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INTRODUCTION: Activated hepatic stellate cells (HSCs) are the primary effector cells in hepatic fibrosis, over depositing extracellular matrix (ECM) proteins. Our previous work found oridonin analog CYD0682 attenuates proliferation, Transforming Growth Factor ß (TGFß)-induced signaling, and ECM production in immortalized HSCs. The underlying mechanism behind these reductions is unclear. The Signal Transduction and Activator of Transcription 3 (STAT3) pathway plays a central role in HSC activation and has been found to be overexpressed in models of hepatic injury. In this study, we will examine the effect of CYD0682 on STAT3 signaling. METHODS: Immortalized human (LX-2) and rat (HSC-T6) HSC lines were treated with CYD0682 or Tanespimycin (17-AAG) with or without TGF-ß. Nuclear and cytosolic proteins were extracted. Protein expression was analyzed with Western blot. DNA binding activity was assessed with STAT3 DNA Binding ELISA. Cell viability was assessed with Alamar blue assay. RESULTS: CYD0682 treatment inhibited STAT3 phosphorylation at tyrosine 705 in a dose-dependent manner in LX-2 and HSC-T6 cells. STAT3 DNA binding activity and STAT3 regulated protein c-myc were significantly decreased by CYD0682. Notably, TGFß-induced STAT3 phosphorylation and ECM protein expression were inhibited by CYD0682. STAT3 is reported to be a Heat Shock Protein 90 (HSP90) client protein. Notably, CYD0682 attenuated the expression of endogenous STAT3 and other HSP90 client proteins FAK, IKKα, AKT and CDK9. HSP90 specific inhibitor 17-AAG suppressed endogenous and TGFß-induced STAT3 phosphorylation and ECM protein production. CONCLUSIONS: CYD0682 attenuates endogenous and TGFß-induced STAT3 activation and ECM production via an HSP90 dependent pathway in HSCs. Further study of this pathway may present new targets for therapeutic intervention in hepatic fibrosis.
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Benzoquinonas , Diterpenos de Tipo Kaurano , Proteínas HSP90 de Choque Térmico , Células Estrelladas Hepáticas , Factor de Transcripción STAT3 , Transducción de Señal , Células Estrelladas Hepáticas/efectos de los fármacos , Células Estrelladas Hepáticas/metabolismo , Factor de Transcripción STAT3/metabolismo , Humanos , Ratas , Animales , Diterpenos de Tipo Kaurano/farmacología , Transducción de Señal/efectos de los fármacos , Proteínas HSP90 de Choque Térmico/metabolismo , Proteínas HSP90 de Choque Térmico/antagonistas & inhibidores , Benzoquinonas/farmacología , Factor de Crecimiento Transformador beta/metabolismo , Línea Celular , Fosforilación/efectos de los fármacos , Lactamas Macrocíclicas/farmacología , Cirrosis Hepática/tratamiento farmacológico , Cirrosis Hepática/metabolismo , Cirrosis Hepática/patologíaRESUMEN
BACKGROUND: Fulminant myocarditis (FM) is a common life-threatening disease in pediatric patients, which can result in sudden cardiac arrest (CA). Whether prolonged cardiopulmonary resuscitation (CPR) is beneficial to FM induced CA is unknown. CASE PRESENTATION: We reported the case of an 8-year-old child with FM. At 14:49 of the day after admission, the ECG monitoring indicated ventricular flutter. The patient was immediately given continuous external cardiac compression. Electric cardioversion (energy 30J) and electric defibrillation (energy 50 J, 100 J, 100 J) were given. Continuous chest compression was conducted until extracorporeal membrane oxygenation (ECMO) successfully placed at 19:30 P.M. The total duration of CPR was 291 min. Nine days later, the ECMO was removed; and 29 days later, the patient was discharged from hospital. In the three years of follow-up, the boy showed a full recovery without neurological sequela. At present, his daily activities have returned to normal and his academic performance at school is excellent. CONCLUSIONS: Prolonged CPR can be used in FM induced in-hospital CA in pediatric patients, especially during preparation for ECMO after the failure of standard resuscitation measures.
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Reanimación Cardiopulmonar , Oxigenación por Membrana Extracorpórea , Paro Cardíaco , Miocarditis , Niño , Estudios de Seguimiento , Paro Cardíaco/etiología , Paro Cardíaco/terapia , Humanos , Masculino , Miocarditis/complicaciones , Miocarditis/terapia , Resultado del TratamientoRESUMEN
Intercropping with guava (Psidium guajava L.) can assist with the management of Asian citrus psyllid (ACP, Diaphorina citri Kuwayama), the insect vector of the huanglongbing pathogen, in citrus orchards. Sulfur volatiles have a repellent activity and physiological effects, as well as being important components of guava volatiles. In this study, we tested whether the sulfur volatiles emitted by guava plants play a role in plant-plant communications and trigger anti-herbivore activities against ACP in sweet orange plants (Citrus sinensis L. Osbeck). Real-time determination using a proton-transfer-reaction mass spectrometer (PTR-MS) showed that guava plants continuously release methanethiol, dimethyl sulfide (DMS), and dimethyl disulfide (DMDS), and the contents increased rapidly after mechanical damage. The exposure of orange plants to DMDS resulted in the suppression of the developmental performance of ACP. The differential elevation of salicylic acid (SA) levels; the expression of phenylalanine ammonia lyase (PAL), salicylate-O-methyl transferase (SMT), and pathogenesis-related (PR1) genes; the activities of defense-related enzymes PAL, polyphenol oxidase (PPO), and peroxidase (POD); and the total polyphenol content were observed in DMDS-exposed orange plants. The emission of volatiles including myrcene, nonanal, decanal, and methyl salicylate (MeSA) was increased. In addition, phenylpropanoid and flavonoid biosynthesis, and aromatic amino acid (such as phenylalanine, tyrosine, and tryptophan) metabolic pathways were induced. Altogether, our results indicated that DMDS from guava plants can activate defense responses in eavesdropping orange plants and boost their herbivore resistance to ACP, which suggests the possibility of using DMDS as a novel approach for the management of ACP in citrus orchards.
