RESUMEN
Exposure to hypoxia results in the development of pulmonary arterial hypertension (PAH). An increase in the intracellular Ca2+ concentration ([Ca2+]i) in pulmonary artery smooth muscle cells (PASMCs) is a major trigger for pulmonary vasoconstriction and proliferation. This study investigated the mechanism by which KMUP-1, a xanthine derivative with phosphodiesterase inhibitory activity, inhibits hypoxia-induced canonical transient receptor potential channel 1 (TRPC1) protein overexpression and regulates [Ca2+]i through store-operated calcium channels (SOCs). Ex vivo PASMCs were cultured from Sprague-Dawley rats in a modular incubator chamber under 1% O2/5% CO2 for 24 h to elucidate TRPC1 overexpression and observe the Ca2+ release and entry. KMUP-1 (1 µM) inhibited hypoxia-induced TRPC family protein encoded for SOC overexpression, particularly TRPC1. KMUP-1 inhibition of TRPC1 protein was restored by the protein kinase G (PKG) inhibitor KT5823 (1 µM) and the protein kinase A (PKA) inhibitor KT5720 (1 µM). KMUP-1 attenuated protein kinase C (PKC) activator phorbol 12-myristate 13-acetate (PMA, 1 µM)-upregulated TRPC1. We suggest that the effects of KMUP-1 on TRPC1 might involve activating the cyclic guanosine monophosphate (cGMP)/PKG and cyclic adenosine monophosphate (cAMP)/PKA pathways and inhibiting the PKC pathway. We also used Fura 2-acetoxymethyl ester (Fura 2-AM, 5 µM) to measure the stored calcium release from the sarcoplasmic reticulum (SR) and calcium entry through SOCs in hypoxic PASMCs under treatment with thapsigargin (1 µM) and nifedipine (5 µM). In hypoxic conditions, store-operated calcium entry (SOCE) activity was enhanced in PASMCs, and KMUP-1 diminished this activity. In conclusion, KMUP-1 inhibited the expression of TRPC1 protein and the activity of SOC-mediated Ca2+ entry upon SR Ca2+ depletion in hypoxic PASMCs.
RESUMEN
Autophagy facilitates the degradation of organelles and cytoplasmic proteins in a lysosome-dependent manner. It also plays a crucial role in cell damage. Whether loganin affects autophagy in chronic constriction injury (CCI)-induced neuropathic pain remains unclear. We investigated the neuroprotective effect of loganin on the autophagic-lysosomal pathway in the rat CCI model. Sprague-Dawley rats were divided into sham, CCI, sham + loganin, and CCI + loganin. Loganin (5 mg/kg/day) was intraperitoneally injected once daily, and rats were sacrificed on day 7 after CCI. This study focused on the mechanism by which loganin modulates autophagic flux after CCI. CCI enhanced the autophagic marker LC3B-II in the ipsilateral spinal cord. The ubiquitin-binding protein p62 binds to LC3B-II and integrates into autophagosomes, which are degraded by autophagy. CCI caused the accumulation of p62, indicating the interruption of autophagosome turnover. Loganin significantly attenuated the expression of Beclin-1, LC3B-II, and p62. Double immunofluorescence staining was used to confirm that LC3B-II and p62 were reduced by loganin in the spinal microglia and astrocytes. Loganin also lessened the CCI-increased colocalization of both proteins. Enhanced lysosome-associated membrane protein 2 (LAMP2) and pro-cathepsin D (pro-CTSD) in CCI rats were also attenuated by loganin, suggesting that loganin improves impaired lysosomal function and autophagic flux. Loganin also attenuated the CCI-increased apoptosis protein Bax and cleaved caspase-3. Loganin prevents CCI-induced neuropathic pain, which could be attributed to the regulation of neuroinflammation, neuronal autophagy, and associated cell death. These data suggest autophagy could be a potential target for preventing neuropathic pain.
