass spectrometry (IP-MS). Next, we verified the interaction of Nsp9 with H2BE by immunoprecipitation and immunofluorescence. H2BE colocalized with Nsp9 in the cytoplasm and nuclei. PEDV Nsp9 upregulated the expression of H2BE by inhibiting the expression of IRX1. We demonstrated that overexpression of H2BE significantly promoted PEDV replication, whereas knockdown of H2BE by small interfering RNA (siRNA) inhibited PEDV replication. Overexpression of H2BE led to significantly inhibited GRP78 expression, phosphorylated PERK (p-PERK), phosphorylated eIF2 (p-eIF2), phosphorylated IRE1 (p-IRE1), and phosphorylated JNK (p-JNK); negatively regulated CHOP and Bax expression and caspase-9 and caspase-3 cleavage; and promoted Bcl-2 production. Knocking down H2BE exerted the opposite effects. Furthermore, we found that after deletion of amino acids 1-28, H2BE did not promote PEDV replication. In conclusion, these studies revealed the mechanism by which H2BE is associated with ER stress-mediated apoptosis to regulate PEDV replication. Nsp9 upregulates H2BE. H2BE plays a role in inhibiting apoptosis and thus facilitating viral replication, which depends on the N-terminal region of H2BE (amino acids 1-28). These findings provide a reference for host-PEDV interactions and offer the possibility for developing strategies for PEDV decontamination and prevention.
Assuntos
Infecções por Coronavirus , Vírus da Diarreia Epidêmica Suína , Doenças dos Suínos , Animais , Suínos , Chlorocebus aethiops , Vírus da Diarreia Epidêmica Suína/fisiologia , Fator de Iniciação 2 em Eucariotos , Proteínas não Estruturais Virais/genética , Replicação Viral , Proteínas Serina-Treonina Quinases , Aminoácidos , Estresse do Retículo Endoplasmático , Apoptose , Infecções por Coronavirus/veterinária , Células VeroRESUMO
INTRODUCTION: Inflammatory pain is a significant global clinical challenge that involves both unpleasant sensory and emotional experiences. The treatment of pain is imminent, and we are committed to seeking new analgesics for pain relief. Transcrocetin meglumine salt (TCMS), a saffron metabolite derived from the crocin apocarotenoids, has exhibited the ability to cross the blood-brain barrier and exert neuroprotective effects. In this study, we aimed to investigate whether TCMS could ameliorate complete Freund's adjuvant (CFA)-induced inflammatory pain in mice and elucidate its underlying mechanisms. METHODS: Here, we established an inflammatory pain model in mice by injecting CFA into the left hind paw. Three days later, we administered intraperitoneal injections of TCMS (10 mg/kg) or saline to the animals. We examined mechanical allodynia, thermal hypersensitivity, and anxiety behavior. Furthermore, the activation of glial cells and proinflammatory cytokines in the spinal cord were detected. RESULTS: Our results showed that TCMS significantly reversed the mechanical allodynia and thermal hypersensitivity in the CFA-injected mice. Furthermore, TCMS administration effectively inhibited the activation of microglia and astrocytes in the spinal cord induced by CFA. Additionally, TCMS suppressed the production and release of spinal proinflammatory cytokines, including TNF-α, IL-1ß, and IL-6, in CFA-injected mice. CONCLUSION: Taken together, our findings demonstrate that TCMS holds promise as an innovative analgesic due to its ability to ameliorate inflammatory reactions.
Assuntos
Citocinas , Hiperalgesia , Camundongos , Animais , Citocinas/metabolismo , Hiperalgesia/induzido quimicamente , Hiperalgesia/tratamento farmacológico , Adjuvante de Freund/toxicidade , Meglumina/efeitos adversos , Dor/tratamento farmacológico , Neuroglia/metabolismo , Inflamação/induzido quimicamente , Inflamação/tratamento farmacológico , Inflamação/metabolismo , Medula EspinalRESUMO
Ferroptosis is a novel form of programmed cell death characterized by the accumulation of iron-dependent lipid peroxides, which causes membrane injury. Under the catalysis of iron ions, cells deficient in glutathione peroxidase (GPX4) cannot preserve the balance in lipid oxidative metabolism, and the buildup of reactive oxygen species on the membrane lipids leads to cell death. An increasing body of evidence suggests that ferroptosis plays a significant role in the development and occurrence of cardiovascular diseases. In this paper, we mainly elaborated on the molecular mechanisms regulating ferroptosis and its impact on cardiovascular disease to lay the groundwork for future studies on the prophylaxis and treatment of this patient population.