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Citrus sinensis , Citrus , Hemípteros , Psidium , Animales , Catecol Oxidasa/metabolismo , Citrus/metabolismo , Citrus sinensis/genética , Disulfuros , Hemípteros/fisiología , Peroxidasas/metabolismo , Fenilalanina/farmacología , Fenilanina Amoníaco-Liasa/metabolismo , Enfermedades de las Plantas/genética , Polifenoles/farmacología , Protones , Psidium/química , Ácido Salicílico/metabolismo , Azufre/metabolismo , Transferasas/metabolismo , Triptófano/metabolismo , Tirosina/metabolismoRESUMEN
BACKGROUND: Hepatic fibrosis is wound-healing response that is the result of hepatic stellate cell (HSC) activation and subsequent excess extracellular matrix deposition. HSCs can be activated by a variety of inflammatory stimuli as well as through the signal transducer and activator of transcription 3 (STAT3) pathway. HJC0416 is a novel, orally bioavailable small-molecule inhibitor of STAT3 that was developed by our team using a fragment-based drug design approach. Previously, our team has shown that HJC0416 has antifibrogenic effects in activated HSCs. Recently, increasing evidence suggests that nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) plays an important role in the activation of HSCs. In the present study, we examined the role of NF-κB inhibition of HSC activation by HJC0416. METHODS: LX-2 (human) and HSC-T6 (rat) cell lines were used. Expression levels of extracellular proteins, NF-κB and STAT3 expression and DNA binding, and inflammatory cytokine levels were determined using western blot, ELISA, and immunofluorescence assay. RESULTS: HJC0416 decreased cell viability in a dose-dependent manner in both cell lines and arrested the cell cycle at the S phase. Increased apoptosis was seen in LX-2 cells through Yo-Pro-1 and propidium iodide immunofluorescent stating. HJC0416 significantly decreased expression of fibronectin and collagen I as well as markedly decreased α-SMA and laminin. HJC0416 inhibited the STAT3 pathway by decreasing phosphorylation of STAT3, as well as signal transduction pathway activation. Notably, HJC0416 also inhibited the classic and alternative pathways of NF-κB activation. HJC0416 inhibited LPS-induced p65 nuclear translocation and DNA binding, as well as prevented phosphorylation and degradation of inhibitory protein IκBα. HJC0416 also prevented phosphorylation of serine residue 536 on p65. CONCLUSIONS: HJC0416, an inhibitor of STAT3, was found to have antifibrogenic properties in activated hepatic stellate cell lines. In addition, HJC0416 was found to inhibit the NF-κB pathway. Owing to this double effect, HJC0416 demonstrates promise for in vivo experimentation as an antifibrosis treatment.
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Benzamidas/uso terapéutico , Células Estrelladas Hepáticas/efectos de los fármacos , Cirrosis Hepática/prevención & control , FN-kappa B/metabolismo , Factor de Transcripción STAT3/metabolismo , Tiofenos/uso terapéutico , Animales , Benzamidas/farmacología , Ciclo Celular/efectos de los fármacos , Línea Celular , Evaluación Preclínica de Medicamentos , Células Estrelladas Hepáticas/metabolismo , Humanos , Ratas , Tiofenos/farmacologíaRESUMEN
Electroacupuncture has been shown to effectively reduce chronic pain in patients with nerve injury. The underlying mechanisms are not well understood. Accumulated evidence suggests that purinergic P2X3 receptors (P2X3Rs) in dorsal root ganglion neurons play a major role in mediating chronic pain associated with nerve injury. The aim of this study is to determine if electroacupuncture stimulation alters P2X3R activity in dorsal root ganglia to produce analgesia under neuropathic pain condition. Peripheral neuropathy was produced by ligation of the left lumbar 5 (L5) spinal nerve in rats. Low-frequency (2 Hz) electrical stimulation was applied to ipsilateral ST36 and BL60 acupoints in rats. The P2X3R agonist (α,ß-meATP)-induced flinch responses were reduced after electroacupuncture treatment. Western analyses showed that P2X3R expression was upregulated in nerve-uninjured lumbar 4 (L4) dorsal root ganglion neurons ipsilateral to the spinal nerve ligation. Electroacupuncture-stimulation reversed the upregulation. In nerve-injured L5 dorsal root ganglia, P2X3R expression was substantially reduced. Electroacupuncture had no effect on the reduction. We also determined the injury state of P2X3R expressing dorsal root ganglion neurons using the neuronal injury marker, activating transcription factor 3 (ATF3). Immunohistochemical assay showed that in L4 dorsal root ganglia, almost all P2X3Rs were expressed in uninjured (ATF3-) neurons. Spinal nerve ligation increased the expression of P2X3Rs. Electroacupuncture reduced the increase in P2X3R expression without affecting the percentage of ATF + neurons. In ipsilateral L5 dorsal root ganglion neurons, spinal nerve ligation reduced the percentage of P2X3R + neurons and markedly increased the percentage of ATF3 + cells. Almost all of P2X3Rs were expressed in damaged (ATF3+) neurons. Electroacupuncture had no effect on spinal nerve ligation-induced changes in the percentage of P2X3R or percentage of ATF3 + cells in L5 dorsal root ganglia. These observations led us to conclude that electroacupuncture effectively reduces injury-induced chronic pain by selectively reducing the expression of P2X3Rs in nerve-uninjured L4 dorsal root ganglion neurons.
