RESUMO
Increased matrix metalloproteinase (MMP) abundance occurs with adverse left ventricular (LV) remodeling in a number of cardiac disease states, including those induced by long-standing arrhythmias. However, whether regionally contained aberrant electrical activation of the LV, with consequent dyskinesia, alters interstitial MMP activation remained unknown. Electrical activation of the LV of pigs (n = 10, 30-35 kg) was achieved by pacing (150 beats/min) at left atrial and LV sites such that normal atrioventricular activation (60 min) was followed by regional early LV activation for 60 min within 1.5 cm of the paced site and restoration of normal atrioventricular pacing for 120 min. Regional shortening (piezoelectric crystals) and interstitial MMP activity (microdialysis with MMP fluorogenic substrate) at the LV pacing site and a remote LV site were monitored at 30-min intervals. During aberrant electrical stimulation, interstitial MMP activity at the paced site was increased (122 +/- 4%) compared with the remote region (100%, P < 0.05). Restoration of atrioventricular pacing after the 60-min period of aberrant electrical activation normalized segmental shortening (8.5 +/- 0.4%), but MMP activity remained elevated (121 +/- 6%, P < 0.05). This study demonstrates that despite the restoration of mechanical function, disturbances in electrical conduction, in and of itself, can cause acute increases in regional in vivo MMP activation and, therefore, contribute to myocardial remodeling.
Assuntos
Arritmias Cardíacas/enzimologia , Sistema de Condução Cardíaco/fisiopatologia , Metaloproteinases da Matriz/metabolismo , Função Ventricular Esquerda , Remodelação Ventricular , Animais , Arritmias Cardíacas/etiologia , Arritmias Cardíacas/fisiopatologia , Pressão Sanguínea , Estimulação Cardíaca Artificial , Modelos Animais de Doenças , Eletrocardiografia , Ativação Enzimática , Frequência Cardíaca , Ventrículos do Coração/enzimologia , Ventrículos do Coração/fisiopatologia , Recuperação de Função Fisiológica , Suínos , Fatores de Tempo , Regulação para Cima , Pressão VentricularRESUMO
BACKGROUND: Periods of ischemia-reperfusion (I/R) during cardiac surgery are associated with transient left ventricular (LV) dysfunction and an inflammatory response. In this study, we examined the potential dose-dependent effects of aprotinin (APRO) on LV contractility and cytokine release in the setting of I/R. METHODS: An index of LV contractility, LV maximal elastance (E(max)), was measured at baseline, 30 min of ischemia, and 60 min of reperfusion by microtransducer volumetry. Mice were randomized as follows: (a) APRO 20,000 kallikrein-inhibiting units (KIU)/kg (n = 11); (b) APRO 4 x 10(4) KIU/kg (n = 10); (c) APRO 8 x 10(4) KIU/kg (n = 10); and (d) vehicle (saline; n = 10). APRO doses were calculated to reflect half, full, and twice the clinical Hammersmith dosing schedule. After I/R, plasma was collected for cytokine measurements. RESULTS: After I/R, E(max) decreased from the baseline value by more than 40% in the vehicle group as well as in the APRO 4 x 10(4) KIU/kg and APRO 8 x 10(4) KIU/kg groups (P < 0.05). However, E(max) returned to near baseline values in the APRO 2 x 10(4) KIU/kg group. Tumor necrosis factor (TNF) increased 10-fold after I/R, but it was reduced with higher APRO doses. CONCLUSIONS: This study demonstrated that a low dose of APRO provided protective effects on LV contractility, whereas higher doses suppressed TNF release. These unique findings suggest that there are distinct and independent mechanisms of action of APRO in the context of I/R.
Assuntos
Aprotinina/farmacologia , Citocinas/metabolismo , Hemostáticos/farmacologia , Contração Miocárdica/efeitos dos fármacos , Traumatismo por Reperfusão Miocárdica/metabolismo , Traumatismo por Reperfusão Miocárdica/fisiopatologia , Função Ventricular Esquerda/efeitos dos fármacos , Animais , Aprotinina/sangue , Arritmias Cardíacas/etiologia , Arritmias Cardíacas/fisiopatologia , Relação Dose-Resposta a Droga , Elasticidade , Hemodinâmica/fisiologia , Hemostáticos/sangue , Cinética , Camundongos , Fator de Necrose Tumoral alfa/metabolismoRESUMO
BACKGROUND: Endothelin-1 (ET-1) is released after hyperkalemic cardioplegic arrest (CA) and reperfusion and may contribute to contractile dysfunction. ET-1 receptor transduction causes activation of protein kinase C (PKC) isoforms, which can cause differential intracellular events. The goal of this study was to determine which PKC isoforms contribute to myocyte contractile dysfunction with ET-1 and CA. METHODS AND RESULTS: Percent shortening (PERSHORT) and the time to 50% relaxation (T50) were measured in porcine (n =22) left ventricular myocytes, randomized (minimum: 30 cells/group) to normothermia: (cell media for 2 hours/37 degrees C), and CA: (2 hours/4 degrees C, 24 mEq K+ solution followed by reperfusion in cell media), ET-1/CA: (100 pM ET-1 during CA). Studies were performed in the presence and absence of PKC inhibitors (500 nM) against the classical (Beta-I, Beta-II, Gamma) and novel (Epsilon, Eta) isoforms (myocytes from a minimum of 3 pigs per inhibitor). CA reduced PERSHORT by approximately 35% from normothermia (P<0.05), which was further reduced with ET-1. PKC-Beta-II or PKC-Gamma inhibition increased PERSHORT from ET-1/CA as well as CA only (P<0.05). CA prolonged T50 by approximately 19% from normothermia (P<0.05) and was further prolonged with ET-1. Inhibition of the classical PKC isoforms reduced T50 from ET-1/CA (P<0.05). Inhibition of novel PKC isoforms did not yield similar effects on either PERSHORT or T50 with ET-1/CA. CONCLUSIONS: Inhibition of the classical PKC isoforms relieved the negative inotropic and lusitropic effects of ET-1 after CA. These findings provide mechanistic support for developing targeted inhibitory strategies with respect to ET-1 signaling and myocyte contractile dysfunction with cardioplegic arrest and reperfusion.
