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1.
Med Sci Monit ; 25: 1769-1779, 2019 Mar 08.
Artigo em Inglês | MEDLINE | ID: mdl-30848248

RESUMO

BACKGROUND Cardiac remote ischemic conditioning (RIC) is a noninvasive cardioprotective method in ischemia-reperfusion injury and acute myocardial infarction (AMI). The aims of this study were to investigate the effects of RIC in a rat model of AMI. MATERIAL AND METHODS Adult male Sprague-Dawley rats included the AMI group that underwent ligation of the left anterior descending (LAD) coronary artery (n=24), the RIC group that consisted the AMI rat model treated with RIC once daily in the left hind limb until days 1, 7 and 14 (n=24), and the sham group (n=24). Myocardial infarct size was measured by routine histology with triphenyltetrazolium chloride (TTC) and Masson's trichrome histochemical staining for myocardial necrosis and fibrosis, respectively. Serum levels of Bcl-2, Bax, caspase-3, and inducible nitric oxide synthase (iNOS) were measured by enzyme-linked immunosorbent assay (ELISA). The apoptosis index was detected using the TUNEL assay. Spectrophotometry of the myocardium was used to identify mitochondrial complexes and myocardial ATP. RESULTS The RIC group showed improved cardiac hemodynamics, reduced the size of the myocardial infarction, upregulated expression of Bcl-2, and down-regulation of the levels of Bax, caspase-3, and iNOS, and reduced cardiac myocyte apoptosis and inhibited the opening of the mitochondrial permeability transition pore (MPTP). CONCLUSIONS In a rat model of AMI, RIC improved the hemodynamic index, reduce the levels of apoptosis and myocardial injury, and improved mitochondrial function.


Assuntos
Precondicionamento Isquêmico/métodos , Infarto do Miocárdio/metabolismo , Traumatismo por Reperfusão/prevenção & controle , Animais , Apoptose , Cardiotônicos , Caspase 3/análise , Caspase 3/sangue , Modelos Animais de Doenças , Traumatismos Cardíacos/prevenção & controle , Hemodinâmica , Masculino , Mitocôndrias/metabolismo , Infarto do Miocárdio/fisiopatologia , Traumatismo por Reperfusão Miocárdica/metabolismo , Miocárdio/metabolismo , Miocárdio/patologia , Proteínas Proto-Oncogênicas c-bcl-2/análise , Proteínas Proto-Oncogênicas c-bcl-2/sangue , Ratos , Ratos Sprague-Dawley , Traumatismo por Reperfusão/terapia , Proteína X Associada a bcl-2/análise , Proteína X Associada a bcl-2/sangue
2.
Zhongguo Ying Yong Sheng Li Xue Za Zhi ; 34(2): 164-168, 2018 Feb 08.
Artigo em Chinês | MEDLINE | ID: mdl-29926683

RESUMO

OBJECTIVES: To investigate the effects of Astragaloside IV (AST) on diastolic function of rat thoracic aorta rings which was injured by microvesicles derived from hypoxia/reoxygenation (H/R)-treated human umbilical vein endothelial cells (HUVECs), and the mechanism of AST. METHODS: H/R-induced endothelial microvesicles (H/R-EMVs) were generated from cultured HUVECs in vitro under the condition of hypoxia for 12 hour/Reoxygenation for 4 hour, H/R-EMVs were stored in D-Hank's solution. Male Wistar rats were underwent thoracotomy, the thoracic aorta with intact endothelium were carefully removed and cut into 3~4 mm rings. The experiment was divided into six groups. H/R-EMVs group:thoracic aortic rings of rats were incubated in culture medium and treated with H/R-EMVs in a final concentration of 10µg/ml; different doses of AST groups:thoracic aortic rings of rats were treated with 10, 20, 40, 60 mg/L AST co-incubated with 10µg/ml H/R-EMVs respectively; control group were treated with the same volume of D-Hank's solution. Duration of incubation was 4 h, each group was tested in five replicate aortic rings. Effects of AST on endothelium-dependent relaxation were detected. The production of nitric oxide (NO) and the level of endothelial NO synthase (eNOS), phosphorylated eNOS (p-eNOS, Ser-1177), serine/threonine kinase (Akt), phosphorylated Akt (p-Akt, Ser-473), extracellular regulated protein kinases (ERK1/2) and phosphorylated ERK1/2 (p-ERK1/2, Thr202/Tyr204) of rat thoracic aortic rings were detected. RESULTS: Tenµg/ml H/R-EMVs could impaire the relaxation of rat thoracic aortic rings significantly (P<0.01). Compared with H/R-EMVs group, relaxation of rat thoracic aortic rings was increased by 20, 40 and 60 mg/L AST in a concentration-dependent manner (P<0.01), the level of NO production was also enhanced (P<0.05, P<0.01). The level of t-eNOS, t-Akt and ERK1/2 was not changed, but the level of p-eNOS, p-Akt and p-ERK1/2 increased by the treatment with AST (P<0.01). CONCLUSIONS: AST could effectively ameliorate endotheliumdependent relaxation of rat thoracic aortic rings impaired by H/R-EMVs in a concentration-dependent manner, the mechanism might involve the increase in production of NO, and the protein level of p-eNOS, p-Akt and p-ERK1/2.