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Regulación hacia Abajo , Electroacupuntura , Ganglios Espinales/metabolismo , Receptores Purinérgicos P2X3/metabolismo , Nervios Espinales/metabolismo , Factor de Transcripción Activador 3/metabolismo , Adenosina Trifosfato/análogos & derivados , Animales , Ganglios Espinales/patología , Hiperalgesia/patología , Ligadura , Vértebras Lumbares/patología , Masculino , Neuronas/patología , Ratas Sprague-DawleyRESUMEN
Itol A, a novel isoryanodane diterpene derived from Itoa orientalis Hemsl., has potent activities against insect pests. This study was conducted to determine the contact toxicity and biochemical effects of itol A on the Nilaparvata lugens. After macropterous females of N. lugens were exposed to itol A from 0.5 to 24â¯h, the mortality and poisoning symptoms were measured. Effects of itol A on the major enzymes activity and oxidative stress level were assessed in dose-response (with LD10-LD70 at 24â¯h) and time-course (with LD50 at 0.5-24â¯h) experiments for the potential toxicity mechanisms. Based on the results, the mortality of N. lugens showed significant dose- and time-dependent effects, with the 24-h LD50 value was 0.58⯵g/insect. The symptoms of excitation, convulsion and paralysis were also observed. However, acetylcholinesterases (AChE) activity was not altered after itol A treatment compared to control. Na+/K+-ATPases, Ca2+-ATPases, Ca2+/Mg2+-ATPases, glutathione S-transferases (GSTs), cytochrome P450 monooxygenases (P450s), superoxide dismutases (SOD) and catalases (CAT) activities were significantly reduced in dose-response and time-course experiments. While acid phosphatases (ACP) and glutathione peroxidases (GPX) activities were significantly increased. We further revealed that itol A exposure resulted in the decrease of GSH/GSSG (reduced to oxidized glutathione) ratio and the increase of hydrogen peroxide (H2O2) and malondialdehyde (MDA) levels in both experiments. The results indicated that the inhibition of Na+/K+-ATPases, Ca2+-ATPases, Ca2+/Mg2+-ATPases, GSTs, P450s, SOD and CAT activities and the induction of oxidative stress was one of the potential biochemical mechanisms of itol A against N. lugens.
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Diterpenos/toxicidad , Inhibidores Enzimáticos/toxicidad , Hemípteros/efectos de los fármacos , Insecticidas/toxicidad , Fosfatasa Ácida/antagonistas & inhibidores , Adenosina Trifosfatasas/antagonistas & inhibidores , Animales , Diterpenos/química , Inhibidores Enzimáticos/química , Femenino , Glutatión/metabolismo , Glutatión Transferasa/antagonistas & inhibidores , Hemípteros/metabolismo , Insecticidas/química , Dosificación Letal Mediana , Malondialdehído/metabolismo , Oxidorreductasas/antagonistas & inhibidores , SalicaceaeRESUMEN
Abstract: Primary sensory neurons are responsible for transmitting sensory information from the peripheral to the central nervous system. Their responses to incoming stimulation become greatly enhanced and prolonged following inflammation, giving rise to exaggerated nociceptive responses and chronic pain. The inflammatory mediator, prostaglandin E2 (PGE2), released from the inflamed tissue surrounding the terminals of sensory neurons contributes to the abnormal pain responses. PGE2 acts on G protein-coupled EP receptors to activate adenylyl cyclase, which catalyzes the conversion of adenosine triphosphate to cyclic adenosine 3',5'-monophosphate (cAMP). Under normal conditions, cAMP activates primarily protein kinase A. After inflammation, cAMP also activates the exchange proteins activated by cAMP (Epacs) to produce exaggerated PGE2-mediated hyperalgesia. The role of cAMP-Epac signaling in the generation of hypersensitivity is the topic of this review.