Assuntos
Endotelina-1/fisiologia , Parada Cardíaca Induzida/efeitos adversos , Contração Miocárdica/fisiologia , Traumatismo por Reperfusão Miocárdica/fisiopatologia , Miócitos Cardíacos/enzimologia , Proteína Quinase C/fisiologia , Animais , Células Cultivadas/efeitos dos fármacos , Células Cultivadas/enzimologia , Células Cultivadas/fisiologia , Endotelina-1/farmacologia , Ativação Enzimática/efeitos dos fármacos , Isoenzimas/antagonistas & inibidores , Isoenzimas/fisiologia , Contração Miocárdica/efeitos dos fármacos , Traumatismo por Reperfusão Miocárdica/enzimologia , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/fisiologia , Peptídeos/farmacologia , Proteína Quinase C/antagonistas & inibidores , Proteína Quinase C beta , Proteína Quinase C-épsilon/antagonistas & inibidores , Distribuição Aleatória , Receptores de Quinase C Ativada , Sus scrofa , TemperaturaRESUMO
LV myocardial remodeling is a structural hallmark of hypertensive hypertrophy, but molecular mechanisms driving this process are not well understood. The matrix metalloproteinases (MMPs) can cause myocardial remodeling in chronic disease states, but how MMP activity is altered with a mechanical load remains unknown. The present study quantified interstitial MMP activity after a discrete increase in LV load and dissected out the contributory role of the angiotensin II Type 1 receptor (AT1R). Pigs (38 kg) were randomized to undergo (1) increased LV load by insertion of an intra-aortic balloon pump (IABP) triggered at systole for 3 hours, then deactivated (n=11); (2) IABP and AT1R blockade (AT1RB; valsartan, 3 ng/kg/hr; n=6). MMP activity was directly measured in the myocardial interstitium using a validated inline digital fluorogenic microdialysis system. IABP engagement increased LV peak pressure from 92+/-3 to 113+/-5 and 123+/-7 mm Hg in the vehicle and AR1RB group, respectively, and remained elevated throughout the IABP period (P<0.05). With IABP disengagement, segmental shortening (% change from baseline of 0) remained depressed in the vehicle group (-32.2+/-11.8%, P<0.05) but returned to baseline in the AT1RB group (2.3+/-12.5%). MMP activity decreased with IABP in both groups. At IABP disengagement, a surge in MMP activity occurred in the vehicle group that was abrogated with AT1RB (3.03+/-0.85 versus 0.07+/-1.55 MMP units/hr, P<0.05). A transient increase in LV load caused a cyclic variation in interstitial MMP activity that is regulated in part by the AT1R. These temporally dynamic changes in MMP activity likely influence myocardial function and structure with increased LV load.
Assuntos
Hipertensão/enzimologia , Metaloproteinases da Matriz/metabolismo , Miocárdio/enzimologia , Receptor Tipo 1 de Angiotensina/fisiologia , Função Ventricular Esquerda , Bloqueadores do Receptor Tipo 1 de Angiotensina II/farmacologia , Animais , Hipertensão/fisiopatologia , Balão Intra-Aórtico , Suínos , Remodelação VentricularRESUMO
BACKGROUND: The matrix metalloproteinases (MMPs) contribute to regional remodeling after prolonged periods of ischemia and reperfusion (I/R), but specific MMP types activated during this process remain poorly understood. A novel class, the membrane-type MMPs (MT-MMPs), has been identified in the myocardium, but activity of these MMP types has not been assessed in vivo, particularly during I/R. METHODS AND RESULTS: Pigs (30 kg, n=8) were instrumented with microdialysis catheters to measure MT1-MMP activity in both ischemic and nonischemic (remote) myocardium. A validated MT1-MMP fluorogenic substrate was infused through the microdialysis system, and changes in fluorescence were reflective of MT1-MMP activity at steady state, during ischemia (90 minutes), and during reperfusion (120 minutes). At peak ischemia, MT1-MMP activity was increased by >40% in the ischemic region, with no change in the remote region, which persisted with reperfusion (P<0.05). After I/R, MT1-MMP abundance was increased by >50% (P<0.05). Differential centrifugation revealed that the endosomal fraction (which contains subcellular organelles) within the ischemic myocardium was associated with a >135% increase in MT1-MMP (P<0.05). Furthermore, in an isolated left ventricular myocyte model of I/R, hypoxia (simulated ischemia) induced a >70% increase in MT1-MMP abundance in myocytes, and confocal microscopy revealed MT1-MMP internalization during this time period and reemergence to the membrane with reperfusion. CONCLUSIONS: These unique results demonstrate that a specific MMP type, MT1-MMP, is increased in abundance and activity with I/R and is likely attributed, at least in part, to changes in intracellular trafficking.