Asunto(s)
Glicósidos Cardíacos , Neuralgia , Animales , Ratas , Autofagia , Constricción , Hiperalgesia/etiología , Hiperalgesia/complicaciones , Glicósidos Iridoides , Neuralgia/tratamiento farmacológico , Neuralgia/etiología , Neuralgia/metabolismo , Ratas Sprague-DawleyRESUMEN
Eugenosedin-A (Eu-A) has been shown to protect against hyperglycemia- and hyperlipidemia-induced metabolic syndrome. We investigated the relationship of KATP channel activities and insulin secretion by Eu-A in vitro in pancreatic ß-cells, and examined the effect of Eu-A on streptozotocin (STZ)/nicotinamide (NA)-induced type 2 diabetes mellitus (T2DM) in vivo. We isolated pancreatic islets from adult male Wistar rats (250-350 g) and identified pancreatic ß-cells by the cell size, capacitance and membrane potential. Perforated patch-clamp and inside-out recordings were used to monitor the membrane potential (current-clamp mode) and channel activity (voltage-clamp mode) of ß-cells. The membrane potential of ß-cells was raised by Eu-A and reversed by the KATP channel activator diazoxide. Eu-A inhibited the KATP channel activity measured at - 60 mV and increased the intracellular calcium concentration ([Ca2+]i), resulting in enhanced insulin secretion. Eu-A also reduced Kir6.2 protein on the cell membrane and scattered in the cytosol under normal glucose conditions (5.6 mM). In our animal study, rats were divided into normal and STZ/NA-induced T2DM groups. Normal rats fed with regular chow were divided into control and control+Eu-A (5 mg/kg/day, i.p.) groups. The STZ/NA-induced diabetic rats fed with a high-fat diet (HFD) were divided into three groups: T2DM, T2DM+Eu-A (5 mg/kg/day, i.p.), and T2DM+glibenclamide (0.5 mg/kg/day, i.p.; a KATP channel inhibitor). Both Eu-A and glibenclamide decreased the rats' blood glucose, prevented weight gain, and enhanced insulin secretion. We found that Eu-A blocked pancreatic ß-cell KATP channels, caused membrane potential depolarization, and stimulated Ca2+ influx, thus increasing insulin secretion. Furthermore, Eu-A decreased blood glucose and increased insulin levels in T2DM rats. These results suggested that Eu-A might have clinical benefits for the control of T2DM and its complications.
Asunto(s)
Diabetes Mellitus Experimental/tratamiento farmacológico , Diabetes Mellitus Tipo 2/tratamiento farmacológico , Hiperglucemia/tratamiento farmacológico , Piperazinas/farmacología , Animales , Glucemia/efectos de los fármacos , Dieta Alta en Grasa , Gliburida/farmacología , Hiperglucemia/etiología , Hipoglucemiantes/farmacología , Secreción de Insulina/efectos de los fármacos , Islotes Pancreáticos/efectos de los fármacos , Islotes Pancreáticos/metabolismo , Canales KATP/metabolismo , Masculino , Obesidad/complicaciones , Ratas , Ratas WistarRESUMEN
Diabetic peripheral neuropathy (DPN) is caused by hyperglycemia, which induces oxidative stress and inflammatory responses that damage nerve tissue. Excessive generation of reactive oxygen species (ROS) and NOD-like receptor protein 3 (NLRP3) inflammasome activation trigger the inflammation and pyroptosis in diabetes. Schwann cell dysfunction further promotes DPN progression. Loganin has been shown to have antioxidant and anti-inflammatory neuroprotective activities. This study evaluated the neuroprotective effect of loganin on high-glucose (25 mM)-induced rat Schwann cell line RSC96 injury, a recognized in vitro cell model of DPN. RSC96 cells were pretreated with loganin (0.1, 1, 10, 25, 50 µM) before exposure to high glucose. Loganin's effects were examined by CCK-8 assay, ROS assay, cell death assay, immunofluorescence staining, quantitative RT-PCR and western blot. High-glucose-treated RSC96 cells sustained cell viability loss, ROS generation, NF-κB nuclear translocation, P2 × 7 purinergic receptor and TXNIP (thioredoxin-interacting protein) expression, NLRP3 inflammasome (NLRP3, ASC, caspase-1) activation, IL-1ß and IL-18 maturation and gasdermin D cleavage. Those effects were reduced by loganin pretreatment. In conclusion, we found that loganin's antioxidant effects prevent RSC96 Schwann cell pyroptosis by inhibiting ROS generation and suppressing NLRP3 inflammasome activation.