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AMP Cíclico/metabolismo , Dinoprostona/metabolismo , Hiperalgesia/metabolismo , Nociceptores/metabolismo , Células Receptoras Sensoriales/metabolismo , Animales , Factores de Intercambio de Guanina Nucleótido/metabolismo , HumanosRESUMEN
Sensitization of purinergic P2X3 receptors (P2X3Rs) contributes to the production of exaggerated nociceptive responses following inflammatory injury. We showed previously that prostaglandin E2 (PGE2) potentiates P2X3R-mediated ATP currents in dorsal root ganglion neurons isolated from both control and complete Freund's adjuvant-induced inflamed rats. PGE2 potentiation of ATP currents depends only on PKA signaling in control neurons, but it depends on both PKA and PKC signaling in inflamed neurons. We further found that inflammation evokes an increase in exchange proteins directly activated by cAMP (Epacs) in dorsal root ganglions. This increase promotes the activation of PKC to produce a much enhanced PGE2 effect on ATP currents and to elicit Epac-dependent flinch nocifensive behavioral responses in complete Freund's adjuvant rats. The link between Epac-PKC signaling and P2X3R sensitization remains unexplored. Here, we show that the activation of Epacs promotes the expression of phosphorylated PKC and leads to an increase in the cytoskeleton, F-actin, expression at the cell perimeter. Depolymerization of F-actin blocks PGE2-enhanced ATP currents and inhibits P2X3R-mediated nocifensive responses after inflammation. Thus, F-actin is dynamically involved in the Epac-PKC-dependent P2X3R sensitization. Furthermore, Epacs induce a PKC-dependent increase in the membrane expression of P2X3Rs. This increase is abolished by F-actin depolymerization, suggesting that F-actin mediates Epac-PKC signaling of P2X3R membrane expression. Thus, after inflammation, an Epac-PKC dependent increase in F-actin in dorsal root ganglion neurons enhances the membrane expression of P2X3Rs to bring about sensitization of P2X3Rs and abnormal pain behaviors.
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Actinas/metabolismo , Ganglios Espinales/metabolismo , Ganglios Espinales/patología , Factores de Intercambio de Guanina Nucleótido/metabolismo , Inflamación/patología , Proteína Quinasa C/metabolismo , Receptores Purinérgicos P2X3/metabolismo , Transducción de Señal , Adenosina Trifosfato/metabolismo , Animales , Membrana Celular/efectos de los fármacos , Membrana Celular/metabolismo , AMP Cíclico/análogos & derivados , AMP Cíclico/farmacología , Citocalasina D/farmacología , Dinoprostona/farmacología , Adyuvante de Freund , Ganglios Espinales/efectos de los fármacos , Hiperalgesia/patología , Inflamación/metabolismo , Masculino , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Neuronas/patología , Ratas Sprague-Dawley , Transducción de Señal/efectos de los fármacos , Acetato de Tetradecanoilforbol/farmacología , Tionucleótidos/farmacologíaRESUMEN
Bitter taste receptors (TAS2Rs) Tas2r108 gene possesses a high abundance in mouse kidney; however, the biological functions of Tas2r108 encoded receptor TAS2Rs member 4 (TAS2R4) are still unknown. In the present study, we found that mouse TAS2R4 (mTAS2R4) signaling was inactivated in chronic high glucose-stimulated mouse podocyte cell line MPC, evidenced by the decreased protein expressions of mTAS2R4 and phospholipase C ß2 (PLCß2), a key downstream molecule of mTAS2R4 signaling. Nonetheless, agonism of mTAS2R4 by quinine recovered mTAS2R4 and PLCß2 levels, and increased podocyte cell viability as well as protein expressions of ZO-1 and nephrin, biomarkers of podocyte slit diaphragm, in high glucose-cultured MPC cells. However, blockage of mTAS2R4 signaling with mTAS2R4 blockers γ-aminobutyric acid and abscisic acid, a Gßγ inhibitor Gallein, or a PLCß2 inhibitor U73122 all abolished the effects of quinine on NLRP3 inflammasome and p-NF-κB p65 as well as the functional podocyte proteins in MPC cells in a high glucose condition. Furthermore, knockdown of mTAS2R4 with lentivirus-carrying Tas2r108 shRNA also ablated the effect of quinine on the key molecules of the above inflammatory signalings and podocyte functions in high glucose-cultured MPC cells. In summary, we demonstrated that activation of TAS2R4 signaling alleviated the podocyte injury caused by chronic high glucose, and inhibition of NF-κB p65 and NLRP3 inflammasome mediated the protective effects of TAS2R4 activation on podocytes. Moreover, activation of TAS2R4 signaling could be an important strategy for prevention and treatment of diabetic kidney disease.
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Glucosa , Podocitos , Receptores Acoplados a Proteínas G , Transducción de Señal , Podocitos/metabolismo , Podocitos/efectos de los fármacos , Podocitos/patología , Animales , Ratones , Receptores Acoplados a Proteínas G/metabolismo , Receptores Acoplados a Proteínas G/genética , Glucosa/toxicidad , Glucosa/farmacología , Transducción de Señal/efectos de los fármacos , Transducción de Señal/fisiología , Línea CelularRESUMEN
Studies of the structural organization and functions of the cell body of a neuron (soma) and its surrounding satellite glial cells (SGCs) in sensory ganglia have led to the realization that SGCs actively participate in the information processing of sensory signals from afferent terminals to the spinal cord. SGCs use a variety ways to communicate with each other and with their enwrapped soma. Changes in this communication under injurious conditions often lead to abnormal pain conditions. "What are the mechanisms underlying the neuronal soma and SGC communication in sensory ganglia?" and "how do tissue or nerve injuries affect the communication?" are the main questions addressed in this review.