Assuntos
Aorta Torácica/efeitos dos fármacos , Micropartículas Derivadas de Células/patologia , Saponinas/farmacologia , Triterpenos/farmacologia , Vasodilatação , Animais , Células Endoteliais da Veia Umbilical Humana , Humanos , Técnicas In Vitro , Sistema de Sinalização das MAP Quinases , Masculino , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos , Ratos Wistar
3.
Eur J Neurosci ; 46(11): 2746-2753, 2017 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-29057540

RESUMO

ATP-sensitive K+ (K-ATP) channels play significant roles in regulating the excitability of dopamine neurons in the substantia nigra zona compacta (SNC). We showed previously that K-ATP channel function is up-regulated by AMP-activated protein kinase (AMPK). This study extended these studies to the neurons adjacent to the SNC in the ventral tegmental area (VTA). Using patch pipettes to record whole-cell currents in slices of rat midbrain, we found that the AMPK activator A769662 increased the amplitude of currents evoked by the K-ATP channel opener diazoxide in presumed dopamine-containing VTA neurons. However, current evoked by diazoxide with A769662 was significantly smaller in VTA neurons compared to SNC neurons. Moreover, a significantly lower proportion of VTA neurons responded to diazoxide with outward current. However, A769662 was able to increase the incidence of diazoxide-responsive neurons in the VTA. In contrast, A769662 did not potentiate diazoxide-evoked currents in presumed non-dopamine VTA neurons. These results show that AMPK activation augments K-ATP currents in presumed dopamine neurons in the VTA and SNC, although diazoxide-evoked currents remain less robust in the VTA. We conclude that K-ATP channels may play important physiological roles in VTA and SNC dopamine neurons.


Assuntos
Adenilato Quinase/metabolismo , Canais KATP/metabolismo , Parte Compacta da Substância Negra/citologia , Parte Compacta da Substância Negra/metabolismo , Área Tegmentar Ventral/citologia , Área Tegmentar Ventral/metabolismo , Animais , Compostos de Bifenilo , Diazóxido/farmacologia , Neurônios Dopaminérgicos/fisiologia , Sinergismo Farmacológico , Masculino , Potenciais da Membrana/efeitos dos fármacos , Potenciais da Membrana/fisiologia , Parte Compacta da Substância Negra/efeitos dos fármacos , Pironas/farmacologia , Ratos , Tiofenos/farmacologia , Área Tegmentar Ventral/efeitos dos fármacos
4.
Oncotarget ; 8(33): 54572-54582, 2017 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-28903365

RESUMO

OBJECTIVE: To investigate the effects of circulating microvesicles derived from myocardial ischemia (I-MVs) on apoptosis in myocardial ischemia/reperfusion (I/R) injury in rats. METHODS: I-MVs from rats undergoing myocardial left anterior descending (LAD) coronary artery ligation were isolated by ultracentrifugation from circulating blood and characterized by flow cytometry. I-MVs were administered intravenously (4.8 mg/kg) at 5 min before reperfusion procedure in I/R injury model which was induced by 30-min of ischemia and 120-min of reperfusion of LAD in rats. RESULTS: Treatment with I-MVssignificantly reduced the size of myocardial infarction, the activities of serum CK-MB and LDH, and the number of apoptotic cardiomyocytes. The activities of caspase 3, caspase 9 and caspase 12 in myocardium were also decreased significantly with I-MVs treatment. Moreover, the expression of Bax was decreased but Bcl-2 was increased. The expression of glucose regulated protein 78 (GRP78), sarco/endoplasmic reticulum Ca2+-ATPase 2 (SERCA2) and phosphorylated phospholamban (p-PLB) were increased after being treated with I-MVs. CONCLUSION: I-MVs could protect hearts from I/R injury in rats through SERCA2 and p-PLB of calcium regulatory proteins to alleviate intrinsic myocardial apoptosis including mitochondrial and endoplasmic reticulum pathways.

5.
Biochem Biophys Res Commun ; 488(2): 278-284, 2017 06 24.
Artigo em Inglês | MEDLINE | ID: mdl-28479248

RESUMO

Recent studies have demonstrated that remote ischemic conditioning (RIC) creates cardioprotection against ischemia/reperfusion injury and myocardial infarction (MI); however, the effects of non-invasive remote ischemic conditioning (nRIC) on prognosis and rehabilitation after MI (post-MI) remain unknown. We successfully established MI models involving healthy adult male Sprague-Dawley rats. The nRIC group repeatedly underwent 5 min of ischemia and 5 min of reperfusion in the left hind limb for three cycles every other day until weeks 4, 6, and 8 after MI. nRIC improved cardiac hemodynamic function and mitochondrial respiratory function through increasing myocardial levels of mitochondrial respiratory chain complexes I, II, III, IV, and adenosine triphosphate (ATP) and decreasing the activity of nitric oxide synthase (NOS). nRIC could inhibit cardiomyocytes apoptosis and reduce myocardium injury through raising the expression of Bcl-2 and reduced the content of creatine kinase-MB, cardiac troponin I and Bax. The results indicated that long-term nRIC could accelerate recovery and improve prognosis and rehabilitation in post-MI rats.