Asunto(s)
Glucosa/efectos adversos , Inflamasomas/metabolismo , Iridoides/metabolismo , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Piroptosis/efectos de los fármacos , Células de Schwann/metabolismo , Animales , Humanos , RatasRESUMEN
Valproate (VPA) is a well-known drug for treating epilepsy and mania, but its action in neuropathic pain is unclear. We used a chronic constriction injury (CCI) model to explore whether VPA prevents neuropathic pain-mediated inflammation and neuronal death. Rats were treated with or without VPA. CCI + VPA rats were intraperitoneally injected with VPA (300 mg/kg/day) from postoperative day (POD) 1 to 14. We measured paw withdrawal latency (PWL) and paw withdrawal threshold (PWT) 1 day before surgery and 1, 3, 7, 14 days after CCI and harvested the sciatic nerves (SN), spinal cord (SC) and dorsal root ganglia (DRG) on POD 3, 7, and 14. PWL and PWT were reduced in CCI rats, but increased in CCI + VPA rats on POD 7 and POD 14. VPA lowered CCI-induced inflammatory proteins (pNFκB, iNOS and COX-2), pro-apoptotic proteins (pAKT/AKT and pGSK-3ß/GSK-3ß), proinflammatory cytokines (TNF-α and IL-1ß) and nuclear pNFκB activation in the SN, DRG and SC in CCI rats. COX-2 and pGSK-3 proteins were decreased by VPA on immunofluorescence analysis. VPA attenuated CCI-induced thermal and mechanical pain behaviors in rats in correlation with anti-neuroinflammation action involving reduction of pNFκB/iNOS/COX-2 activation and inhibition of pAKT/pGSK-3ß-mediated neuronal death from injury to peripheral nerves.
Asunto(s)
Antiinflamatorios/farmacología , Neuralgia/fisiopatología , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Fármacos Neuroprotectores/farmacología , Ácido Valproico/farmacología , Animales , Biomarcadores , Muerte Celular/efectos de los fármacos , Citocinas/metabolismo , Modelos Animales de Enfermedad , Glucógeno Sintasa Quinasa 3 beta/metabolismo , Mediadores de Inflamación/metabolismo , FN-kappa B/metabolismo , Neuralgia/tratamiento farmacológico , Neuralgia/etiología , Proteínas Proto-Oncogénicas c-akt/metabolismo , RatasRESUMEN
BACKGROUND: Diabetes mellitus is a metabolic disorder characterized by chronic hyperglycemia accompanied by impaired vascular and endothelial function. Activation of ATP-sensitive potassium (KATP) channels can protect endothelial function against hypertension and hyperglycemia. KMUP-1, a xanthine derivative, has been demonstrated to modulate K+-channel activity in smooth muscles. This study investigated protective mechanisms of KMUP-1 in impaired mesenteric artery (MA) reactivity in streptozotocin (STZ)-induced diabetic rats. METHODS: Rats were divided into three groups: control, STZ (65â¯mg/kg, ip) and STZâ¯+â¯KMUP-1 (5 or 10â¯mg/kg/day, ip). MA reactivity was measured by dual wire myograph. MA smooth muscle cells (MASMCs) were enzymatically dissociated and the KATP currents recorded by a whole-cell patch-clamp technique. RESULTS: STZ decreased MA KATP currents in a time-course dependent manner and achieved steady inhibition at day 14. In the MASMCs of STZ-treated rats, KMUP-1 partially recovered the KATP currents, suggesting that vascular KATP channels were activated by KMUP-1. K+ (80â¯mM KCl)-induced MA contractions in STZ-treated rats were higher than those of control rats. KMUP-1 significantly attenuated STZ-stimulated MA contractions in response to high K+, suggesting that KMUP-1 may partly restore the vascular reactivity of MAs. In addition, STZ decreased the expression of endothelial nitric oxide synthase (eNOS) and this effect was reversed by KMUP-1, suggesting that KMUP-1 could improve STZ-induced vascular endothelial dysfunction. CONCLUSION: KMUP-1 prevents STZ impairment of MA reactivity, eNOS levels and KATP channels, and accordingly protects against vascular dysfunction in diabetic rats.