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Comunicación Celular/fisiología , Ganglios Sensoriales/citología , Neuroglía/fisiología , Neuronas/fisiología , AnimalesRESUMEN
BACKGROUND: Phosphorylation sites in the C-terminus of mu-opioid receptors (MORs) are known to play critical roles in the receptor functions. Our understanding of their participation in opioid analgesia is mostly based on studies of opioid effects on mutant receptors expressed in in vitro preparations, including cell lines, isolated neurons and brain slices. The behavioral consequences of the mutation have not been fully explored due to the complexity in studies of mutant receptors in vivo. To facilitate the determination of the contribution of phosphorylation sites in MOR to opioid-induced analgesic behaviors, we expressed mutant and wild-type human MORs (hMORs) in sensory dorsal root ganglion (DRG) neurons, a major site for nociceptive (pain) signaling and determined morphine- and the full MOR agonist, DAMGO,-induced effects on heat-induced hyperalgesic behaviors and potassium current (IK) desensitization in these rats. FINDINGS: A mutant hMOR DNA with the putative phosphorylation threonine site at position 394 replaced by an alanine (T394A), i.e., hMOR-T, or a plasmid containing wild type hMOR (as a positive control) was intrathecally delivered. The plasmid containing GFP or saline was used as the negative control. To limit the expression of exogenous DNA to neurons of DRGs, a neuron-specific promoter was included in the plasmid. Following a plasmid injection, hMOR-T or hMOR receptors were expressed in small and medium DRG neurons. Compared with saline or GFP rats, the analgesic potency of morphine was increased to a similar extent in hMOR-T and hMOR rats. Morphine induced minimum IK desensitization in both rat groups. In contrast, DAMGO increased analgesic potency and elicited IK desensitization to a significantly less extent in hMOR-T than in hMOR rats. The development and extent of acute and chronic tolerance induced by repeated morphine or DAMGO applications were not altered by the T394A mutation. CONCLUSIONS: These results indicate that phosphorylation of T394 plays a critical role in determining the potency of DAMGO-induced analgesia and IK desensitization, but has limited effect on morphine-induced responses. On the other hand, the mutation contributes minimally to both DAMGO- and morphine-induced behavioral tolerance. Furthermore, the study shows that plasmid gene delivery of mutant receptors to DRG neurons is a useful strategy to explore nociceptive behavioral consequences of the mutation.
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Analgésicos Opioides/farmacología , Tolerancia a Medicamentos/genética , Receptores Opioides mu/agonistas , Receptores Opioides mu/metabolismo , Células Receptoras Sensoriales/efectos de los fármacos , Células Receptoras Sensoriales/metabolismo , Animales , Encefalina Ala(2)-MeFe(4)-Gli(5)/farmacología , Humanos , Masculino , Fosforilación , Ratas , Ratas Sprague-Dawley , Receptores Opioides mu/genéticaRESUMEN
Oxidative stress is involved in the development and progression of disease. Because sodium aescinate has been reported to have immunity enhancing and antioxidative effects, we investigated its activity by employing a hepatocellular carcinoma (HCC) mouse model. Sixty BALB/c mice were randomly divided into four groups, including a 1.4 mg/kg treated group (n = 15), a 2.8 mg/kg treated group (n = 15), an untreated hepatocellular carcinoma control group (n = 15) and a normal control group (n = 15). After H22 cells were cultured for one week, we collected 2 × 106 cells and injected them subcutaneously as 0.2 mL cell suspensions in sterile saline into the right shoulder region of every mouse. The animals were monitored for changes in activity, physical condition and body weight during the experiment. The next day after injection of H22 cells, animals in these test groups received one intraperitoneal injection of drug or physiological saline for 13 days. Results showed that in the sodium aescinate injection liquid (SAIL)-treated HCC mice, serum interleukin-1 beta (IL-1ß), interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-α), interferon-gamma (IFN-γ), Gamma-glutamyltransferase (γ-GT), alanine transaminase (ALT), aspartate transaminase (AST) and alkaline phosphatase (ALP) levels were significantly decreased compared with normal control mice. In addition, treatment with sodium aescinate injection liquid significantly decreased blood and liver malondialdehyde (MDA) levels, increased glutathione (GSH) levels, and antioxidant enzyme [superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSH-Px)] activities in a dose-dependent manner. We conclude that sodium aescinate injection liquid can decrease oxidative injury and enhance immunity functions in HCC mice.