Assuntos
Precondicionamento Isquêmico Miocárdico , Infarto do Miocárdio/reabilitação , Infarto do Miocárdio/terapia , Animais , Masculino , Infarto do Miocárdio/metabolismo , Ratos , Ratos Sprague-Dawley
6.
Neuroscience ; 330: 219-28, 2016 08 25.
Artigo em Inglês | MEDLINE | ID: mdl-27267246

RESUMO

AMP-activated protein kinase (AMPK) is a master enzyme that regulates ATP-sensitive K(+) (K-ATP) channels in pancreatic beta-cells and cardiac myocytes. We used patch pipettes to record currents and potentials to investigate effects of AMPK on K-ATP currents in substantia nigra compacta (SNC) dopamine neurons in slices of rat midbrain. When slices were superfused repeatedly with the K-ATP channel opener diazoxide, we were surprised to find that diazoxide currents gradually increased in magnitude, reaching 300% of the control value 60min after starting whole-cell recording. However, diazoxide current increased significantly more, to 472% of control, when recorded in the presence of the AMPK activator A769662. Moreover, superfusing the slice with the AMPK blocking agent dorsomorphin significantly reduced diazoxide current to 38% of control. Control experiments showed that outward currents evoked by the K-ATP channel opener NN-414 also increased over time, but not currents evoked by the GABAB agonist baclofen. Delaying the application of diazoxide after starting whole-cell recording correlated with augmentation of current. Loose-patch recording showed that diazoxide produced a 34% slowing of spontaneous firing rate that did not intensify with repeated applications of diazoxide. However, superfusion with A769662 significantly augmented the inhibitory effect of diazoxide on firing rate. We conclude that K-ATP channel function is augmented by AMPK, which is activated during the process of making whole-cell recordings. Our results suggest that AMPK and K-ATP interactions may play an important role in regulating dopamine neuronal excitability.


Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Neurônios Dopaminérgicos/metabolismo , Canais KATP/metabolismo , Parte Compacta da Substância Negra/metabolismo , Proteínas Quinases Ativadas por AMP/antagonistas & inibidores , Potenciais de Ação/efeitos dos fármacos , Potenciais de Ação/fisiologia , Animais , Compostos de Bifenilo , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Fármacos do Sistema Nervoso Central/farmacologia , Óxidos S-Cíclicos/farmacologia , Diazóxido/farmacologia , Neurônios Dopaminérgicos/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Masculino , Parte Compacta da Substância Negra/efeitos dos fármacos , Técnicas de Patch-Clamp , Pirazóis/farmacologia , Pirimidinas/farmacologia , Pironas/farmacologia , Ratos Sprague-Dawley , Tiofenos/farmacologia , Técnicas de Cultura de Tecidos
7.
Zhongguo Ying Yong Sheng Li Xue Za Zhi ; 32(2): 97-101, 2016 Feb 08.
Artigo em Chinês | MEDLINE | ID: mdl-29931856

RESUMO

OBJECTIVE: To investigate the effects of circulating microvesicles (MVs) derived from ischemic preconditioning (IPC) on myocardial ischemia/reperfusion (I/R) injury in rats and explore the underlying mechanism. METHODS: To establish the IPC model, the rats were subjected to brief cycles of left anterior descending (LAD) coronary occlusion and reperfusion. The blood was drawn from abdominal aorta once the operation was finished. IPC-MVs were isolated by ultracentrifugation from the peripheral blood and characterized by flow cytometry. The myocardial I/R model of rats was established in vivo. Rats were injected via the femoral vein with IPC-MVs at 7 mg/kg. Morphological changes of myocardium were observed microscopically after HE staining. Apoptosis of myocardial cells was detected with TUNEL assay. Myocardial infarct size was detected by TTC staining. Moreover, activity of plasma lactate dehydrogenase (LDH) was tested by colorimetry. The activity of caspase 3 in myocardium was assayed with spectrophotometry. Expression levels of Bcl-2 and Bax protein were examined with Western blot. RESULTS: The concentration of IPC-MVs, which was detected by flow cytometry, was 4380±745 cells/µl. Compared with I/R group, IPC-MVs alleviated the damage of tissues in I/R injured rats significantly. The myocardial infarct size and the cardiomyocyte apoptotic index were obviously decreased after IPC-MVs treatment (P<0.01, respectively). The activity of plasma LDH was significantly decreased in IPC-MVs treated rats (P<0.01). Moreover, the activity of caspase 3 was markedly decreased after IPC-MVs treatment (P<0.01). In addition, the expression of Bcl-2 was increased (P<0.01), the expression of Bax was decreased (P<0.01), the ratio of Bcl-2/Bax was significantly increased after IPC-MVs treatment (P<0.01). CONCLUSIONS: IPC-MVs protected myocardial against I/R injury by up-regulating the expression of Bcl-2 protein, down-regulating the expression of Bax protein, increasing the ratio of Bcl-2/Bax and decreasing cleavage of caspase 3.


Assuntos
Micropartículas Derivadas de Células , Precondicionamento Isquêmico Miocárdico , Traumatismo por Reperfusão Miocárdica/terapia , Animais , Apoptose , Caspase 3/metabolismo , Infarto do Miocárdio/patologia , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Ratos , Proteína X Associada a bcl-2/metabolismo
8.
Brain Res ; 1603: 1-7, 2015 Apr 07.
Artigo em Inglês | MEDLINE | ID: mdl-25656790

RESUMO

Recent studies suggest that selective block of extrasynaptic N-methyl-d-aspartate (NMDA) receptors might protect against neurodegeneration. We recorded whole-cell currents with patch pipettes to characterize the ability of memantine, a low-affinity NMDA channel blocker, to block synaptic and extrasynaptic NMDA receptors in substantia nigra zona compacta (SNC) dopamine neurons in slices of rat brain. Pharmacologically isolated NMDA receptor-mediated EPSCs were evoked by electrical stimulation, whereas synaptic and extrasynaptic receptors were activated by superfusing the slice with NMDA (10 µM). Memantine was 15-fold more potent for blocking currents evoked by bath-applied NMDA compared to synaptic NMDA receptors. Increased potency for blocking bath-applied NMDA currents was shared by the GluN2C/GluN2D noncompetitive antagonist DQP-1105 but not by the high-affinity channel blocker MK-801. Our data suggest that memantine causes a selective block of extrasynaptic NMDA receptors that are likely to contain GluN2C/2D subunits. Our results justify further investigations on the use of memantine as a neuroprotective agent in Parkinson's disease.