Asunto(s)
Diabetes Mellitus Experimental/complicaciones , Diabetes Mellitus Experimental/fisiopatología , Canales KATP/metabolismo , Arterias Mesentéricas/fisiopatología , Piperidinas/farmacología , Enfermedades Vasculares/complicaciones , Enfermedades Vasculares/prevención & control , Xantinas/farmacología , Animales , Diabetes Mellitus Experimental/metabolismo , Relación Dosis-Respuesta a Droga , Masculino , Potenciales de la Membrana/fisiología , Arterias Mesentéricas/metabolismo , Miocitos del Músculo Liso/fisiología , Óxido Nítrico Sintasa de Tipo III/biosíntesis , Potasio/farmacología , Ratas , Factores de Tiempo , Enfermedades Vasculares/metabolismo , Enfermedades Vasculares/fisiopatologíaRESUMEN
This study examined the effects of eugenosedin-A (Eu-A) in a streptozotocin (STZ)/nicotinamide-induced rat model of type II diabetes mellitus (T2DM). Six-week-old Sprague-Dawley rats were randomly divided into three groups: (1) RD group, normal rats fed a regular diet (RD), (2) DM group, T2DM rats fed a high-fat diet, and (3) Eu-A group, T2DM rats fed a high fat diet plus oral Eu-A (5 mg/kg/day). After 30 days, the DM group had higher body weight, higher blood glucose and lower insulin levels than the RD group. The DM group also had increased protein expression of glycogen synthase kinase (GSK) in liver and skeletal muscle and decreased protein expression of insulin receptor (IR), insulin receptor substrate-1 (IRS-1), IRS-2, AMP-activated protein kinase (AMPK), glucose transporter-4 (GLUT-4), glucokinase (GCK), and peroxisome proliferator-activated receptor γ (PPAR-γ). STZ/nicotinamide-induced T2DM increased the expression of mitogen-activated protein kinases (MAPKs: p38, ERK, JNK) and inflammatory p65 protein. In the Eu-A treated T2DM rats, however, blood glucose was attenuated and the insulin concentration stimulated. Changes in IR, IRS-1 and IRS-2 proteins as well as AMPK, GLUT-4, GCK, GSK, PPAR-γ, MAPKs, and inflammatory p65 proteins were ameliorated. These results suggested that Eu-A alleviates STZ/nicotinamide-induced hyperglycemia by improving insulin levels and glucose metabolism, and inhibiting the MAPKs- and p65-mediated inflammatory pathway.
Asunto(s)
Diabetes Mellitus Experimental/tratamiento farmacológico , Hiperglucemia/tratamiento farmacológico , Hipoglucemiantes/farmacología , Proteínas Quinasas Activadas por Mitógenos/genética , Piperazinas/farmacología , Animales , Glucemia/metabolismo , Diabetes Mellitus Experimental/inducido químicamente , Diabetes Mellitus Experimental/genética , Diabetes Mellitus Experimental/patología , Dieta Alta en Grasa/efectos adversos , Regulación de la Expresión Génica , Glucoquinasa/genética , Glucoquinasa/metabolismo , Transportador de Glucosa de Tipo 4/genética , Transportador de Glucosa de Tipo 4/metabolismo , Glucógeno Sintasa Quinasas/genética , Glucógeno Sintasa Quinasas/metabolismo , Hiperglucemia/inducido químicamente , Hiperglucemia/genética , Hiperglucemia/patología , Insulina/sangre , Proteínas Sustrato del Receptor de Insulina/genética , Proteínas Sustrato del Receptor de Insulina/metabolismo , Hígado/efectos de los fármacos , Hígado/metabolismo , Hígado/patología , Masculino , Proteínas Quinasas Activadas por Mitógenos/antagonistas & inhibidores , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Músculo Esquelético/efectos de los fármacos , Músculo Esquelético/metabolismo , Músculo Esquelético/patología , Niacinamida , PPAR gamma/genética , PPAR gamma/metabolismo , Ratas , Ratas Sprague-Dawley , Receptor de Insulina/genética , Receptor de Insulina/metabolismo , Transducción de Señal , Estreptozocina , Factor de Transcripción ReIA/genética , Factor de Transcripción ReIA/metabolismoRESUMEN