Assuntos
Doenças Cardiovasculares , Ferroptose , Humanos , Peroxidação de Lipídeos , Apoptose , Ferro/metabolismoRESUMO
Glutathione S-transferases (GSTs) are one of the three detoxification enzyme families. The constitutive and inducible overexpression of GSTs genes plays an important role in insecticide resistance. Previous study showed that malathion resistance was polygenic, and elevated GSTs activity was one of the important factor participating in malathion resistance of Bactrocera dorsalis (Hendel), a serious economic pest worldwide. BdGSTd5 overexpression was inducible upon exposure to malathion. However, the involvement of BdGSTd5 in malathion resistance has not been clarified. In this study, we found that BdGSTd5 sequence harbored the conserved region of delta class GSTs, which were overexpressed in malathion resistant strain of B. dorsalis compared to malathion susceptible strain. The highest mRNA expression level of BdGSTd5 was found in 1-day-old adult, and the levels decreased with aging. The dsBdGSTd5 injection effectively silenced (73.4% reduction) the expression of BdGSTd5 and caused significant increase in susceptibility to malathion with a cumulative mortality increasing of 13.5% at 72 h post malathion treatment (p < 0.05). Cytotoxicity assay demonstrated that BdGSTd5 was capable of malathion detoxification. Molecular docking analysis further indicated the interactive potential of BdGSTd5 with malathion and its toxic oxide malaoxon. The recombinant BdGSTd5 exhibited glutathione-conjugating activity toward 1-chloro-2, 4-dinitrobenzene and malathion and malaoxon metabolic capacity with significant reduction (p < 0.05) of the peak areas by 90.0% and 73.1%, respectively. Taken together, the overexpressed BdGSTd5 contributes to malathion metabolism and resistance, which detoxify the malathion in B. dorsalis via directly depleting malathion and malaoxon.
Assuntos
Inseticidas , Tephritidae , Animais , Malation/toxicidade , Inseticidas/farmacologia , Glutationa Transferase/genética , Glutationa Transferase/metabolismo , Simulação de Acoplamento Molecular , Tephritidae/genética , Resistência a Inseticidas/genéticaRESUMO
Carboxylesterases (CarEs) are a multifunctional superfamily of enzymes and play an important role in detoxification of various insecticides in insects. The oriental fruit fly, Bactrocera dorsalis, is one of the most destructive agricultural pests and has developed different degrees of resistance to organophosphates in field. However, the involvement of BdCarEs in tolerance or resistance to other alternative insecticides are still unclear. In the present study, 33 BdCarEs genes were identified based on the genome database of B. dorsalis. Phylogenetic analysis demonstrated that they were classified into nine clades, with abundance of α-esterases. Meanwhile, the sequence characterization and the chromosome distribution were also analyzed. The spatiotemporal expression analysis of BdCarEs genes suggested that the diversity of potential function in different physiological processes. With the exception of BdCarE21, all BdCarEs genes responded to at least one insecticide exposure, and BdCarE20 was found to be up-regulated after exposure to all five tested insecticides individually. Eight BdCarEs genes were overexpressed in MR strain when compared to that in SS strain. Subsequently, knockdown the expression of representative BdCarEs genes significantly increased the susceptibility of the oriental fruit fly to corresponding insecticides, which indicated that the tested BdCarEs genes contributed to one or multiple insecticide detoxification. These findings provide valuable insights into the potential role in respond to tolerance or resistance to insecticides with different mode of action, and will facilitate development of efficiency management strategy for B. dorsalis.