Assuntos
Neurônios Dopaminérgicos/efeitos dos fármacos , Antagonistas de Aminoácidos Excitatórios/farmacologia , Memantina/farmacologia , Receptores de N-Metil-D-Aspartato/antagonistas & inibidores , Substância Negra/efeitos dos fármacos , Animais , Maleato de Dizocilpina/farmacologia , Neurônios Dopaminérgicos/fisiologia , Relação Dose-Resposta a Droga , Agonistas de Aminoácidos Excitatórios/farmacologia , Masculino , Potenciais da Membrana/efeitos dos fármacos , N-Metilaspartato/farmacologia , Técnicas de Patch-Clamp , Pirazóis/farmacologia , Quinolonas/farmacologia , Ratos Sprague-Dawley , Receptores de N-Metil-D-Aspartato/agonistas , Receptores de N-Metil-D-Aspartato/metabolismo , Substância Negra/fisiologia , Técnicas de Cultura de Tecidos
9.
Zhongguo Ying Yong Sheng Li Xue Za Zhi ; 31(6): 524-31, 2015 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-27215020

RESUMO

OBJECTIVE: To establish a flow cytometric method to detect the alteration of phenotypes and concentration of circulating microvesicles (MVs) from myocardial ischemic preconditioning (IPC) treated rats (IPC-MVs), and to investigate the effects of IPC-MVs on ischemia/reperfusion (I/R) injury in rats. METHODS: Myocardial IPC was elicited by three.cycles of 5-min ischemia and 5-min reperfusion of the left anterior descending (LAD) coronary artery. Platelet-free plasma (PFP) was isolated through two steps of centrifugation at room temperature from the peripheral blood, and IPC-MVs were isolated by ultracentrifugation from PFR PFP was incubated with anti-CD61, anti-CD144, anti-CD45 and anti-Erythroid Cells, and added 1, 2 µm latex beads to calibrate and absolutely count by flow cytometry. For functional research, I/R injury was induced by 30-min ischemia and 120-min reperfusion of LAD. IPC-MVs 7 mg/kg were infused via the femoral vein in myocardial I/R injured rats. Mean arterial blood pressure (MAP), heart rate (HR) and ST-segment of electro-cardiogram (ECG) were monitored throughout the experiment. Changes of myocardial morphology were observed after hematoxylin-eosin (HE) staining. The activity of plasma lactate dehydrogenase (LDH) was tested by Microplate Reader. Myocardial infarct size was measured by TTC staining. RESULTS: Total IPC-MVs and different phenotypes, including platelet-derived MVs (PMVs), endothelial cell-derived MVs (EMVs), leucocyte-derived MVs (LMVs) and erythrocyte-derived MVs (RMVs) were all isolated which were identified membrane vesicles (<1 Vm) with corresponding antibody positive. The numbers of PMVs, EMVs and RMVs were significantly increased in circulation of IPC treated rats (P<0.05, respectively). In addition, at the end of 120-min reperfusion in I/R injured rats, IPC-MVs markedly increased HR (P<0.01), decreased ST-segment and LDH activity (P < 0.05, P < 0.01). The damage of myocardium was obviously alleviated and myocardial infarct size was significantly lowered after IPC-MVs treatment (P < 0.01). CONCLUSION: The method of flow cytometry was successfully established to detect the phenotypes and concentration alteration of IPC-MVs, including PMVs, EMVs, LMVs and RMVs. Furthermore, circulating IPC-MVs protected myocardium against I/R injury in rats.


Assuntos
Micropartículas Derivadas de Células/metabolismo , Precondicionamento Isquêmico Miocárdico , Infarto do Miocárdio/fisiopatologia , Traumatismo por Reperfusão Miocárdica/fisiopatologia , Animais , Vasos Coronários/patologia , Citometria de Fluxo , Frequência Cardíaca , Miocárdio/patologia , Fenótipo , Ratos
10.
Artigo em Inglês | MEDLINE | ID: mdl-26016367

RESUMO

OBJECTIVE: To investigate the effects of microvesicles (MVs) derived from hypoxia/reoxygenation (H/R)-treated human umbilical vein endothelial cells (HUVECs) on endothelium-dependent relaxation of rat thoracic aortic rings. METHODS: H/R injury model was established to induce HUVECs to release H/R-EMVs. H/R-EMVs from HUVECs were isolated by ultracentrifugation from the conditioned culture medium. H/R-EMVs were characterized using 1 µm latex beads and anti-PE-CD144 by flow cytometry. Thoracic aortic rings of rats were incubated with 2.5, 5, 10, 20 µg/ml H/R-EMVs derived from H/R-treated HUVECs for 4 hours, and their endothelium-dependent relaxation in response to acetylcholine (ACh) or endothelium-independent relaxation in response to sodium nitroprusside (SNP) was recorded in vitro. The nitric oxide (NO) production of ACh-treated thoracic aortic rings of rats was measured using Griess reagent. The expression of endothelial NO synthase (eNOS) and phosphorylated eNOS (p-eNOS, Ser-1177) in the thoracic aortic rings of rats was detected by Western blotting. Furthermore, the levels of SOD and MDA in H/R-EMVs-treated thoracic aortic rings of rats were measured using SOD and MDA kit. RESULTS: H/R-EMVs were induced by H/R-treated HUVECs and isolated by ultracentrifugation. The membrane vesicles (< 1 µm) induced by H/R were CD144 positive. ACh-induced relaxation and NO production of rat thoracic aortic rings were impaired by H/R-EMVs treatment in a concentration-dependent manner (P < 0.05, P < 0.01). The expression of total eNOS (t-eNOS) was not affected by H/R-EMVs. However, the expression of p-eNOS decreased after treated with H/R-EMVs. The activity of SOD decreased and the level of MDA increased in H/R-EMVs treated rat thoracic aortic rings (P < 0.01). CONCLUSION: ACh induced endothelium-dependent relaxation of thoracic aortic rings of rats was impaired by H/R-EMVs in a concentration-dependent manner. The mechanisms included a decrease in NO production, p-eNOS expression and an increase in oxidative stress.