Assuntos
Metaloendopeptidases/metabolismo , Isquemia Miocárdica/enzimologia , Traumatismo por Reperfusão Miocárdica/enzimologia , Miócitos Cardíacos/enzimologia , Transporte Proteico , Animais , Hipóxia Celular , Endossomos/enzimologia , Ventrículos do Coração , Metaloproteinases da Matriz Associadas à Membrana , Metaloendopeptidases/análise , Microdiálise , Microscopia Confocal , Isquemia Miocárdica/patologia , Traumatismo por Reperfusão Miocárdica/patologia , Miocárdio/enzimologia , Miócitos Cardíacos/ultraestrutura , Frações Subcelulares/enzimologia , Sus scrofa , Inibidor Tecidual de Metaloproteinase-1/análise , Inibidor Tecidual de Metaloproteinase-2/análise , Inibidor Tecidual de Metaloproteinase-3/análise , Inibidores Teciduais de Metaloproteinases/análise , Inibidor Tecidual 4 de MetaloproteinaseRESUMO
BACKGROUND: Whether mechanical restraint of the left ventricle (LV) can influence remodeling after myocardial infarction (MI) remains poorly understood. This study surgically placed a cardiac support device (CSD) over the entire LV and examined LV and myocyte geometry and function after MI. METHODS AND RESULTS: Post-MI sheep (35 to 45 kg; MI size, 23+/-2%) were randomized to placement of the CorCap CSD (Acorn Cardiovascular, Inc) (MI+CSD; n=6) or remained untreated (MI only; n=5). Uninstrumented sheep (n=10) served as controls. At 3 months after MI, LV end-diastolic volume (by MRI) was increased in the MI only group compared with controls (98+/-8 versus 43+/-4 mL; P<0.05). In the MI+CSD group, LV end-diastolic volume was lower than MI only values (56+/-7 mL; P<0.05) but remained higher than controls (P<0.05). Isolated LV myocyte shortening velocity was reduced by 35% from control values (P<0.05) in both MI groups. LV myocyte beta-adrenergic response was reduced with MI but normalized in the MI+CSD group. LV myocyte length increased in the MI group and was reduced in the MI+CSD group. Relative collagen content was increased and matrix metalloproteinase-9 was decreased within the MI border region of the CSD group. CONCLUSIONS: A CSD beneficially modified LV and myocyte remodeling after MI through both cellular and extracellular mechanisms. These findings provide evidence that nonpharmacological strategies can interrupt adverse LV remodeling after MI.
Assuntos
Coração Auxiliar , Infarto do Miocárdio/cirurgia , Miocárdio/patologia , Remodelação Ventricular , Actinas/análise , Animais , Colágeno/análise , Matriz Extracelular/fisiologia , Masculino , Metaloproteinase 2 da Matriz/análise , Metaloproteinase 9 da Matriz/análise , Contração Miocárdica , Infarto do Miocárdio/patologia , Infarto do Miocárdio/fisiopatologia , Miócitos Cardíacos/patologia , Miócitos Cardíacos/fisiologia , Ovinos , Inibidores Teciduais de Metaloproteinases/análise , Função Ventricular EsquerdaRESUMO
BACKGROUND: A cause-and-effect relationship exists between matrix metalloproteinase (MMP) induction and left ventricular (LV) remodeling after myocardial infarction (MI). Whether broad-spectrum MMP inhibition is necessary and the timing at which MMP inhibition should be instituted after MI remain unclear. This study examined the effects of MMP-1 and MMP-7-sparing inhibition (sMMPi) on regional and global LV remodeling when instituted before or after MI. METHODS AND RESULTS: Pigs instrumented with coronary snares and radiopaque markers within the area at risk were randomized to MI only (n=11) or sMMPi (PGE-530742, 10 mg/kg PO TID) begun 3 days before MI (n=11) or 3 days after MI (n=10). Eleven weight-matched noninstrumented pigs served as reference controls. At 10 days after MI, infarct size was similar between groups (47+/-3% of the area at risk). Marker area increased from baseline in the MI-only group (10+/-3%, P<0.05) but was unchanged with sMMPi. LV end-diastolic volume increased in the MI-only group (82+/-3 mL) compared with controls (56+/-3 mL, P<0.05) but was attenuated with pre-MI and post-MI sMMPi (69+/-3 and 69+/-4 mL, respectively, P<0.05). Collagen content increased in the infarct zone of the MI-only group (34+/-5%) compared with control (2+/-1%, P<0.05) but was reduced with pre-MI and post-MI sMMPi (24+/-1% and 23+/-2%, P<0.05). Collagen content increased in the border zone (12+/-2%) and decreased in the remote zone (3+/-1%) of the pre-MI sMMPi group compared with post-MI sMMPi values (7+/-1% and 5+/-1%, P<0.05). CONCLUSIONS: Inhibition of MMP-1 and -7 is not required to favorably influence LV remodeling after MI. Moreover, a temporal difference exists with respect to the timing of sMMPi and regional and global myocardial remodeling patterns after MI.