Assuntos
Inseticidas , Tephritidae , Animais , Inseticidas/toxicidade , Carboxilesterase/genética , Malation/farmacologia , Filogenia , Resistência a Inseticidas/genética , Tephritidae/genéticaRESUMO
Infection with the porcine epidemic diarrhoea virus (PEDV) causes severe enteric disease in suckling piglets, causing massive economic losses in the swine industry worldwide. Tripartite motif-containing 56 (TRIM56) has been shown to augment type I IFN response, but whether it affects PEDV replication remains uncharacterized. Here we investigated the role of TRIM56 in Marc-145 cells during PEDV infection. We found that TRIM56 expression was upregulated in cells infected with PEDV. Overexpression of TRIM56 effectively reduced PEDV replication, while knockdown of TRIM56 resulted in increased viral replication. TRIM56 overexpression significantly increased the phosphorylation of IRF3 and NF-κB P65, and enhanced the IFN-ß antiviral response, while silencing TRIM56 did not affect IRF3 activation. TRIM56 overexpression increased the protein level of TRAF3, the component of the TLR3 pathway, thereby significantly activating downstream IRF3 and NF-κB signalling. We demonstrated that TRIM56 overexpression inhibited PEDV replication and upregulated expression of IFN-ß, IFN-stimulated genes (ISGs) and chemokines in a dose-dependent manner. Moreover, truncations of the RING domain, N-terminal domain or C-terminal portion on TRIM56 were unable to induce IFN-ß expression and failed to restrict PEDV replication. Together, our results suggested that TRIM56 was upregulated in Marc-145 cells in response to PEDV infection. Overexpression of TRIM56 inhibited PEDV replication by positively regulating the TLR3-mediated antiviral signalling pathway. These findings provide evidence that TRIM56 plays a positive role in the innate immune response during PEDV infection.
Assuntos
Infecções por Coronavirus , Vírus da Diarreia Epidêmica Suína , Animais , Antivirais , Interferon beta/genética , Interferon beta/metabolismo , NF-kappa B/genética , NF-kappa B/metabolismo , Suínos , Fator 3 Associado a Receptor de TNF/genética , Fator 3 Associado a Receptor de TNF/metabolismo , Receptor 3 Toll-Like/genética , Receptor 3 Toll-Like/metabolismo , Replicação ViralRESUMO
BACKGROUND: Mitochondrial autophagy maintains mitochondrial function and cellular homeostasis and plays a critical role in the pathological process of cerebral ischemia/reperfusion injury (CIRI). Whether Gypenoside XVII (GP17) has regulatory effects on mitochondrial autophagy against CIRI remains unclear. The purpose of this study was to investigate the pharmacodynamic effects and mechanisms of GP17 on mitochondrial autophagy after CIRI. METHODS: A rat middle cerebral artery occlusion/reperfusion (MCAO/R) model was used to assess the effects of GP17 against CIRI and to explore the underlying mechanisms. An oxygen-glucose deprivation/reoxygenation (OGD/R) cell model was used to verify the ameliorative effects on mitochondrial damage and to probe the autophagy pathways involved in combating neural injuries. RESULTS: The in vivo results showed that GP17 significantly improved mitochondrial metabolic functions and suppressed cerebral ischemic injury, possibly via the autophagy pathway. Further research revealed that GP17 maintains moderate activation of autophagy under ischemic and OGD conditions, producing neuroprotective effects against CIRI, and that the regulation of mitochondrial autophagy is associated with crosstalk between the SIRT1-FOXO3A and Hif1a-BNIP3 signalling pathway that is partially eliminated by the specific inhibitors AGK-7 and 2-ME. CONCLUSION: Overall, this work offers new insights into the mechanisms by which GP17 protects against CIRI and highlights the potential of therapy with Notoginseng leaf triterpene compounds as a novel clinical strategy in humans.