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Antioxidantes/farmacología , Carcinoma Hepatocelular/tratamiento farmacológico , Factores Inmunológicos/farmacología , Neoplasias Hepáticas/tratamiento farmacológico , Estrés Oxidativo/efectos de los fármacos , Compuestos de Sodio/farmacología , Alanina Transaminasa/metabolismo , Animales , Aspartato Aminotransferasas/metabolismo , Humanos , Interferón gamma/metabolismo , Interleucina-1beta/metabolismo , Interleucina-6/análisis , Ratones , Ratones Endogámicos BALB C , Distribución Aleatoria , Compuestos de Sodio/administración & dosificación , Superóxido Dismutasa/metabolismo , Células Tumorales Cultivadas , Factor de Necrosis Tumoral alfa/análisis , Ensayos Antitumor por Modelo de Xenoinjerto , gamma-Glutamiltransferasa/metabolismoRESUMEN
Intercropping can reduce agricultural pest incidence and represents an important sustainable alternative to conventional pest control methods. Citrus intercropped with guava (Psidium guajava L.) has a lower incidence of Asian citrus psyllid (ACP, Diaphorina citri Kuwayama) and huanglongbing disease (HLB), but the mechanisms are still unknown. In this study, we tested whether volatile organic compounds (VOCs) emitted by guava plants play a role in plant-plant communications and trigger defense responses in sweet orange (Citrus sinensis L. Osbeck) in the laboratory. The results showed that the behavioral preference and developmental performance of ACP on citrus plants that were exposed to guava VOCs were suppressed. The expression of defense-related pathways involved in early signaling, jasmonate (JA) biosynthesis, protease inhibitor (PI), terpenoid, phenylpropanoid, and flavonoid biosynthesis was induced in guava VOC-exposed citrus plants. Headspace analysis revealed that guava plants constitutively emit high levels of (E)-ß-caryophyllene and (E)-4,8-dimethyl-1,3,7-nonatriene (DMNT), which can induce the accumulation of JA and promote stronger defense responses of citrus to ACP feeding. In addition, exposure to guava VOCs also increased the indirect defense of citrus by attracting the parasitic wasp Tamarixia radiata. Together, our findings indicate that citrus plants can eavesdrop on the VOC cues emitted by neighboring intact guava plants to boost their JA-dependent anti-herbivore activities. The knowledge gained from this study will provide mechanisms underlying citrus-guava intercropping for the ecological management of insect pests.
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Purinergic ionotropic P2X7 receptors (P2X7Rs) are closely associated with excitotoxicity and nociception. Inhibition of P2X7R activation has been considered as a potentially useful strategy to improve recovery from spinal cord injury and reduce inflammatory damage to trauma. The physiological functions of P2X7Rs, however, are poorly understood, even though such information is essential for making the P2X7R an effective therapeutic target. We show here that P2X7Rs in satellite cells of dorsal root ganglia tonically inhibit the expression of P2X3Rs in neurons. Reducing P2X7R expression using siRNA or blocking P2X7R activity by antagonists elicits P2X3R up-regulation, increases the activity of sensory neurons responding to painful stimuli, and evokes abnormal nociceptive behaviors in rats. Thus, contrary to the notion that P2X7R activation is cytotoxic, P2X7Rs in satellite cells play a crucial role in maintaining proper P2X3R expression in dorsal root ganglia. Studying the mechanism underlying the P2X7R-P2X3R control, we demonstrate that activation of P2X7Rs evokes ATP release from satellite cells. ATP in turn stimulates P2Y1 receptors in neurons. P2Y1 receptor activation appears to be necessary and sufficient for the inhibitory control of P2X3R expression. We further determine the roles of the P2X7R-P2Y1-P2X3R inhibitory control under injurious conditions. Activation of the inhibitory control effectively prevents the development of allodynia and increases the potency of systemically administered P2X7R agonists in inflamed rats. Thus, direct blocking P2X7Rs, as proposed before, may not be the best strategy for reducing pain or lessening neuronal degeneration because it also disrupts the protective function of P2X7Rs.
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Regulación hacia Abajo/genética , Neuroglía/metabolismo , Dolor , Receptores Purinérgicos P2/genética , Receptores Purinérgicos P2/fisiología , Células Satélites Perineuronales/metabolismo , Animales , Ganglios Espinales , Masculino , Ratas , Ratas Sprague-Dawley , Receptores Purinérgicos P2/metabolismo , Receptores Purinérgicos P2X3 , Receptores Purinérgicos P2X7 , Células Receptoras Sensoriales/químicaRESUMEN
INTRODUCTION: Severe burns lead to marked impairment of gastrointestinal motility, such as delayed gastric emptying and small and large intestinal ileus. However, the cellular mechanism of these pathologic changes remains largely unknown. METHODS: Male Sprague Dawley rats approximately 3 months old and weighing 300-350 g were randomized to either a 60% total body surface area full-thickness scald burn or sham procedure and were sacrificed 24 h after the procedure. Gastric emptying, gastric antrum contractility ileal smooth muscle contractility, and colonic contractility were measured. Muscularis externa was isolated from the ileal segment to prepare smooth muscle protein extracts for Western blot analysis. RESULTS: Compared with sham controls, the baseline rhythmic contractile activities of the antral, ileal, and colonic smooth muscle strips were impaired in the burned rats. Simultaneously, our data showed that ileal muscularis ECM proteins fibronectin and laminin were significantly up-regulated in burned rats compared with sham rats. TGF-ß signaling is an important stimulating factor for ECM protein expression. Our results revealed that TGF-ß signaling was activated in the ileal muscle of burned rats evidenced by the activation of Smad2/3 expression and phosphorylation. In addition, the total and phosphorylated AKT, which is an important downstream factor of ECM signaling in smooth muscle cells, was also up-regulated in burned rats' ileal muscle. Notably, these changes were not seen in the colonic or gastric tissues. CONCLUSION: Deposition of fibrosis-related proteins after severe burn is contributors to decreased small intestinal motility.