Assuntos
Aorta Torácica/fisiologia , Endotélio Vascular/fisiologia , Células Endoteliais da Veia Umbilical Humana/citologia , Acetilcolina/farmacologia , Animais , Hipóxia Celular , Humanos , Técnicas In Vitro , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo , Nitroprussiato/farmacologia , Estresse Oxidativo , Ratos
11.
Zhongguo Ying Yong Sheng Li Xue Za Zhi ; 29(6): 559-64, 2013 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-24654540

RESUMO

OBJECTIVE: To investigate the effects of endothelial microvesicles (EMVs) induced by calcium ionophore A23187 on H9c2 cardiomyocytes. METHODS: Human umbilical vein endothelial cells (HUVECs) were treated with 10 micromol/L A23187 for 30 min. EMVs from HUVECs were isolated by ultracentrifugation from the conditioned culture medium. EMVs were characterized using 1 and 2 microm latex beads and anti-PE-CD144 antibody by flow cytometry. For functional research, EMVs at different concentrations were cocultured with H9c2 cardiomyocytes for 6 h. Cell viability of H9c2 cells and the activity of LDH leaked from H9c2 cells were tested by colorimetry. Moreover, apoptosis of H9c2 cells was observed through Hoechst 33258 staining and tested by FITC-Annexin V/PI double staining. RESULTS: EMVs were induced by A23187 on HUVECs, and isolated by ultracentrifugation. We identified the membrane vesicles (< 1 microm) induced by A23187 were CD144 positive. In addition, the EMVs could significantly reduce the viability of H9c2 cells, and increase LDH leakage from H9c2 cells in a dose dependent manner (P < 0.05). Condensed nuclei could be observed with the increasing concentrations of EMVs through Hoechst 33258 staining. Furthermore, increased apoptosis rates of H9c2 cells could be assessed through FITC-Annexin V/PI double staining by flow cytometry. CONCLUSION: Microvesicles could be released from HUVECs after induced by A23187 through calcium influx, and these EMVs exerted a pro-apoptotic effect on H9c2 cells by induction of apoptosis.


Assuntos
Apoptose , Calcimicina/farmacologia , Membrana Celular/efeitos dos fármacos , Miócitos Cardíacos/efeitos dos fármacos , Anexina A5 , Cálcio/metabolismo , Linhagem Celular , Técnicas de Cocultura , Citometria de Fluxo , Fluoresceína-5-Isotiocianato/análogos & derivados , Células Endoteliais da Veia Umbilical Humana , Humanos , Coloração e Rotulagem
12.
Neurotoxicology ; 33(3): 429-35, 2012 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-22521663

RESUMO

Previous patch-clamp studies by our laboratory showed that acute exposure to the pesticide rotenone augments inward currents evoked by N-methyl-d-aspartate (NMDA) in substantia nigra zona compacta (SNC) dopamine neurons in slices of rat brain. The present experiments were done to search for histological evidence of increased neurotoxicity produced by combined rotenone and NMDA treatments. In horizontal slices of rat midbrain, we found that a 30 min superfusion with 100 nM rotenone caused significant injury to tyrosine hydroxylase (TH)-positive proximal dendrites in dorsal and ventral regions of the SNC and ventral tegmental area (VTA). Moreover, treatment with 100 µM NMDA potentiated rotenone toxicity. In contrast, treatment with 30 µM NMDA protected against rotenone-induced injury to dendrites in the ventral SNC and ventral VTA. Interestingly, treatment with 30 µM NMDA-alone produced an apparent increase in proximal dendrite scores in ventral SNC and dorsal VTA. We conclude that NMDA has concentration-dependent actions on rotenone toxicity that differ according to regional subtype of dopamine neuron.


Assuntos
Dopamina/metabolismo , Neurônios Dopaminérgicos/efeitos dos fármacos , N-Metilaspartato/farmacologia , Rotenona/toxicidade , Substância Negra/efeitos dos fármacos , Área Tegmentar Ventral/efeitos dos fármacos , Animais , Citoproteção , Dendritos/efeitos dos fármacos , Dendritos/metabolismo , Dendritos/patologia , Neurônios Dopaminérgicos/metabolismo , Neurônios Dopaminérgicos/patologia , Relação Dose-Resposta a Droga , Imuno-Histoquímica , Técnicas In Vitro , Masculino , N-Metilaspartato/toxicidade , Ratos , Ratos Sprague-Dawley , Substância Negra/metabolismo , Substância Negra/patologia , Tirosina 3-Mono-Oxigenase/metabolismo , Área Tegmentar Ventral/metabolismo , Área Tegmentar Ventral/patologia
13.
J Surg Res ; 174(1): 176-83, 2012 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-21195427