Assuntos
Isquemia Encefálica , Traumatismo por Reperfusão , Humanos , Ratos , Animais , Sirtuína 1 , Traumatismo por Reperfusão/complicações , Autofagia , Infarto da Artéria Cerebral Média/complicações , Mitocôndrias/metabolismo , Isquemia Encefálica/complicações , Apoptose , Proteínas de Membrana , Proteínas Proto-Oncogênicas , Subunidade alfa do Fator 1 Induzível por HipóxiaRESUMO
Sevoflurane inhalation is prone to initiate cognitive deficits in infants. The early growth response-2 (Egr-2) gene is DNA-binding transcription factor, involving in cognitive function. In this study we explored the molecular mechanisms underlying the vulnerability to cognitive deficits after sevoflurane administration. Six-day-old (young) and 6-week-old (early adult) mice received anesthesia with 3% sevoflurane for 2 h daily for 3 days. We showed that multiple exposures of sevoflurane induced significant learning ability impairment in young but not early adult mice, assessed in Morris water maze test on postnatal days 65. The integrated differential expression analysis revealed distinct transcription responses of Egr family members in the hippocampus of the young and early adult mice after sevoflurane administration. Particularly, Egr2 was significantly upregulated after sevoflurane exposure only in young mice. Microinjection of Egr2 shRNA recombinant adeno-associated virus into the dentate gyrus alleviated sevoflurane-induced cognitive deficits, and abolished sevoflurane-induced dendritic spins loss and BDNF downregulation in young mice. On the contrary, microinjection of the Egr2 overexpression virus in the dentate gyrus aggravated learning ability impairment induced by sevoflurane in young mice but not early adult mice. Furthermore, we revealed that sevoflurane markedly upregulated the nuclear factors of activated T-cells NFATC1 and NFATC2 in young mice, which were involved in Egr2 regulation. In conclusion, Egr2 serves as a critical factor for age-dependent vulnerability to sevoflurane-induced cognitive deficits.
Assuntos
Anestésicos Inalatórios , Disfunção Cognitiva , Proteína 2 de Resposta de Crescimento Precoce , Éteres Metílicos , Animais , Camundongos , Anestésicos Inalatórios/toxicidade , Animais Recém-Nascidos , Cognição , Disfunção Cognitiva/induzido quimicamente , Proteína 2 de Resposta de Crescimento Precoce/genética , Proteína 2 de Resposta de Crescimento Precoce/metabolismo , Hipocampo/metabolismo , Aprendizagem em Labirinto , Sevoflurano/efeitos adversosRESUMO
CONTEXT: Folium Ginkgo extract and tetramethylpyrazine sodium chloride injection (Xingxiong injection) is a compound preparation commonly used for treating cerebral ischaemia/reperfusion injury in ischaemic stroke in China. However, its potential mechanisms on ischaemic stroke remain unknown. OBJECTIVE: This study explores the potential mechanisms of Xingxiong injection in vivo or in vitro. MATERIALS AND METHODS: Sprague-Dawley (SD) rats were randomly assigned to five groups: the sham (normal saline), the model (normal saline) and the Xingxiong injection groups (12.5, 25 or 50 mL/kg). The rats were subjected to 2 h of middle cerebral artery occlusion (MCAO) followed by reperfusion for 14 d. Xingxiong injection was administered via intraperitoneal (i.p.) injection immediately after ischaemia induction for 14 d. Afterwards, rats were sacrificed at 14 d induced by administration of Xingxiong injection. RESULTS: Xingxiong injection significantly reduces infarct volume (23%) and neurological deficit scores (93%) compared with the MCAO/R group. Additionally, Xingxiong injection inhibits the loss in mitochondrial membrane potential (43%) and reduces caspase-3 level (44%), decreases NOX (41%), protein carbonyl (29%), 4-HNE (40%) and 8-OhdG (41%) levels, inhibits the expression of inflammatory factors, such as TNF-α (26%), IL-1ß (34%), IL-6 (39%), MCP-1 (36%), CD11a (41%) and ICAM-1 (43%). Moreover, Xingxiong injection can increase p-Akt/Akt (35%) and Nrf2 (47%) protein expression and inhibit NLRP3 (42%) protein expression. CONCLUSIONS: Xingxiong injection prevents cerebral ischaemia/reperfusion injury via activating the Akt/Nrf2 pathway and inhibiting NLRP3 inflammasome. These findings provide experimental evidence for clinical use of drugs in the treatment of ischaemic stroke.
