Targeted inactivation of Gαi does not alter cardiac function or β-adrenergic sensitivity

2001 ◽  
Vol 280 (2) ◽  
pp. H569-H575 ◽  
Author(s):  
Mohit Jain ◽  
Chee Chew Lim ◽  
Kohzo Nagata ◽  
Vannessa M. Davis ◽  
David S. Milstone ◽  
...  
Keyword(s):  
Cardiac Function ◽  
Ventricular Myocytes ◽  
Ex Vivo ◽  
Cardiac Response ◽  
Similar Degree ◽  
Wild Type ◽  
Isolated Hearts ◽  
Targeted Inactivation ◽  
Contractile Performance

Inhibitory Gαi protein increases in the myocardium during hypertrophy and has been associated with β-adrenergic receptor (β-AR) desensitization, contractile dysfunction, and progression of cardiac disease. The role of Gαi proteins in mediating basal cardiac function and β-AR response in nonpathological myocardium, however, is uncertain. Transgenic mice with targeted inactivation of Gαi2 or Gαi3 were examined for in vivo cardiac function with the use of conscious echocardiography and for ex vivo cardiac response to inotropic stimulation with the use of Langendorff blood-perfused isolated hearts and adult ventricular cardiomyocytes. Echocardiography revealed that percent fractional shortening and heart rate were similar among wild-type, Gαi2 -null, and Gαi3 -null mice. Comparable baseline diastolic and contractile performance was also observed in isolated hearts and isolated ventricular myocytes from wild-type mice and mice lacking Gαi proteins. Isoproterenol infusion enhanced diastolic and contractile performance to a similar degree in wild-type, Gαi2 -null, and Gαi3 -null mice. These data demonstrate no observable role for inhibitory G proteins in mediating basal cardiac function or sensitivity to β-AR stimulation in nonpathological myocardium.

scholarly journals Chronic coexistence of two troponin T isoforms in adult transgenic mouse cardiomyocytes decreased contractile kinetics and caused dilatative remodeling

2012 ◽  
Vol 303 (1) ◽  
pp. C24-C32 ◽  
Author(s):  
Zhi-Bin Yu ◽  
Hongguang Wei ◽  
J.-P. Jin
Keyword(s):  
Transgenic Mouse ◽  
Troponin T ◽  
Ex Vivo ◽  
Cellular Level ◽  
Wild Type ◽  
Negative Effects ◽  
Adult Cardiomyocytes ◽  
Cross Bridges ◽  
Contractile Performance

Our previous in vivo and ex vivo studies suggested that coexistence of two or more troponin T (TnT) isoforms in adult cardiac muscle decreased cardiac function and efficiency (Huang QQ, Feng HZ, Liu J, Du J, Stull LB, Moravec CS, Huang X, Jin JP, Am J Physiol Cell Physiol 294: C213–C22, 2008; Feng HZ, Jin JP, Am J Physiol Heart Circ Physiol 299: H97–H105, 2010). Here we characterized Ca2+-regulated contractility of isolated adult cardiomyocytes from transgenic mice coexpressing a fast skeletal muscle TnT together with the endogenous cardiac TnT. Without the influence of extracellular matrix, coexistence of the two TnT isoforms resulted in lower shortening amplitude, slower shortening and relengthening velocities, and longer relengthening time. The level of resting cytosolic Ca2+ was unchanged, but the peak Ca2+ transient was lowered and the durations of Ca2+ rising and decaying were longer in the transgenic mouse cardiomyocytes vs. the wild-type controls. Isoproterenol treatment diminished the differences in shortening amplitude and shortening and relengthening velocities, whereas the prolonged durations of relengthening and Ca2+ transient in the transgenic cardiomyocytes remained. At rigor state, a result from depletion of Ca2+, resting sarcomere length of the transgenic cardiomyocytes became shorter than that in wild-type cells. Inhibition of myosin motor diminished this effect of TnT function on cross bridges. The length but not width of transgenic cardiomyocytes was significantly increased compared with the wild-type controls, corresponding to longitudinal addition of sarcomeres and dilatative remodeling at the cellular level. These dominantly negative effects of normal fast TnT demonstrated that chronic coexistence of functionally distinct variants of TnT in adult cardiomyocytes reduces contractile performance with pathological consequences.