RESUMO

BACKGROUND: To study the protection offered by noninvasive delayed limb ischemic preconditioning (NDLIP) against cerebral ischemia reperfusion (I/R) injury in rats. MATERIALS AND METHODS: Healthy male Wistar rats were randomly divided into four groups. The delayed protection offered by NDLIP was estimated in light of changes in the neural behavior marker and cerebral tissue antioxidative ability. Neurological functions were studied by observing neural behavior. Total superoxide dismutase (T-SOD), manganese-superoxide dismutase (Mn-SOD), glutathione peroxidase (GSH-PX), and xanthine oxidase (XOD) activity in cerebral tissue and malonaldehyde (MDA) content were detected using a spectrophotometer. Mn-SOD mRNA was measured by the reverse transcription polymerase chain reaction method. RESULTS: Cerebral infarct size was diminished in the early cerebral ischemia preconditioning (ECIP)+I/R and NDLIP+I/R groups compared with the I/R group (P < 0.05). The cortical and hippocampal antioxidant enzyme activity and Mn-SOD expression were increased in the ECIP+I/R and NDLIP+I/R groups. In contrast, the cortical and hippocampal XOD activity and MDA content decreased in the ECIP+I/R and NDLIP+I/R groups. CONCLUSIONS: NDLIP decreased cerebral infarct size, increased cerebral antioxidative ability after I/R injury, and decreased peroxidative damage. The antioxidative protection offered by NDLIP was as effective as that offered by ECIP.


Assuntos
Antioxidantes/metabolismo , Isquemia Encefálica/metabolismo , Extremidades/irrigação sanguínea , Precondicionamento Isquêmico , Traumatismo por Reperfusão/prevenção & controle , Animais , Glutationa Peroxidase/metabolismo , Masculino , Malondialdeído/análise , RNA Mensageiro/análise , Ratos , Ratos Wistar , Traumatismo por Reperfusão/metabolismo , Superóxido Dismutase/genética , Superóxido Dismutase/metabolismo
14.
Artigo em Chinês | MEDLINE | ID: mdl-22097737

RESUMO

OBJECTIVE: To investigate whether Astragaloside IV(AST) protects H9c2 cells against H2O2-induced oxidative injury partly through ERK1/2 signaling pathway. METHODS: H9c2 cells oxidative injury was induced by 200 tmol/L H2O2 for 6 hours to establish the H2O2-induced injury model of H9c2 cells. The viability of H9c2 cells was detected using MTf method. Activity of lactate dehydrogenase(LDH), total-superoxide dismutase (T-SOD), manganese-superoxide dismutase (Mn-SOD) and content of MDA (malondialdehyde) in the culture medium were detected using colorimetric method. Western blot was performed to exam expression of p-ERK1/2 and ERK1/2 in H9c2 cells respectively. RESULTS: Under 200 micromol/L H2O2 treatment for 6 hours, the vaibility of H9c2 cells was suitable for the following study. Compared with H2O2 group, the cell viability was increased significantly in AST10 + H2O2 and AST2O + H2O2 groups (P < 0.01). The activity of LDH in the culture medium was decreased significantly (P < 0.01). The activity of T-SOD and Mn-SOD was increased significantly (P < 0.01), the content of MDA was decreased significantly (P < 0.01). Treated with 10 mg/L or 20 mg/L of AST, expression of p-ERK1/2 in H9c2 cells injured from H2O2 was increased significantly (P < 0.01), when PD98059 (inhibitor of ERK1/2) was added, the effects of AST were cancelled. CONCLUSION: AST protects H9c2 cells against H2O2-induced oxidative injury partly through ERK1/2 signaling pathway.


Assuntos
Peróxido de Hidrogênio/toxicidade , Sistema de Sinalização das MAP Quinases/fisiologia , Mioblastos Cardíacos/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Saponinas/farmacologia , Triterpenos/farmacologia , Animais , Antioxidantes/farmacologia , Linhagem Celular , Mioblastos Cardíacos/metabolismo , Mioblastos Cardíacos/patologia , Substâncias Protetoras/farmacologia , Ratos
15.
Brain Res ; 1395: 86-93, 2011 Jun 13.
Artigo em Inglês | MEDLINE | ID: mdl-21550595

RESUMO

Mild uncoupling of oxidative phosphorylation has been reported to reduce generation of reactive oxygen species (ROS) and therefore may be neuroprotective. We reported previously that the mitochondrial poison rotenone enhanced currents evoked by N-methyl-D-aspartate (NMDA) by a ROS-dependent mechanism in rat midbrain dopamine neurons. Thus, rotenone, which produces a model of Parkinson's disease in rodents, may increase the risk of dopamine neuron excitotoxicity. The purpose of this study was to test the hypothesis that oxidative phosphorylation uncoupling agents would antagonize the effect of rotenone on NMDA current. We used patch pipettes to record whole-cell currents under voltage-clamp (-60 mV) in substantia nigra dopamine neurons in slices of rat brain. Rotenone, NMDA and uncoupling agents were added to the brain slice superfusate. Inward currents evoked by NMDA (30 µM) more than doubled in amplitude after slices were superfused for 30 min with 100 nM rotenone. Continuous superfusion with the uncoupling agent carbonyl cyanide-p-trifluoromethoxy-phenylhydrazone (1-3 nM) or 2,4-dinitrophenol (100 nM) significantly antagonized and delayed the ability of rotenone to potentiate NMDA currents. Coenzyme Q10 (1-10 nM), which has been reported to facilitate uncoupling protein activity, also antagonized this action of rotenone. These results suggest that mild uncoupling of oxidative phosphorylation may protect dopamine neurons against injury from mitochondrial poisons such as rotenone.