Assuntos
Isquemia Encefálica/tratamento farmacológico , Medicamentos de Ervas Chinesas/farmacologia , Ginkgo biloba/química , Traumatismo por Reperfusão/tratamento farmacológico , Animais , Relação Dose-Resposta a Droga , Medicamentos de Ervas Chinesas/administração & dosagem , Medicamentos de Ervas Chinesas/química , Infarto da Artéria Cerebral Média , Inflamassomos/metabolismo , AVC Isquêmico/tratamento farmacológico , Masculino , Fator 2 Relacionado a NF-E2/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Pirazinas/química , Distribuição Aleatória , Ratos , Ratos Sprague-DawleyRESUMO
BACKGROUND: Breast cancer (BC) has a marked tendency to spread to the bone, resulting in significant skeletal complications and mortality. Recently, circular RNAs (circRNAs) have been reported to contribute to cancer initiation and progression. However, the function and mechanism of circRNAs in BC bone metastasis (BC-BM) remain largely unknown. METHODS: Bone-metastatic circRNAs were screened using circRNAs deep sequencing and validated using in situ hybridization in BC tissues with or without bone metastasis. The role of circIKBKB in inducing bone pre-metastatic niche formation and bone metastasis was determined using osteoclastogenesis, immunofluorescence and bone resorption pit assays. The mechanism underlying circIKBKB-mediated activation of NF-κB/bone remodeling factors signaling and EIF4A3-induced circIKBKB were investigated using RNA pull-down, luciferase reporter, chromatin isolation by RNA purification and enzyme-linked immunosorbent assays. RESULTS: We identified that a novel circRNA, circIKBKB, was upregulated significantly in bone-metastatic BC tissues. Overexpressing circIKBKB enhanced the capability of BC cells to induce formation of bone pre-metastatic niche dramatically by promoting osteoclastogenesis in vivo and in vitro. Mechanically, circIKBKB activated NF-κB pathway via promoting IKKß-mediated IκBα phosphorylation, inhibiting IκBα feedback loop and facilitating NF-κB to the promoters of multiple bone remodeling factors. Moreover, EIF4A3, acted acting as a pre-mRNA splicing factor, promoted cyclization of circIKBKB by directly binding to the circIKBKB flanking region. Importantly, treatment with inhibitor eIF4A3-IN-2 reduced circIKBKB expression and inhibited breast cancer bone metastasis effectively. CONCLUSION: We revealed a plausible mechanism for circIKBKB-mediated NF-κB hyperactivation in bone-metastatic BC, which might represent a potential strategy to treat breast cancer bone metastasis.
Assuntos
Neoplasias Ósseas/secundário , Remodelação Óssea/genética , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Quinase I-kappa B/genética , NF-kappa B/metabolismo , RNA Circular , Transdução de Sinais , Animais , Neoplasias Ósseas/diagnóstico , Neoplasias Ósseas/metabolismo , Linhagem Celular Tumoral , RNA Helicases DEAD-box/metabolismo , Modelos Animais de Doenças , Fator de Iniciação 4A em Eucariotos/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Camundongos , Modelos Biológicos , Inibidor de NF-kappaB alfa/metabolismo , Osteogênese/genética , Osteólise , Transdução de Sinais/efeitos dos fármacos , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Chronic inflammation pain is a debilitating disease, and its mechanism still remains poorly understood. This study attempted to illuminate the metabolic mechanism of chronic inflammation pain induced by complete Freund's adjuvant (CFA) injection, especially at spinal level. The chronic inflammation pain model was established by CFA administration. Behavioral testing including mechanical allodynia and thermal hyperalgesia was performed. Meanwhile, a liquid chromatography-mass spectrometry-based metabolomics approach was applied to analyze potential metabolic biomarkers. The orthogonal partial least squares discrimination analysis mode was employed for determining metabolic changes, and a western blot was performed to detect the protein expression change. The results showed that 27 metabolites showed obviously abnormal expression and seven metabolic pathways were significantly enriched, comprising aminoacyl-tRNA biosynthesis, arginine and proline metabolism, histidine metabolism, purine metabolism, phenylalanine, tyrosine and tryptophan biosynthesis, glutathione metabolism, and phenylalanine metabolism. Meanwhile, the results showed that the expression of arginase I and nitric oxide levels were elevated in the CFA group compared with the control group, while the argininosuccinate synthetase and argininosuccinatelyase proteins were not significantly different between the groups. These findings demonstrate that metabolic changes of the spinal cord may be implicated in neurotransmitter release and pain conductivity following CFA administration.