Assuntos
Inibidores de Metaloproteinases de Matriz , Infarto do Miocárdio/tratamento farmacológico , Inibidores de Proteases/uso terapêutico , Animais , Sistemas de Liberação de Medicamentos , Hemodinâmica , Imuno-Histoquímica , Isoenzimas/antagonistas & inibidores , Metaloproteinase 2 da Matriz/análise , Metaloproteinase 2 da Matriz/imunologia , Metaloproteinase 2 da Matriz/metabolismo , Infarto do Miocárdio/enzimologia , Infarto do Miocárdio/patologia , Infarto do Miocárdio/fisiopatologia , Miocárdio/patologia , Inibidores de Proteases/administração & dosagem , Suínos , Fatores de Tempo , Função Ventricular Esquerda , Remodelação Ventricular , CicatrizaçãoRESUMO
BACKGROUND: A mechanism for myocardial dysfunction after ischemia and reperfusion is Na(+)/H(+) exchanger activation. Although past in vivo models of limited ischemia and reperfusion intervals demonstrate that Na(+)/H(+) exchanger inhibition confers myocardial protection when administered at the onset of ischemia, the effect of Na(+)/H(+) exchanger inhibition on myocardial function after prolonged ischemia and reperfusion remains unknown. This investigation tested the hypothesis that Na(+)/H(+) exchanger inhibition instituted at reperfusion and after prolonged coronary occlusion in pigs would influence myocardial contractility independent of myocardial viability. METHODS: A coronary snare and sonomicrometry crystals were placed in pigs (n = 21, 32 kg). Coronary occlusion was instituted for 120 minutes followed by reperfusion for 180 minutes. At 105 minutes of ischemia, pigs were randomized to ischemia and reperfusion only (saline solution, n = 11) or Na(+)/H(+) exchanger inhibition (HOE-642, 3 mg/kg intravenously, n = 10). Myocardial injury was determined by tissue staining and measurement of plasma myocyte-specific enzymes. Myocardial contractility was determined by calculation of the regional end-systolic pressure-dimension relation (millimeters of mercury per centimeter) and by assessment of interregional shortening. RESULTS: Infarct size was not different between groups (39% +/- 6%, P =.26). Moreover, at 180 minutes of reperfusion, plasma troponin-I and creatine kinase MB values had increased to identical levels in the ischemia and reperfusion-only and Na(+)/H(+) exchanger inhibition groups (300 +/- 35 and 50 +/- 6 ng/mL, respectively). At 90 minutes of ischemia, regional end-systolic pressure-dimension relation decreased from baseline (5.7 +/- 0.5 versus 2.7 +/- 0.3, P <.05) in the area at risk. By 30 minutes of reperfusion, regional end-systolic pressure-dimension relation decreased further in the ischemia and reperfusion-only group (1.6 +/- 0.2, P <.05), but improved with Na(+)/H(+) exchanger inhibition (4.4 +/- 0.7, P <.05). CONCLUSIONS: Na(+)/H(+) exchanger inhibition instituted at reperfusion improved contractility independent of myocardial viability as assessed by absolute infarct size and myocyte-specific enzyme release. Thus, modulation of Na(+)/H(+) exchanger activity in the setting of prolonged ischemia and reperfusion may hold therapeutic potential.