We have demonstrated that KMUP-1 (7-[2-[4-(2-chlorobenzene)piperazinyl]ethyl]-1,3-dimethylxanthine) blunts monocrotaline-induced pulmonary arterial hypertension by altering Ca2+ sensitivity, K+-channel function, endothelial nitric oxide synthase activity, and RhoA/Rho kinase (ROCK) expression. This study further investigated whether KMUP-1 impedes uridine 5'-triphosphate (UTP)-inhibited delayed rectifying K+ (KDR) current in rat pulmonary arteries involved the RhoA/ROCK signaling. Pulmonary artery smooth muscle cells (PASMCs) were enzymatically dissociated from rat pulmonary arteries. KMUP-1 (30µM) attenuated UTP (30µM)-mediated membrane depolarization and abolished UTP-enhanced cytosolic Ca2+ concentration. Whole-cell patch-clamp electrophysiology was used to monitor KDR currents. A voltage-dependent KDR current was isolated and shown to consist of a 4-aminopyridine (5mM)-sensitive component and an insensitive component. The 4-aminopyridine sensitive KDR current was suppressed by UTP (30µM). The ROCK inhibitor Y27632 (30µM) abolished the ability of UTP to inhibit the KDR current. Like Y27632, KMUP-1 (30µM) similarly abolished UTP-inhibited KDR currents. Superfused protein kinase A and protein kinase G inhibitors (KT5720, 300nM and KT5823, 300nM) did not affect UTP-inhibited KDR currents, but the currents were restored by adding KMUP-1 (30µM) to the superfusate. KMUP-1 reversal of KDR current inhibition by UTP predominantly involves the ROCK inhibition. The results indicate that the RhoA/ROCK signaling pathway plays a key role in eliciting PASMCs depolarization caused by UTP, which would result in pulmonary artery constriction. KMUP-1 blocks UTP-mediated PASMCs depolarization, suggesting that it would prevent abnormal pulmonary vasoconstriction.
Asunto(s)
Canales de Potasio de Tipo Rectificador Tardío/metabolismo , Miocitos del Músculo Liso/metabolismo , Óxido Nítrico/metabolismo , Piperidinas/farmacología , Arteria Pulmonar/citología , Uridina Trifosfato/farmacología , Xantinas/farmacología , Quinasas Asociadas a rho/metabolismo , Proteína de Unión al GTP rhoA/metabolismo , 4-Aminopiridina/farmacología , Amidas/farmacología , Animales , Calcio/metabolismo , Citosol/metabolismo , Regulación hacia Abajo/efectos de los fármacos , Femenino , Activación del Canal Iónico/efectos de los fármacos , Miocitos del Músculo Liso/efectos de los fármacos , Piridinas/farmacología , Ratas Sprague-Dawley , Transducción de Señal/efectos de los fármacosRESUMEN
In this study the effects of low-dose aspirin (5 mg/kg) on adhesion molecule and chemokine expression in a hyperlipidemic rat model. Six-week-old Sprague-Dawley (SD) rats were assigned to two control groups receiving either a regular diet or high-fat diet (HFD) and a treatment group fed HFD with 5 mg/kg aspirin for a 10-week period. Compared with the regular diet control group, the HFD control group had higher body weight, lower levels of high-density lipoprotein, higher concentrations of insulin, triglyceride, total cholesterol, and low-density lipoprotein, but no differences in blood glucose and glycated hemoglobin. The prothrombin time (PT) and activated partial thromboplastin time (aPTT) were clearly shortened in the HFD group. That group also had increased expression of intercellular adhesion molecule-1 (ICAM-1), ICAM-2, ICAM-3, vascular cell adhesion molecule (VCAM), platelet endothelial cell adhesion molecule (PECAM) and P-selectin in platelets and vascular adhesion protein-1 in lymphocyte and in aorta increased expressions of ICAM-1, ICAM-2, ICAM-3, VCAM, PECAM, E-selectin, monocyte chemoattractant protein-1 (MCP-1) and CCR2. The HFD rats also had increased PKCα, IκB kinase α (IKKα), p65, mitogen-activated protein kinases (MAPKs) (p38, c-Jun N-terminal kinases 1, extracellular signal-regulated kinase 1/2), and their phosphorylated forms. Low-dose aspirin improved HFD-induced hyperinsulinemia and hyperlipidemia, recovered PT and aPTT, inhibited upregulation of adhesion molecules and chemokines and reduced expression of PKCα, IKKα, p65, and MAPKs. Low-dose aspirin ameliorates HFD-induced hyperlipidemia and hyperinsulinemia, and prevents HFD-induced expression of adhesion molecules and chemokine formation.