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Quemaduras/metabolismo , Proteínas de la Matriz Extracelular/metabolismo , Íleon/metabolismo , Seudoobstrucción Intestinal/metabolismo , Contracción Muscular/fisiología , Músculo Liso/metabolismo , Animales , Quemaduras/complicaciones , Quemaduras/fisiopatología , Colon/metabolismo , Colon/fisiopatología , Modelos Animales de Enfermedad , Proteínas de la Matriz Extracelular/biosíntesis , Fibronectinas/biosíntesis , Fibronectinas/metabolismo , Fibrosis/etiología , Fibrosis/metabolismo , Fibrosis/fisiopatología , Vaciamiento Gástrico/fisiología , Motilidad Gastrointestinal/fisiología , Íleon/fisiopatología , Ileus/metabolismo , Ileus/fisiopatología , Inflamación/etiología , Inflamación/metabolismo , Inflamación/fisiopatología , Seudoobstrucción Intestinal/etiología , Seudoobstrucción Intestinal/fisiopatología , Laminina/biosíntesis , Laminina/metabolismo , Masculino , Músculo Liso/fisiopatología , Fosforilación , Antro Pilórico/metabolismo , Antro Pilórico/fisiopatología , Ratas , Ratas Sprague-Dawley , Transducción de Señal/fisiología , Estómago/fisiopatologíaRESUMEN
Pluripotent or multipotent stem cells isolated from human embryos or adult central nervous system (CNS) may provide new neurons to ameliorate neural disorders. A major obstacle, however, is that the majority of such cells do not differentiate into neurons when grafted into non-neurogenic areas of the adult CNS. Here we report a new in vitro priming procedure that generates a nearly pure population of neurons from fetal human neural stem cells (hNSCs) transplanted into adult rat CNS. Furthermore, the grafted cells differentiated by acquiring a cholinergic phenotype in a region-specific manner. This technology may advance stem cell-based therapy to replace lost neurons in neural injury or neurodegenerative disorders.
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Trasplante de Tejido Encefálico/métodos , Técnicas de Cultivo de Célula/métodos , Supervivencia de Injerto/fisiología , Enfermedades Neurodegenerativas/terapia , Neuronas/citología , Trasplante de Células Madre/métodos , Células Madre/citología , Acetilcolina/metabolismo , Animales , Células Cultivadas , Colina O-Acetiltransferasa/metabolismo , Feto , Supervivencia de Injerto/efectos de los fármacos , Heparina/farmacología , Humanos , Inmunohistoquímica , Laminina/farmacología , Masculino , Factores de Crecimiento Nervioso/farmacología , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Fenotipo , Ratas , Células Madre/efectos de los fármacos , Células Madre/metabolismoRESUMEN
The exchange proteins activated by cAMP (Epacs) have been shown to play important roles in producing inflammation-induced nociception. Transient receptor potential vanilloid type 1 (TRPV1) is a major receptor processing thermal and chemosensitive nociceptive information. The role of Epacs in modulating the activity of TRPV1 has yet to be determined. Studying the effect of complete Freund adjuvant (CFA)-induced inflammation on capsaicin-activated TRPV1 nociceptive responses in dorsal root ganglia (DRG), we found that CFA produced a large increase in capsaicin-induced responses. The increase was inhibited by Epac1 and Epac2 antagonists. Thus, activation of Epacs is critical in producing enhancement in TRPV1-mediated responses under inflammatory conditions. In addition, the inflammation-induced enhancement of TRPV1 responses was blocked by PKCα and PKCε inhibitors, suggesting the essential roles of these PKCs in enhancing TRPV1 responses. To determine the mechanism underlying the Epac actions on TRPV1, we studied the effects of the Epac activator, 8-(4-chlorophenylthio)-2-O-methyl-cAMP (CPT), on capsaicin-induced nociceptive behavioral responses, capsaicin-activated currents, expression and membrane trafficking of PKC and TRPV1 in DRG. CPT was found to enhance capsaicin-induced nociception and ionic currents. The enhancement was inhibited by PKCα and PKCε inhibitors. In addition, CPT increased the expression of phosphorylated PKCα (pPKCα) and membrane TRPV1 expression in DRG. Studying the colocalization of TRPV1 and pPKCα or pPKCε in DRG slices prepared from CFA-treated rats, we found that pPKCα or pPKCε expressed with TRPV1 in different-sized neurons to exert differential influences on TRPV1 activity. Thus, Epac-PKC signaling is critically important in producing inflammation-induced potentiation of TRPV1 functions.
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Acetilcisteína/análogos & derivados , Eritromicina/análogos & derivados , Hiperalgesia/fisiopatología , Inflamación/enzimología , Proteína Quinasa C-epsilon/metabolismo , Transducción de Señal/fisiología , Canales Catiónicos TRPV/metabolismo , Acetilcisteína/metabolismo , Acetilcisteína/farmacología , Potenciales de Acción/efectos de los fármacos , Potenciales de Acción/fisiología , Animales , Biotinilación , Capsaicina/toxicidad , Células Cultivadas , Modelos Animales de Enfermedad , Inhibidores Enzimáticos/farmacología , Eritromicina/metabolismo , Eritromicina/farmacología , Adyuvante de Freund/toxicidad , Ganglios Espinales/citología , Hiperalgesia/patología , Inflamación/inducido químicamente , Masculino , Neuronas/efectos de los fármacos , Neuronas/fisiología , Técnicas de Placa-Clamp , Proteína Quinasa C-alfa/metabolismo , Ratas , Ratas Sprague-Dawley , Receptores Purinérgicos P2X4/metabolismoRESUMEN
Due to complex pest control scenarios and the needs of agricultural production, different neonicotinoids may be used in certain agricultural applications. Consequently, honeybees may be exposed to these substances through distribution throughout plant tissues via the vascular system through several pathways, such as surface water, the exudates excreted from plants, and air pollution via drift of dust as well as contaminated pollen and nectar. In the current study, the single and combined toxicity of clothianidin, dinotefuran, and thiamethoxam to honeybees was examined after 48 h exposure by the acute oral method and combination index (CI)-isobologram equation. At the 48 h interval, our results showed that 1) the order of toxicities for the single insecticides was ranked as clothianidin > thiamethoxam > dinotefuran and that 2) all binary and ternary combinations showed synergism or additive effect at the effect (fa) 0.5. Therefore, our results not only provided meaningful guidelines in evaluating the safety risk of the mixtures of the three neonicotinoids towards honeybees but also suggested that there is a significant interest in the study of mixture toxicities of neonicotinoids against honeybees because risk assessment of neonicotinoids against honeybees conducted only in individual insecticides may underestimate the realistic toxicity.