Assuntos
Guanidinas/farmacologia , Contração Miocárdica/efeitos dos fármacos , Infarto do Miocárdio/patologia , Traumatismo por Reperfusão Miocárdica/prevenção & controle , Reperfusão Miocárdica/métodos , Trocadores de Sódio-Hidrogênio/antagonistas & inibidores , Sulfonas/farmacologia , Animais , Biópsia por Agulha , Modelos Animais de Doenças , Feminino , Testes de Função Cardíaca , Hemodinâmica , Infusões Intravenosas , Masculino , Contração Miocárdica/fisiologia , Infarto do Miocárdio/cirurgia , Reperfusão Miocárdica/efeitos adversos , Distribuição Aleatória , Recuperação de Função Fisiológica , Valores de Referência , Sensibilidade e Especificidade , Trocadores de Sódio-Hidrogênio/farmacologia , Suínos , Função Ventricular Esquerda/efeitos dos fármacos , Função Ventricular Esquerda/fisiologiaRESUMO
OBJECTIVE: Myocyte death occurs by necrosis and caspase-mediated apoptosis in the setting of myocardial infarction. In vitro studies suggest that caspase activation within myocytes causes contractile protein degradation without inducing cell death. Thus, caspase activation may evoke left ventricular remodeling through 2 independent processes post-myocardial infarction. However, the effects of caspase activation on left ventricular geometry post-myocardial infarction remain unclear. This project applied broad-spectrum caspase inhibition to a chronic porcine model of myocardial infarction. METHODS: Coronary snares and sonomicrometry crystals in remote and area-at-risk regions were placed in pigs (n = 22, 34 kg). Geometric measurements at end diastole and end systole, including left ventricular area by echocardiography and interregional distance by sonomicrometry, were obtained at baseline. Coronary occlusion was instituted for 60 minutes, followed by reperfusion and repeated geometric measurements at 7 days, including left ventriculography. At reperfusion, pigs were randomized to saline (n = 12) or caspase inhibition (n = 10, IDN6734, 2 mg/kg intravenously, then 2 mg x kg x h for 24 hours) at a dose that achieved desired plasma concentrations (790 +/- 142 ng/mL) as predicted by prior pharmacokinetic studies. RESULTS: Infarct size and 24-hour troponin-I values were not significantly different between the saline and caspase inhibition groups (51% +/- 8% vs 42% +/- 6% and 189 +/- 20 ng/mL vs 152 +/- 26 ng/mL, respectively, P >.10). At 7 days, end-diastole volume was increased in both groups compared with reference control values (47 +/- 1 mL, P <.05), but it was decreased with caspase inhibition (72 +/- 4 mL) compared with saline (84 +/- 4 mL, P <.05). Similarly, end-diastole and end-systole areas increased by 32% +/- 3% and 81% +/- 16% in the saline group but were attenuated with caspase inhibition (19% +/- 3% and 31% +/- 10%, respectively, P <.05). End-diastole interregional distance increased by 30% +/- 7% in the saline group but was attenuated with caspase inhibition (12% +/- 5%, P <.05). CONCLUSION: Despite equivalent degrees of myocardial injury, caspase inhibition reduced post-myocardial infarction left ventricular remodeling as evidenced by multiple, independent assessments of left ventricular dilation. Thus, caspase activation alters left ventricular geometry in the absence of significant effects on myocardial injury.
Assuntos
Inibidores de Caspase , Inibidores Enzimáticos/farmacologia , Infarto do Miocárdio/fisiopatologia , Miocárdio/enzimologia , Remodelação Ventricular/fisiologia , Animais , Apoptose , Caspases/metabolismo , Caspases/fisiologia , Circulação Coronária , Ecocardiografia , Contração Miocárdica , Infarto do Miocárdio/diagnóstico por imagem , Infarto do Miocárdio/enzimologia , Suínos , Função Ventricular EsquerdaRESUMO
BACKGROUND: Ischemia-reperfusion (IR) injury causes myocardial dysfunction in part through intracellular calcium overload. A recently described pharmacologic compound, MCC-135 (5-methyl-2-[1-piperazinyl] benzenesulfonic acid monohydrate, Mitsubishi Pharma Corporation), alters intracellular calcium levels. This project tested the hypothesis that MCC-135 would influence regional myocardial contractility when administered at reperfusion and after a prolonged period of ischemia. METHODS: A circumflex snare and sonomicrometry crystals within remote and area-at-risk regions were placed in pigs (n = 18, 32 kg). Coronary occlusion was instituted for 120 minutes followed by 180 minutes of reperfusion. At 105 minutes of ischemia pigs were randomly assigned to IR only (n = 11) or MCC-135 (IR-MCC [300 microg. kg(-1). h(-1), n = 7]) administered intravenously. Regional myocardial contractility was determined by calculation of the regional end-systolic pressure-dimension relation (RESPDR [mm Hg/cm]). Myocardial injury was determined by measurement of plasma levels of myocyte-specific enzymes. RESULTS: At 90 minutes ischemia, mean troponin-I was 35 +/- 8 ng/mL with no significant difference between groups. At 180 minutes reperfusion, heart rate was increased by 18% +/- 5% in the IR only group (p < 0.05) and was reduced by 11% +/- 4% with IR-MCC (p < 0.05). At 90 minutes ischemia RESPDR was reduced from baseline by 51% +/- 6% (p < 0.05). By 30 minutes reperfusion, reductions in RESPDR were attenuated with IR-MCC compared with IR only values. The CK-MB levels were increased at 180 minutes reperfusion in the IR only group (52 +/- 9 ng/mL) compared with baseline (6 +/- 1 ng/mL, p < 0.05) but were attenuated with IR-MCC (24 +/- 4 ng/mL, p < 0.05) compared with IR only values. CONCLUSIONS: Despite similar degrees of injury at 90 minutes ischemia MCC-135 improved regional contractility and reduced the egress of CK-MB. Moreover MCC-135 was associated with decreased heart rate, a determinant of myocardial oxygen demand. Pharmacologic modulation of calcium transport ameliorates myocardial dysfunction in the acute IR period.