Assuntos
Dopamina/fisiologia , Neurônios/efeitos dos fármacos , Rotenona/antagonistas & inibidores , Substância Negra/efeitos dos fármacos , Desacopladores/farmacologia , Animais , Interações Medicamentosas , Masculino , Neurônios/fisiologia , Técnicas de Cultura de Órgãos , Ratos , Ratos Sprague-Dawley , Rotenona/toxicidade , Substância Negra/metabolismo , Substância Negra/fisiopatologia , Desacopladores/uso terapêutico
16.
Free Radic Res ; 45(2): 201-10, 2011 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-20942563

RESUMO

This study was to explore whether repeated non-invasive limb ischemic pre-conditioning (NLIP) can confer an equivalent cardioprotection against myocardial ischemia-reperfusion (I/R) injury in acute diabetic rats to the extent of conventional myocardial ischemic pre-conditioning (MIP) and whether or not the delayed protection of NLIP is mediated by reducing myocardial oxidative stress after ischemia-reperfusion. Streptozotocin-induced diabetic rats were randomized to four groups: Sham group, the I/R group, the MIP group and the NLIP group. Compared with the I/R group, both the NLIP and MIP groups showed an amelioration of ventricular arrhythmia, reduced myocardial infarct size, increased activities of total superoxide dismutase (SOD), manganese-SOD and glutathione peroxidase, increased expression of manganese-SOD mRNA and decreased xanthine oxidase activity and malondialdehyde concentration (All p < 0.05 vs I/R group). It is concluded that non-invasive limb ischemic pre-conditioning reduces oxidative stress and attenuates myocardium ischemia-reperfusion injury in diabetic rats.


Assuntos
Diabetes Mellitus Experimental/metabolismo , Precondicionamento Isquêmico/métodos , Infarto do Miocárdio/metabolismo , Traumatismo por Reperfusão Miocárdica/metabolismo , Animais , Arritmias Cardíacas/metabolismo , Arritmias Cardíacas/fisiopatologia , Pressão Sanguínea , Diabetes Mellitus Experimental/fisiopatologia , Extremidades/irrigação sanguínea , Extremidades/fisiopatologia , Expressão Gênica , Glutationa Peroxidase/metabolismo , Hemodinâmica , Masculino , Malondialdeído/metabolismo , Modelos Animais , Infarto do Miocárdio/fisiopatologia , Traumatismo por Reperfusão Miocárdica/fisiopatologia , Traumatismo por Reperfusão Miocárdica/prevenção & controle , Miocárdio/metabolismo , Estresse Oxidativo , Ratos , Ratos Wistar , Estreptozocina/administração & dosagem , Superóxido Dismutase/genética , Superóxido Dismutase/metabolismo , Xantina Oxidase/metabolismo
17.
J Surg Res ; 164(1): 162-8, 2010 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-19726056

RESUMO

BACKGROUND: Transient limb ischemia induces remote early preconditioning that protects the myocardium from ischemia/reperfusion (I/R). However, it is unknown whether limb ischemia induces remote late preconditioning and whether it induces the same magnitude of cardioprotection compared with cardial ischemic preconditioning (CIP). We tested the hypothesis that late remote preconditioning of noninvasive limb ischemia (NLIP) offers the same magnitude of cardioprotection against myocardium I/R injury. METHODS: Thirty Wistar rats weighing 240-260 g each were randomly divided into three groups: I/R, CIP, and NLIP. The mean arterial pressure (MAP), heart rate (HR), ST-segment, ventricular arrhythmia, and CK-MB, cTnI, and superoxide dismutase (SOD) activity were measured after 0 and 30 min of ischemia and after 120 min of reperfusion. Myocardial infarct size, histologic examination, MMP-2, MMP-9, and TIMP-1 protein expression were determined at the end of the experiment. RESULTS: Compared with I/R groups, CIP and NLIP reduced ST-segment elevation (P<0.01), decreased incidence and duration of ventricular arrhythmia (P<0.01) during ischemia, decreased CK-MB (P<0.05), and cTnI (P<0.01) activity, and increased SOD (P<0.05) activity after reperfusion. The myocardial infarct size (P<0.01) was significantly reduced, and cell injury was attenuated in the CIP and NLIP groups compared with the I/R group. MMP-2 and MMP-9 protein expression was significantly decreased in the CIP and NLIP groups (P<0.01), while TIMP-1 expression was significantly increased in the CIP and NLIP groups compared with the I/R group (P<0.01). CONCLUSION: Remote preconditioning via NLIP has late cardioprotection against myocardium I/R injury and has a similar magnitude of cardioprotection compared with CIP in rats.


Assuntos
Membro Posterior/irrigação sanguínea , Precondicionamento Isquêmico Miocárdico/métodos , Infarto do Miocárdio/prevenção & controle , Traumatismo por Reperfusão Miocárdica/prevenção & controle , Animais , Pressão Sanguínea , Creatina Quinase Forma MB/sangue , Eletrocardiografia , Frequência Cardíaca , Masculino , Metaloproteinase 2 da Matriz/sangue , Metaloproteinase 9 da Matriz/sangue , Infarto do Miocárdio/metabolismo , Infarto do Miocárdio/patologia , Traumatismo por Reperfusão Miocárdica/metabolismo , Traumatismo por Reperfusão Miocárdica/patologia , Ratos , Ratos Wistar , Superóxido Dismutase/sangue , Inibidor Tecidual de Metaloproteinase-1/sangue , Troponina I/sangue
18.
Neurotoxicology ; 30(2): 320-5, 2009 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-19428506