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Guanidinas/toxicidad , Insecticidas/toxicidad , Nitrocompuestos/toxicidad , Oxazinas/toxicidad , Tiazoles/toxicidad , Animales , Abejas , Neonicotinoides , Plantas , Polen , Investigación , Tiametoxam , Pruebas de ToxicidadRESUMEN
Sensitization of purinergic P2X3 receptors (P2X3Rs) is a major mechanism contributing to injury-induced exaggerated pain responses. We showed in a previous study that cyclic adenosine monophosphate (cAMP)-dependent guanine nucleotide exchange factor 1 (Epac1) in rat sensory dorsal root ganglia (DRGs) is upregulated after inflammatory injury, and it plays a critical role in P2X3R sensitization by activating protein kinase C epsilon (PKCε) inside the cells. protein kinase C epsilon has been established as the major PKC isoform mediating injury-induced hyperalgesic responses. On the other hand, the role of PKCα in receptor sensitization was seldom considered. Here, we studied the participation of PKCα in Epac signaling in P2X3R-mediated hyperalgesia. The expression of both Epac1 and Epac2 and the level of cAMP in DRGs are greatly enhanced after complete Freund adjuvant (CFA)-induced inflammation. The expression of phosphorylated PKCα is also upregulated. Complete Freund adjuvant (CFA)-induced P2X3R-mediated hyperalgesia is not only blocked by Epac antagonists but also by the classical PKC isoform inhibitors, Go6976, and PKCα-siRNA. These CFA effects are mimicked by the application of the Epac agonist, 8-(4-chlorophenylthio)-2 -O-methyl-cAMP (CPT), in control rats, further confirming the involvement of Epacs. Because the application of Go6976 prior to CPT still reduces CPT-induced hyperalgesia, PKCα is downstream of Epacs to mediate the enhancement of P2X3R responses in DRGs. The pattern of translocation of PKCα inside DRG neurons in response to CPT or CFA stimulation is distinct from that of PKCε. Thus, in contrast to prevalent view, PKCα also plays an essential role in producing complex inflammation-induced receptor-mediated hyperalgesia.
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Factores de Intercambio de Guanina Nucleótido/metabolismo , Hiperalgesia/metabolismo , Inflamación/metabolismo , Proteína Quinasa C-alfa/metabolismo , Receptores Purinérgicos P2X3/metabolismo , Transducción de Señal/fisiología , Animales , Carbazoles/farmacología , Células Cultivadas , AMP Cíclico/metabolismo , Inhibidores Enzimáticos/farmacología , Ganglios Espinales/efectos de los fármacos , Ganglios Espinales/metabolismo , Factores de Intercambio de Guanina Nucleótido/antagonistas & inhibidores , Masculino , Proteína Quinasa C-alfa/antagonistas & inhibidores , Ratas , Ratas Sprague-Dawley , Transducción de Señal/efectos de los fármacosRESUMEN
P2X3 and P2X2/3 receptors in dorsal root ganglia (DRG) appear to participate in producing nociceptive responses after nerve injury. However, the mechanisms underlying the receptor-mediated nociception in the neuropathic state remain unclear. Using spared nerve injury (SNI) rats, we found that allodynic and nocifensive (flinch) behavioral responses developed after injury can be reversed by P2X receptor antagonists, indicating an involvement of P2X receptors. Immunocytochemical studies revealed that P2X3 receptors are expressed in small and medium but rarely in large DRG neurons of both normal and SNI rats. Thus, contrary to the conventional view that only large A beta cells mediate allodynia, small and medium cells are intimately involved in P2X3 receptor-mediated allodynia. Measuring ATP levels in the subcutaneous space of the rat paw, we showed that ATP release does not change after SNI. On the other hand, the P2X receptor agonist, alpha beta-methylene ATP produces 3.5-fold larger flinch responses at a 8.0-fold lower dose. Thus, sensitization of P2X3 receptors rather than a change in ATP release is responsible for the neuropathic pain behaviors. We further demonstrated that sensitization of P2X3 receptors arises from an increase in receptor function. ATP-induced P2X3 receptor-mediated currents in DRG neurons is 2.5-fold larger after SNI. The expression of P2X3 receptors on the cell membrane is significantly enhanced while the total expression of P2X3 receptors remained unchanged. Thus, the enhancement of trafficking of P2X3 receptors is likely an important mechanism contributing to the increase in receptor function after nerve injury.