Assuntos
Cálcio/metabolismo , Creatina Quinase/sangue , Isoenzimas/sangue , Contração Miocárdica , Traumatismo por Reperfusão Miocárdica/fisiopatologia , Animais , Benzenossulfonatos/farmacologia , Transporte Biológico/efeitos dos fármacos , Creatina Quinase Forma MB , Frequência Cardíaca/efeitos dos fármacos , Contração Miocárdica/efeitos dos fármacos , Traumatismo por Reperfusão Miocárdica/metabolismo , Piperazinas/farmacologia , Suínos , Troponina I/sangueRESUMO
BACKGROUND: Exposure of left ventricular (LV) myocytes to simulated hyperkalemic cardioplegic arrest (HCA) has been demonstrated to perturb ionic homeostasis and adversely affect myocyte contractility on rewarming. Altered ionic homeostasis can cause cytosolic activation of the caspases. While caspases participate in apoptosis, these proteases can degrade myocyte contractile proteins, and thereby alter myocyte contractility. Accordingly, this study tested the hypothesis that caspase inhibition during HCA would attenuate the degree of myocyte contractile dysfunction upon rewarming, independent of a loss in myocyte viability. METHODS: Porcine (n = 8) LV myocytes were isolated and assigned to the following treatment groups: normothermic control: incubation in cell culture media for 2 hours at 37 degrees C; HCA only: incubation for 2 hours in hypothermic HCA solution (4 degrees C, 24 mEq K(+)); or incubation in hypothermic HCA solution supplemented with 10 microM of the caspase inhibitor, z-VAD (z-Val-Ala-Asp-fluoromethyl-ketone, HCA+zVAD). Myocyte viability, assayed as a function of mitochondrial function, was determined to be similar in the normothermic and both HCA groups. RESULTS: The HCA caused a significant reduction in myocyte shortening velocity compared with normothermic control values (41 +/- 6 versus 86 +/- 8 microm/s, p < 0.05). The HCA+zVAD group had significantly improved myocyte shortening velocity compared with the HCA only group (63 +/- 7 microm/s, p < 0.05). CONCLUSIONS: Independent of changes in viability, caspase inhibition attenuated myocyte contractile dysfunction after HCA and rewarming. Thus, caspase activation during HCA contributes, at least in part, to impaired myocyte contractility with rewarming. Supplementation of HCA with caspase inhibitors may provide a means to preserve myocyte contractile function after cardioplegic arrest.
Assuntos
Clorometilcetonas de Aminoácidos/farmacologia , Inibidores de Caspase , Parada Cardíaca Induzida , Células Musculares/fisiologia , Contração Miocárdica/efeitos dos fármacos , Animais , Soluções Cardioplégicas , Caspases/fisiologia , Sobrevivência Celular , Células Cultivadas , Parada Cardíaca Induzida/métodos , Humanos , Hiperpotassemia/fisiopatologia , Hipotermia Induzida , Soluções Isotônicas , Células Musculares/efeitos dos fármacos , Contração Miocárdica/fisiologia , Distribuição Aleatória , Reaquecimento , Solução de Ringer , SuínosRESUMO
The matrix metalloproteinases (MMPs), in particular, membrane type 1 MMP (MT1-MMP), are increased in the context of myocardial ischemia and reperfusion (I/R) and likely contribute to myocardial dysfunction. One potential upstream induction mechanism for MT1-MMP is endothelin (ET) release and subsequent protein kinase C (PKC) activation. Modulation of ET and PKC signaling with respect to MT1-MMP activity with I/R has yet to be explored. Accordingly, this study examined in vivo MT1-MMP activation during I/R following modification of ET signaling and PKC activation. With the use of a novel fluorogenic microdialysis system, myocardial interstitial MT1-MMP activity was measured in pigs (30 kg; n = 9) during I/R (90 min I/120 min R). Local ET(A) receptor antagonism (BQ-123, 1 microM) and PKC inhibition (chelerythrine, 1 microM) were performed in parallel microdialysis probes. MT1-MMP activity was increased during I/R by 122 +/- 10% (P < 0.05) and was unchanged from baseline with ET antagonism and/or PKC inhibition. Selective PKC isoform induction occurred such that PKC-betaII increased by 198 +/- 31% (P < 0.05). MT1-MMP phosphothreonine, a putative PKC phosphorylation site, was increased by 121 +/- 8% (P < 0.05) in the I/R region. These studies demonstrate for the first time that increased interstitial MT1-MMP activity during I/R is a result of the ET/PKC pathway and may be due to enhanced phosphorylation of MT1-MMP. These findings identify multiple potential targets for modulating a local proteolytic pathway operative during I/R.