RESUMO

Rotenone is a pesticide that has been successfully used to produce a rodent model of Parkinson's disease. We reported previously that rotenone potently augmented N-methyl-D-aspartate (NMDA)-evoked currents in rat dopamine neurons via a tyrosine kinase-dependent mechanism. In this study, we investigated the effect of rotenone on the current-voltage relationship of NMDA-induced currents in substantia nigra zona compacta neurons recorded with whole-cell patch pipettes in slices of rat brain. In a physiologic concentration of extracellular Mg(2+) (1.2mM), a 30min perfusion with rotenone (100nM) produced a marked increase in current evoked by bath application of NMDA (30microM), especially when measured at relatively hyperpolarized currents. In the presence of rotenone, NMDA currents lost the characteristic region of negative-slope conductance that is normally produced by voltage-dependent block by Mg(2+). The voltage-dependent effect of rotenone on NMDA currents was mimicked by a low extracellular concentration of Mg(2+) (0.2mM) and was antagonized by a high level of Mg(2+) (6.0mM). Moreover, we report that the tyrosine kinase inhibitor genistein blocked the ability of rotenone to augment NMDA receptor currents. These results suggest that rotenone potentiates NMDA currents by a tyrosine kinase-dependent process that attenuates voltage-dependent Mg(2+) block of NMDA-gated channels. These results support the hypothesis that an excitotoxic mechanism might participate in rotenone-induced toxicity of midbrain dopamine neurons.


Assuntos
Dopamina , Magnésio/farmacologia , Neurônios/efeitos dos fármacos , Receptores de N-Metil-D-Aspartato/antagonistas & inibidores , Rotenona/farmacologia , Substância Negra/efeitos dos fármacos , Animais , Dopamina/fisiologia , Agonistas de Dopamina/farmacologia , Antagonistas de Aminoácidos Excitatórios/farmacologia , Magnésio/antagonistas & inibidores , Masculino , Neurônios/fisiologia , Ratos , Ratos Sprague-Dawley , Receptores de N-Metil-D-Aspartato/fisiologia , Substância Negra/fisiologia
19.
Zhongguo Ying Yong Sheng Li Xue Za Zhi ; 25(4): 478-82, 2009 Nov.
Artigo em Chinês | MEDLINE | ID: mdl-21158037

RESUMO

AIM: To study the protective effects of ramipril in combination with BQ-123 on myocardial ischemia/reperfusion (I/R) injury in vivo in anesthetized rats. METHODS: Healthy male Wistar rats were divided into 5 groups randomly and subjected to 30 min of myocardial ischemia followed by 120 min reperfusion. Ramipril, BQ-123 and their combination were given to rats respectively. To observe the protection of their combination against myocardial I/R injury. HR, MAP and the change of ST-segment were observed. Ventricular arrthymias were monitored. The activity of creatine kinase (CK) and lactate dehydrogenase (LDH) in plasma, the infarct size and morphologic change were examined. RESULTS: Compared with I/R group, the elevation of ST-segment was decreased. Onsets of VPC and VT were delayed, durations of VPC and VT were shortened, especially their combination. Incidences of VPC, VT and VF were decreased. Activity of plasma CK and LDH was decreased, especially their combination. IS, IS/AAR and the morphology of myocardium were improved, especially their combination. CONCLUSION: Ramipril, BQ-123 and combined using these two agents protected myocardium from I/R injury in vivo. The protective effects on delaying onset of VA, shortening duration of VA, decreasing the activities of CK and LDH, decreasing infrarct size and improving morphology of myocardium were better than using ramipril and BQ-123 alone.


Assuntos
Cardiotônicos/farmacologia , Traumatismo por Reperfusão Miocárdica/prevenção & controle , Peptídeos Cíclicos/farmacologia , Ramipril/farmacologia , Animais , Arritmias Cardíacas/prevenção & controle , Creatina Quinase/sangue , Sinergismo Farmacológico , L-Lactato Desidrogenase/sangue , Masculino , Ratos , Ratos Wistar
20.
Artigo em Chinês | MEDLINE | ID: mdl-21158039

RESUMO

AIM: To investigate the effects of ramipril on myocardial ischemia/reperfusion injury in diabetic rats, and to explore its mechanism according to the observation on myocardial ultrastructure. METHODS: Streptozotocin induced diabetic rats were divided randomly into three groups (n = 16): ischemia/reperfusion (I/R), ischemic preconditioning (IPC) and ramipril (RAM) group. Rats in RAM group were administered by RAM(1 mg x kg(-1) x d(-1)) orally for 4 weeks, the others were administered by normal saline. Then all rats were subjected to myocardial ischemia/ reperfusion injury. Rats in IPC group were preconditioned before ischemia. The ECG and the infarct size were examined. The changes of myocardial morphology were examined by light and electron microscopes. RESULTS: Compared with I/R group, the elevation of ST segment and the incidence of ventricular tachycardia and ventricular fibrillation during ischemia were significantly decreased, the infarct size at the end of reperfusion was remarkably reduced, the myocardial morphology were significantly improved, special structure of myofilaments and mitochondria remained clearly, blood vessels were unobstructed, injury of endothelium were decreased in PC and RAM groups. CONCLUSION: Ramipril administered for 4 weeks induces myocardial protection in diabetic rats, which is similar to that of IPC. The mechanism may be involved in protection of cardiocytes and mitochondria, and improvement of endothelial function.


Assuntos
Diabetes Mellitus Experimental/complicações , Precondicionamento Isquêmico Miocárdico/métodos , Traumatismo por Reperfusão Miocárdica/prevenção & controle , Miocárdio/ultraestrutura , Ramipril/farmacologia , Animais , Cardiotônicos/farmacologia , Traumatismo por Reperfusão Miocárdica/patologia , Ratos
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