Assuntos
Adjuvante de Freund , Inflamação , Metaboloma/efeitos dos fármacos , Metabolômica/métodos , Dor , Animais , Comportamento Animal/efeitos dos fármacos , Modelos Animais de Doenças , Adjuvante de Freund/efeitos adversos , Adjuvante de Freund/farmacologia , Hiperalgesia , Inflamação/induzido quimicamente , Inflamação/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Óxido Nítrico/análise , Óxido Nítrico/metabolismo , Dor/induzido quimicamente , Dor/metabolismo , Medula Espinal/química , Medula Espinal/metabolismoRESUMO
Host factors play multiple essential roles in the replication and pathogenesis of mammalian neurotropic viruses. However, the cellular proteins of the central nervous system (CNS) involved in avian neurotropic virus infection have not been completely elucidated. Here, we employed a gene microarray to identify caspase recruitment domain-containing protein 11 (CARD11), a lymphoma-associated scaffold protein presenting brain-specific upregulated expression in a virulent neurotropic Newcastle disease virus (NDV)-infected natural host. Chicken primary neuronal cells infected with NDV appeared slightly syncytial and died quickly. CARD11 overexpression inhibited viral replication and delayed cytopathic effects; conversely, depletion of CARD11 enhanced viral replication and cytopathic effects in chicken primary neuronal cells. The inhibition of viral replication by CARD11 could not be blocked with CARD11-Bcl10-MALT1 (CBM) signalosome and NF-κB signaling inhibitors. CARD11 was found to interact directly with the viral phosphoprotein (P) through its CC1 domain and the X domain of P; this X domain also mediated the interaction between P and the viral large polymerase protein (L). The CARD11 CC1 domain and L competitively bound to P via the X domain that hindered the P-L interaction of the viral ribonucleoprotein (RNP) complex, resulting in a reduction of viral polymerase activity in a minigenome assay and inhibition of viral replication. Animal experiments further revealed that CARD11 contributed to viral replication inhibition and neuropathology in infected chicken brains. Taken together, our findings identify CARD11 as a brain-specific antiviral factor of NDV infection in avian species.IMPORTANCE Newcastle disease virus (NDV) substantially impacts the poultry industry worldwide and causes viral encephalitis and neurological disorders leading to brain damage, paralysis, and death. The mechanism of interaction between this neurotropic virus and the avian central nervous system (CNS) is largely unknown. Here, we report that host protein CARD11 presented brain-specific upregulated expression that inhibited NDV replication, which was not due to CARD11-Bcl10-MALT1 (CBM) complex-triggered activation of its downstream signaling pathways. The inhibitory mechanism of viral replication is through the CARD11 CC1 domain, and the viral large polymerase protein (L) competitively interacts with the X domain of the viral phosphoprotein (P), which hampers the P-L interaction, suppressing the viral polymerase activity and viral replication. An in vivo study indicated that CARD11 alleviated neuropathological lesions and reduced viral replication in chicken brains. These results provide insight into the interaction between NDV infection and the host defense in the CNS and a potential antiviral target for viral neural diseases.