Asunto(s)
Antiinflamatorios no Esteroideos/uso terapéutico , Aspirina/uso terapéutico , Moléculas de Adhesión Celular/sangre , Quimiocinas/sangre , Dieta Alta en Grasa/efectos adversos , Hiperlipidemias/prevención & control , Animales , Antiinflamatorios no Esteroideos/administración & dosificación , Aorta Torácica/efectos de los fármacos , Aorta Torácica/metabolismo , Aspirina/administración & dosificación , Aterosclerosis/etiología , Aterosclerosis/inmunología , Aterosclerosis/metabolismo , Aterosclerosis/prevención & control , Plaquetas/inmunología , Plaquetas/metabolismo , Peso Corporal/efectos de los fármacos , Moléculas de Adhesión Celular/biosíntesis , Quimiocinas/inmunología , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Hiperlipidemias/sangre , Hiperlipidemias/complicaciones , Hiperlipidemias/inmunología , Lípidos/sangre , Linfocitos/inmunología , Linfocitos/metabolismo , Masculino , Ratas Sprague-Dawley , Aumento de Peso/efectos de los fármacosRESUMEN
This study investigated whether KMUP-1, a xanthine-based derivative, inhibits L-type Ca(2+) currents (I(Ca,L)) in rat basilar artery smooth muscle cells (RBASMCs). We used whole cell patch-clamp recording to monitor Ba(2+) currents (I(Ba)) through L-type Ca(2+) channels (LTCCs). Under voltage-clamp conditions, holding at -40 mV, KMUP-1 (1, 3, 10 µM) inhibited I(Ba) in a concentration-dependent manner and its IC(50) value was 2.27 ± 0.45 µM. A high concentration of KMUP-1 (10 µM) showed without modifying the I(Ba) current-voltage relationship. On the other hand, the protein kinase C (PKC) activator phorbol 12-myristate 13-acetate (PMA, 1 µM) increase I(Ba) was inhibited by KMUP-1. Pretreatment with the PKC inhibitor chelerythrine (5 µM) intensified KMUP-1-inhibited I(Ba). However, the Rho kinase inhibitor Y-27632 (30 µM) failed to affect the I(Ba) inhibition by KMUP-1. In light of these results, we suggest that KMUP-1 inhibition of LTCCs in concentration- and voltage-dependent manners in RBASMCs may be due, at least in part, to its modulation of the PKC pathway.
Asunto(s)
Arteria Basilar/citología , Bloqueadores de los Canales de Calcio/farmacología , Canales de Calcio Tipo L/metabolismo , Miocitos del Músculo Liso/efectos de los fármacos , Piperidinas/farmacología , Proteína Quinasa C/metabolismo , Xantinas/farmacología , Animales , Compuestos de Bario/farmacología , Benzofenantridinas/farmacología , Cloruros/farmacología , Femenino , Potenciales de la Membrana/efectos de los fármacos , Miocitos del Músculo Liso/enzimología , Miocitos del Músculo Liso/metabolismo , Técnicas de Placa-Clamp , Proteína Quinasa C/antagonistas & inhibidores , Ratas , Ratas Sprague-Dawley , Transducción de Señal , Quinasas Asociadas a rho/metabolismoRESUMEN
OBJECTIVES: Eugenosedin-A is a serotonin (5-hydroxytryptamine; 5-HT) 5-HT(1B/2A) and alpha(1)/alpha(2)/beta(1)-adrenoceptor blocker with anti-oxidative, anti-inflammatory and free-radical scavenging activities. Previous reports demonstrated that 5-HT(2A) blockers could diminish hyperlipidaemia. This study therefore aimed to investigate the possible uses and mechanisms of eugenosedin-A and other agents in treating hyperlipidaemia. METHODS: C57BL/6J mice were randomly divided into seven groups, fed a regular diet or a high-fat diet alone or supplemented with one of five agents: eugenosedin-A, ketanserin, prazosin, propranolol or atorvastatin (5 mg/kg p.o.) for 8 weeks. KEY FINDINGS: Compared with the regular diet, the mice fed the high-fat diet had significantly higher body weight and glucose, insulin and lipid levels. Brain malondialdehyde concentration was increased and liver glutathione peroxidase activity decreased. Addition of eugenosedin-A