Assuntos
Endotelinas/fisiologia , Metaloproteinase 14 da Matriz/metabolismo , Traumatismo por Reperfusão Miocárdica/fisiopatologia , Transdução de Sinais/fisiologia , Animais , Antagonistas do Receptor de Endotelina B , Corantes Fluorescentes , Imunoprecipitação , Isoenzimas/metabolismo , Microdiálise , Traumatismo por Reperfusão Miocárdica/enzimologia , Peptídeos Cíclicos/farmacologia , Fosforilação , Proteína Quinase C/metabolismo , Receptor de Endotelina B/metabolismo , Volume Sistólico/fisiologia , Suínos , Treonina/metabolismoRESUMO
BACKGROUND: Left ventricular (LV) remodeling after myocardial infarction (MI) commonly causes infarct expansion (IE). This study sought to interrupt IE through microinjections of a biocompatible composite material into the post-MI myocardium. METHODS: MI was created in 21 pigs (coronary ligation). Radiopaque markers (2-mm diameter) were placed for IE (fluoroscopy). Pigs were randomized for microinjections (25 injections; 2- x 2-cm array; 200 microL/injection) at 7 days post-MI of a fibrin-alginate composite (Fib-Alg; fibrinogen, fibronectin, factor XIII, gelatin-grafted alginate, thrombin; n = 11) or saline (n = 10). RESULTS: At 7 days after injection (14 days post-MI), LV posterior wall thickness was higher in the Fib-Alg group than in the saline group (1.07 +/- 0.11 vs 0.69 +/- 0.07 cm, respectively, p = 0.002). At 28 days post-MI, the area within the markers (IE) increased from baseline (1 cm2) in the saline (1.71 +/- 0.13 cm2, p = 0.010) and Fib-Alg groups (1.44 +/- 0.23 cm2, p < 0.001). However, the change in IE at 21 and 28 days post-MI was reduced in the Fib-Alg group (p=0.043 and p=0.019). Total collagen content within the MI region was similar in the saline and Fib-Alg groups (12.8 +/- 1.7 and 11.6 +/- 1.5 microg/mg, respectively, p = NS). However, extractable collagen, indicative of solubility, was lower in the Fib-Alg group than the saline group (59.1 +/- 3.5 vs 71.0 +/- 6.1 microg/mL, p = 0.020). CONCLUSIONS: Targeted myocardial microinjection of the biocomposite attenuated the post-MI decrease in LV wall thickness and infarct expansion. Thus, intraoperative microinjections of biocompatible material may provide a novel approach for interrupting post-MI LV remodeling.
Assuntos
Materiais Biocompatíveis/administração & dosagem , Infarto do Miocárdio/fisiopatologia , Infarto do Miocárdio/terapia , Remodelação Ventricular , Animais , Modelos Animais de Doenças , Feminino , Injeções Intralesionais , Masculino , Microinjeções , Infarto do Miocárdio/patologia , Probabilidade , Distribuição Aleatória , Valores de Referência , Sensibilidade e Especificidade , Suínos , Remodelação Ventricular/fisiologiaRESUMO
BACKGROUND: A cause-effect relationship has been established between MMP activation and left ventricular (LV) remodeling following myocardial infarction. The goal of the present study was to examine a selective MMP inhibitor (sMMPi) strategy that effectively spared MMP-1, -3, and -7 with effect to regional and global left ventricular remodeling in a pig model of myocardial infarction. METHODS AND RESULTS: Pigs instrumented with coronary snares and radiopaque markers within the area at risk were randomized to myocardial infarction-only (n = 10) or sMMPi (PGE-530742, 1 mg/kg TID) begun 3 days prior to myocardial infarction. Ten weight-matched noninstrumented pigs served as reference controls. Left ventricular end-diastolic volume in the myocardial infarction-only group was increased from baseline (81 +/- 3 mL versus 55 +/- 4 mL, respectively, P < 0.05) but was attenuated with sMMPi (67 +/- 3 mL, P < 0.05). Fractional area of shortening of marker area was decreased in the myocardial infarction-only group (change from baseline -63 +/- 10%, P < 0.05) but this effect was attenuated with sMMPi (-28 +/- 14%, P < 0.05), indicative of less dyskinesis of the infarct region with sMMPi. Wall stress was reduced within both the septal and posterior wall regions with sMMPi. Myocardial MMP-2 activity was decreased in both remote and border areas of sMMPi-treated samples compared with myocardial infarction-only values, consistent with pharmacologic MMP inhibition. CONCLUSIONS: Selective MMP inhibition favorably affected regional myocardial geometry and decreased left ventricular dilation post-myocardial infarction. This study suggests that a strategy of selective MMP inhibition of a limited array of MMPs may be an achievable goal in preventing pathologic left ventricular remodeling post-myocardial infarction.