Assuntos
Proteínas Adaptadoras de Sinalização CARD/antagonistas & inibidores , Guanilato Ciclase/antagonistas & inibidores , Neurônios/virologia , Vírus da Doença de Newcastle/efeitos dos fármacos , Replicação Viral/efeitos dos fármacos , Animais , Proteína 10 de Linfoma CCL de Células B/metabolismo , Ligação Competitiva , Encéfalo/patologia , Encéfalo/virologia , Proteínas Adaptadoras de Sinalização CARD/genética , Proteínas Adaptadoras de Sinalização CARD/metabolismo , Galinhas , Técnicas de Silenciamento de Genes , Guanilato Ciclase/genética , Guanilato Ciclase/metabolismo , Humanos , Proteína de Translocação 1 do Linfoma de Tecido Linfoide Associado à Mucosa/metabolismo , Doença de Newcastle/virologia , Receptor EphB2 , Transdução de SinaisRESUMO
Molybdenum complexes are versatile and efficient for liquid phase olefin epoxidation reactions. Rational design of catalysts is critical to achieve high atom efficiency during epoxidation processes. Although liquid phase epoxidation has been a popular topic for decades, three key issues, (a) rational control of morphology of molybdenum nanoparticles, (b) manipulating metal-support interaction and (c) altering electronic configuration at molybdenum center remains unsolved in this area. Therefore, in this paper, we have critically revised recent research progress on heterogeneous molybdenum catalysts for facile liquid phase olefin epoxidation in terms of catalyst synthesis, surface characterization, catalytic performance and structure-function relationship. Furthermore, plausible reaction mechanisms will be systematically discussed with the aim to provide insights into fundamental understanding on novel epoxidation chemistry.
RESUMO
Porcine growth hormone (pGH) is a class of peptide hormones secreted from the pituitary gland, which can significantly improve growth and feed utilization of pigs. However, it is unstable and volatile in vitro. It needs to be encapsulated in liposomes when feeding livestock, whose high cost greatly limits its application in pig industry. Therefore we attempted to express pGH as intracellular soluble protein in Pichia pastoris and feed these yeasts with partial wall-breaking for swine, which could release directly pGH in intestine tract in case of being degraded in intestinal tract with low cost. In order to improve the intracellular soluble expression of pGH protein in Pichia pastoris and stability in vitro, we optimized the pGH gene, and screened molecular chaperones from E. coli and Pichia pastoris respectively for co-expressing with pGH. In addition, we had also explored conditions of mechanical crushing and fermentation. The results showed that the expression of intracellular soluble pGH protein was significantly increased after gene optimized and co-expressed with Ssa1-Sis1 chaperone from Pichia pastoris. Meanwhile, the optimal conditions of partial wall-breaking and fermentation of Pichia pastoris were confirmed, the data showed that the intracellular expression of the optimized pGH protein co-expressed with Ssa1-Sis1 could reach 340 mg/L with optimal conditions of partial wall-breaking and fermentation. Animal experiments verified that the optimized pGH protein co-expression with Ssa1-Sis1 had the best promoting effects on the growth of piglets. Our study demonstrated that Ssa1-Sis1 could enhance the intracellular soluble expression of pGH protein in Pichia pastoris and that partial wall-breaking of yeast could prevent pGH from degradation in vitro, release targetedly in the intestine and play its biological function effectively. Our study could provide a new idea to cut the cost effectively, establishing a theoretical basis for the clinic application of unstable substances in vitro.