to the high-fat diet resulted in less weight gain and reduced hyperglycaemia, hyperinsulinaemia and hyperlipidaemia. Lipid and glucose homeostasis were related to decreased hepatic lipogenesis mRNAs and proteins (sterol regulatory element binding protein 1a, fatty acid synthase, sterol-CoA desaturase) and restored adipose peroxisome proliferator-activated receptor gamma expression. Eugenosedin-A also enhanced low-density lipoprotein receptor mRNA expression. CONCLUSIONS: Eugenosedin-A may improve plasma lipid metabolism by increasing low-density lipoprotein receptor and peroxisome proliferator-activated receptor gamma expression and diminishing sterol regulatory element binding protein 1a, fatty acid synthase and sterol-CoA desaturase. Reduction of plasma glucose and lipid levels may, in turn, reduce insulin concentration, which would explain the marked improvement in obesity-related hyperglycaemia and hyperlipidaemia. Furthermore, eugenosedin-A affected malondialdehyde concentration and glutathione peroxidase activity, suggesting it may have anti-peroxidation effects in mice fed a high-fat diet.
Asunto(s)
Grasas de la Dieta/farmacología , Hiperglucemia/prevención & control , Hiperlipidemias/prevención & control , Peroxidación de Lípido/efectos de los fármacos , Piperazinas/farmacología , Tejido Adiposo/metabolismo , Animales , Atorvastatina , Glucemia/efectos de los fármacos , Peso Corporal/efectos de los fármacos , Encéfalo/metabolismo , Femenino , Glutatión Peroxidasa/metabolismo , Ácidos Heptanoicos/farmacología , Insulina/sangre , Ketanserina/farmacología , Lipogénesis/genética , Hígado/metabolismo , Malondialdehído/metabolismo , Ratones , Ratones Endogámicos C57BL , PPAR gamma/genética , PPAR gamma/metabolismo , Prazosina/farmacología , Propranolol/farmacología , Pirroles/farmacología , Superóxido Dismutasa/metabolismo , Aumento de Peso/efectos de los fármacosRESUMEN
Eugenosedin-B is able to block serotonin (5-HT) and alpha/beta receptors and to inhibit platelet aggregation. In Wistar rats, intravenous injections of eugenosedin-B (2.4, 7.2, 12 micromoL/kg) caused a dose-dependent decrease in blood pressure and heart rate. In contrast, intracisternal injection of eugenosedin-B (0.3, 0.03 micromoL) and an alpha2-antagonist yohimbine (0.03 micromoL) increased blood pressure and heart rate. Eugenosedin-B and yohimbine prevented hypotension induced by intracisternal injection of an alpha2-agonist clonidine (38 pmol). In in vitro experiments, eugenosedin-B (10, 10, 10 M) competitively antagonized norepinephrine-, clonidine-, and 5-HT (10 to 10 M)-induced vasocontractions in isolated rat aorta. It also competitively antagonized the isoproterenol (10 to 10 M)-induced positive inotropic effects in isolated rat atrium. These findings clearly suggest that eugenosedin-B possesses alpha1, alpha2, beta1, and 5-HT2A receptor blocking activities. In isolated rabbit ear artery sensitized with 16 mM K, eugenosedin-B antagonized 5-nonyloxytryptamine- and 5-HT-induced vasocontractions, indicating it also blocked 5-HT1B and 5-HT2A receptors. In radioligand-binding experiments, eugenosedin-B had significant binding affinities on alpha1, alpha2, beta1, 5-HT1B, and 5-HT2A receptors. In human platelets, eugenosedin-B inhibited epinephrine and 5-HT-induced aggregations. It also had competitive binding effects in human platelet with [H]yohimbine (alpha2), [H]ketanserin (5-HT2A). We conclude that hypotensive and vasorelaxant effects of eugenosedin-B can be attributed to its multiple actions on the blockade of 5-HT1B, 5-HT2A, alpha1/2 and beta1 receptors, and its ability to reduce platelet aggregation attributed to its blockade of alpha2 and 5-HT2A receptors.