Assuntos
Inibidores de Metaloproteinases de Matriz , Metaloproteinases da Matriz/fisiologia , Infarto do Miocárdio/tratamento farmacológico , Inibidores de Proteases/uso terapêutico , Inibidores Teciduais de Metaloproteinases/uso terapêutico , Remodelação Ventricular/efeitos dos fármacos , Animais , Modelos Animais de Doenças , Ecocardiografia , Fluoroscopia , Infarto do Miocárdio/complicações , Infarto do Miocárdio/enzimologia , Infarto do Miocárdio/patologia , Infarto do Miocárdio/fisiopatologia , Miocárdio/patologia , Inibidores de Proteases/administração & dosagem , Distribuição Aleatória , Suínos , Fatores de Tempo , Inibidores Teciduais de Metaloproteinases/farmacocinética , Inibidores Teciduais de Metaloproteinases/farmacologia , Remodelação Ventricular/fisiologiaRESUMO
BACKGROUND: Increased myocardial interstitial levels of endothelin (ET) occur during cardioplegic arrest (CA) and may contribute to contractile dysfunction. Endothelin receptor transduction involves the protein kinase-C (PKC) family comprised of multiple isoforms with diverse functions. Which PKC isoforms may be involved in ET-induced contractile dysfunction after CA remains unknown. METHODS: Shortening velocity was measured in isolated left ventricular porcine myocytes and randomized (minimum of 30 per group): normothermia (cell culture media for 2 hours at 37 degrees C); CA (2 hours in CA solution [4 degrees C, 24 mEq K+] followed by reperfusion in cell media); ET/CA (100 pM ET incubated during CA and reperfusion). These studies were carried out in the presence and absence of PKC inhibitors (500 nM) and directed against members of the classical PKC subfamily (beta I, beta II, gamma) and the novel subfamily (epsilon, eta). RESULTS: Cardiac arrest reduced shortening velocity by approximately 50%, which was further reduced in the presence of ET. Inhibition of either the beta II or gamma PKC isoform significantly increased shortening velocity from ET/CA as well as CA only values. In separate studies (n = 3), total beta II and phosphorylated beta II increased by over 150% with ET/CA (p < 0.05). Taken together, these results suggest that a predominant intracellular effector for the negative contractile effects mediated by ET in the context of CA is the PKC isoform beta II. CONCLUSIONS: Targeted inhibition of specific PKC isoforms relieves the negative inotropic effects of ET after simulated CA. These findings provide important mechanistic support for the development of targeted inhibitory strategies with respect to ET signaling and myocyte contractile dysfunction in the context of CA and reperfusion.
Assuntos
Endotelinas/farmacologia , Parada Cardíaca Induzida , Isoenzimas/fisiologia , Contração Miocárdica/efeitos dos fármacos , Miócitos Cardíacos/fisiologia , Proteína Quinase C/fisiologia , Animais , Ativação Enzimática , Reperfusão Miocárdica , Miócitos Cardíacos/enzimologia , Receptores de Quinase C Ativada , Receptores de Superfície Celular/fisiologia , SuínosRESUMO
Hyperkalemic cardioplegic arrest (HCA) and rewarming evokes postoperative myocyte contractile dysfunction, a phenomenon of particular importance in settings of preexisting left ventricular (LV) failure. Caspases are intracellular proteolytic enzymes recently demonstrated to degrade myocardial contractile proteins. This study tested the hypothesis that myocyte contractile dysfunction induced by HCA could be ameliorated with caspase inhibition in the setting of compromised myocardial function. LV myocytes were isolated from control pigs (n = 9, 30 kg) or pigs with LV failure induced by rapid pacing (n = 6, 240 bpm for 21 days) and were randomized to the following: (1) normothermia (2003 myocytes), incubation in cell culture medium for 2 hours at 37 degrees C; (2) HCA only (506 myocytes), incubation for 2 hours in hypothermic HCA solution (4 degrees C, 24 mEq K); or (3) HCA + z-VAD, incubation in hypothermic HCA solution supplemented with 10 microM of the caspase inhibitor z-VAD (z-Val-Ala-Asp-fluoromethyl-ketone, 415 myocytes). Inotropic responsiveness was examined using beta-adrenergic stimulation (25 nM isoproterenol). Ambient normothermic myocyte shortening velocity (microm/s) was reduced with LV failure compared with control values (54 +/- 2 versus 75 +/- 2, respectively, P < 0.05). Following HCA, shortening velocity decreased in the LV failure and control groups (27 +/- 5 and 45 +/- 3, P < 0.05). Institution of z-VAD increased myocyte shortening velocity following HCA in both the LV failure and control groups (49 +/- 5 and 65 +/- 5, P < 0.05). Moreover, HCA supplementation with z-VAD increased beta-adrenergic responsiveness in both groups compared with HCA-only values. This study provides proof of concept that caspase activity contributes to myocyte contractile dysfunction following simulated HCA. Pharmacologic caspase inhibition may hold particular relevance in the execution of cardiac surgical procedures requiring HCA in the context of preexisting LV failure.