Assuntos
Proteínas Fúngicas/metabolismo , Hormônio do Crescimento/biossíntese , Chaperonas Moleculares/metabolismo , Pichia/metabolismo , Suínos/crescimento & desenvolvimento , Animais , Clonagem Molecular , Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Fermentação , Pichia/genética , Proteínas Recombinantes/biossínteseRESUMO
Retinol-binding protein 4 (RBP4) is known as a highly conserved adipokine for immune activation. Aeromonas hydrophila (A. hydrophila) is the most common zoonotic pathogen in aquaculture, which causes serious economic losses to aquaculture, especially to bighead carp (Hypophthalmichthys nobilis, H. nobilis) and silver carp (Hypophthalmichthys molitrix, H. molitrix). Recent studies along with our previous findings have shown that synthetic oligodeoxynucleotides containing CpG motifs (CpG ODN) can play a good role in aquatic animals against infection. In order to clarify the relationship between CpG ODN and RBP4 under A. hydrophila infection, firstly, full-length RBP4 cDNAs from H. nobilis and H. molitrix were cloned. And characteristics of RBP4, including sequence and structure, tissue distribution and genetic evolution were analyzed. In addition, mRNA expression levels of RBP4, cytokine, toll-like receptors (TLRs), morbidity and survival rates of H. nobilis and H. molitrix were observed post CpG ODN immunization or following challenge. The results indicated that hn/hm_RBP4 (RBP4 genes obtained from H. nobilis and H. molitrix) had the highest homology with Megalobrama amblycephala. Distribution data showed that the expression level of hn_RBP4 mRNA was higher than that of hm_RBP4. After CpG ODN immunization followed by A.hydrophila challenge, significantly higher survival was observed in both carps, together with up-regulated RBP4 expression. Meanwhile, hn/hm_IL-1ß level was relatively flat (and decreased), hn/hm_IFN-γ, hn/hm_TLR4 and hn/hm_TLR9 levels increased significantly, but hn/hm_STRA6 showed no significant change, compared with control. Moreover, CpG ODN immunization could induce stronger immune protective responses (higher IFN-γ/gentle IL-1ß level and lower morbidity/higher survival rate) against A. hydrophila in H. nobilis, along with higher RBP4 level, when compared with that in H. molitrix. These results demonstrated that RBP4 was well involved in the immune protection of CpG ODN. Based on the results, we speculated that in the case of A. hydrophila infection, TLR9 signaling pathway was activated by CpG ODN. Subsequently, CpG ODN up-regulated RBP4, and RBP4 activated TLR4 signaling pathway. Then TLR4 and TLR9 synergistically improved the anti-infection responses. Our findings have good significance for improving resistance to pathogen infection in freshwater fish.
Assuntos
Carpas/genética , Carpas/microbiologia , Infecções por Bactérias Gram-Negativas/veterinária , Imunização/veterinária , Oligodesoxirribonucleotídeos/administração & dosagem , Proteínas Celulares de Ligação ao Retinol/genética , Aeromonas hydrophila/patogenicidade , Animais , Carpas/imunologia , DNA Complementar , Infecções por Bactérias Gram-Negativas/imunologia , Infecções por Bactérias Gram-Negativas/prevenção & controle , Oligodesoxirribonucleotídeos/imunologia , Proteínas Celulares de Ligação ao Retinol/química , Proteínas Celulares de Ligação ao Retinol/imunologia , Regulação para CimaRESUMO
Up to now, many previous reports have emphasized that Annexins (ANX) family played an important role in immune responses. Aeromonas hydrophila (A. hydrophila), the most common zoonotic pathogenic bacteria of yellow catfish (Pelteobagrus fulvidraco), can cause serious economic loss, especially to yellow catfish with high economic value. In our previous work, we demonstrated that synthetic oligodeoxynucleotides containing CpG motifs (CpG ODN) owned powerful immunostimulatory activity. However, the relationship among Pelteobagrus fulvidraco Annexins (Pf_ANX), CpG ODN and A. hydrophila is unknown. Therefore, we cloned Pf_ANX1-6 genes and analyzed its sequences, structures, genetic evolution, post-translation modifications (PTMs), Ca2+ ion binding sites and tissue distribution to reveal the relevance. In addition, we investigated the responses of ANXA1-6 and cytokines in intestine and spleen as well as morbidity/survival rate of fish post CpG ODN immunization and/or A. hydrophila infection. The results showed that compared with challenge alone (challenge-CK) group, the CpG immunization following challenge (CpG-challenge) group displayed relatively flat IL-1ß level throughout in both organs. Meanwhile, the expression of IFN-γ and morbidity/survival rate of fish in CpG-challenge group showed a great improvement compared with the challenge-CK group. Our results indicated that CpG ODN could improve morbidity/survival by up-regulating Pf_ANXA 1, 2 and 5 in the intestine and spleen to ameliorate inflammatory responses and promote anti-infective responses. Our findings offer some important insights into ANX related to the immunity of fish infection and lay a theoretical basis for the prevention and treatment of fish infections.