Asunto(s)
Antagonistas Adrenérgicos alfa/farmacología , Antagonistas Adrenérgicos beta/farmacología , Plaquetas/efectos de los fármacos , Piperazinas/farmacología , Inhibidores de Agregación Plaquetaria/farmacología , Antagonistas de la Serotonina/farmacología , Estirenos/farmacología , Vasodilatadores/farmacología , Antagonistas Adrenérgicos alfa/administración & dosificación , Antagonistas Adrenérgicos beta/administración & dosificación , Animales , Aorta Torácica/efectos de los fármacos , Aorta Torácica/fisiología , Unión Competitiva , Plaquetas/metabolismo , Presión Sanguínea/efectos de los fármacos , Oído/irrigación sanguínea , Humanos , Hipotensión/inducido químicamente , Hipotensión/prevención & control , Técnicas In Vitro , Inyecciones Intravenosas , Inyecciones Intraventriculares , Masculino , Contracción Miocárdica/efectos de los fármacos , Piperazinas/administración & dosificación , Inhibidores de Agregación Plaquetaria/administración & dosificación , Conejos , Ensayo de Unión Radioligante , Ratas , Ratas Wistar , Receptor de Serotonina 5-HT1B/metabolismo , Receptor de Serotonina 5-HT2A/metabolismo , Receptores Adrenérgicos alfa 1/fisiología , Receptores Adrenérgicos alfa 2/fisiología , Antagonistas del Receptor de Serotonina 5-HT1 , Antagonistas del Receptor de Serotonina 5-HT2 , Antagonistas de la Serotonina/administración & dosificación , Estirenos/administración & dosificación , Vasoconstricción/efectos de los fármacos , Vasodilatadores/administración & dosificaciónRESUMEN
In this study, we investigate the protective effects of eugenosedin-A on p38 mitogen-activated protein kinase (MAPK), inflammatory nitric oxide (NO) and cyclooxygenase-2 (COX-2) pathways in a rat model of endotoxin shock. Rats were pretreated with eugenosedin-A, trazodone, yohimbine (1 mg kg(-1), i.v.), aminoguanidine or ascorbic acid (15 mg kg(-1), i.v.) 30 min before endotoxin challenge. Endotoxaemia was induced by a single i.v. injection of lipopolysaccharide (LPS, 10 mg kg(-1)). In rats not treated with eugenosedin-A, LPS increased plasma concentrations of NO and prostaglandin E(2) (PGE(2)), and levels of p38 MAPK, inducible NO synthase (iNOS) and COX-2 proteins in the liver, lung, aorta and lymphocytes. In the pre-treated rats, eugenosedin-A not only inhibited the LPS-induced NO and PGE(2) levels but also attenuated the LPS-induced increase in p38 MAPK and iNOS levels in the liver, aorta and lymphocytes. Eugenosedin-A also reduced LPS-induced COX-2 proteins in the aorta and lymphocytes. Likewise, aminoguanidine, ascorbic acid, yohimbine and trazodone were also found to decrease NO and PGE(2) concentrations after endotoxin challenge. While aminoguanidine and ascorbic acid also attenuated the LPS-induced increase in p38 MAPK, iNOS and COX-2 proteins in the aorta and lymphocytes, trazodone and yohimbine inhibited only the increase in p38 MAPK, iNOS and COX-2 proteins in lymphocytes. Finally, eugenosedin-A (10(-10)-10(-8) M) significantly inhibited the biphasic response induced by hydrogen peroxide (10(-6)-3 x 10(-5) M) in rat denudated aorta. Taken together, the results of this study indicate that eugenosedin-A, as well as ascorbic acid, can attenuate free-radical-mediated aortic contraction and relaxation. It may therefore be able to reduce the damage caused by septic shock by inhibiting formation of p38 MAPK, iNOS, COX-2 and free radicals.