Attenuated cardiac function degradation in ex vivo pig hearts

2019 ◽  
Vol 43 (3) ◽  
pp. 173-179
Author(s):  
Benjamin Kappler ◽  
Sjoerd van Tuijl ◽  
Bülent Ergin ◽  
Louis Fixsen ◽  
Marco Stijnen ◽  
...  
Keyword(s):  
Cardiac Function ◽  
Ex Vivo ◽  
Heart Function ◽  
Blood Parameters ◽  
Diagnostic Tools ◽  
Control Group ◽  
Isolated Hearts ◽  
Cardiac Hemodynamics ◽  
Cardiac Pump Function

Isolated hearts offer the opportunity to evaluate heart function, treatments, and diagnostic tools without in vivo factor interference. However, the early loss of cardiac function and edema occur over time and do limit the duration of the experiment. This research focuses on delaying these limitations using optimal blood control. This study examines whether blood conditioning by means of the combination of blood predilution and hemodialysis can significantly reduce cardiac function degradation. Slaughterhouse porcine hearts were revived in the PhysioHeart™ platform to restore physiological cardiac performance. Twelve hearts were divided into a control group and a dialysis group; in the latter group, hemodialysis was attached to the blood reservoir. Cardiac hemodynamics and blood parameters were recorded and evaluated. Blood conditioning significantly reduced the loss of cardiac pump function (control group vs dialysis group, −14.9 ± 6.3%/h vs −9.7 ± 2.7%/h) and loss of cardiac output (control group vs dialysis group, −11.8 ± 3.4%/h vs −5.9 ± 2.0%/h). Hemodialysis resulted in physiological and stable blood parameters, whereas in the control group ions reached pathological values, while interstitial edema still occurred. The combination of blood predilution and hemodialysis significantly attenuated ex vivo cardiac function degradation and delayed the loss of cardiac hemodynamics. We hypothesized that besides electrolyte and metabolic control, the hemodialysis-accompanied increase in hematocrit resulted in improved oxygen transport. This could have temporarily compensated the deleterious effect of an increased oxygen-diffusion distance due to edema in the dialysis group and resulted in less progression of cell decay. Clinically validated measures delaying edema might improve the effectiveness of the PhysioHeart™ platform.

scholarly journals Platelet necrosis mediates ischemic stroke outcome in mice

Blood ◽  
2020 ◽  
Vol 135 (6) ◽  
pp. 429-440 ◽  
Author(s):  
Frederik Denorme ◽  
Bhanu Kanth Manne ◽  
Irina Portier ◽  
Alicia S. Eustes ◽  
Yasuhiro Kosaka ◽  
...  
Keyword(s):  
Ischemic Stroke ◽  
Ischemia Reperfusion Injury ◽  
Ex Vivo ◽  
Infarct Volume ◽  
Ischemia Reperfusion ◽  
Similar Degree ◽  
Cyclophilin D ◽  
Wild Type ◽  
Neutrophil Depletion

Abstract Dysregulated platelet functions contribute to the development and progression of ischemic stroke. Utilizing mice with a platelet-specific deletion of cyclophilin D (CypD), a mediator of necrosis, we found that platelet necrosis regulates tissue damage and outcomes during ischemic stroke in vivo. Mice with loss of CypD in platelets (CypDplt−/−mice) exhibited significantly enhanced cerebral blood flow, improved neurological and motor functions, and reduced ischemic stroke infarct volume after cerebral ischemia-reperfusion injury. These effects were attributable, at least in part, to platelet-neutrophil interactions. Twenty-four hours after stroke, significantly more circulating platelet-neutrophil aggregates (PNAs) were found in CypDplt+/+ mice. Underscoring the role of platelet necrosis in PNA formation, we observed a significant number of phosphatidylserine (PS)+ platelets in PNAs in CypDplt+/+ mice. In contrast, significantly fewer platelets in PNAs were PS+ in CypDplt−/− counterparts. Accordingly, mice with CypD-deficient platelets had fewer neutrophils and PNAs recruited to their brain following stroke relative to wild-type counterparts. Neutrophil depletion in wild-type mice conferred protection from ischemic stroke to a similar degree as observed in mice with CypD-deficient platelets. Neutrophil depletion in CypDplt−/− mice did not further reduce infarct size. Transmission electron microscopy of ex vivo–formed PNAs revealed a propensity of necrotic platelets to interact with neutrophils. These results suggest that necrotic platelets interact with neutrophils to exacerbate brain injury during ischemic stroke. Because inhibiting platelet necrosis does not compromise hemostasis, targeting platelet CypD may be a potential therapeutic strategy to limit brain damage following ischemic stroke.

scholarly journals Differential effect of anesthetics on mucociliary clearance in vivo in mice

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Kyle S. Feldman ◽  
Eunwon Kim ◽  
Michael J. Czachowski ◽  
Yijen Wu ◽  
Cecilia W. Lo ◽  
...  
Keyword(s):  
Mucociliary Clearance ◽  
Defense Mechanism ◽  
Ex Vivo ◽  
Beat Frequency ◽  
Ciliary Beat Frequency ◽  
Wild Type ◽  
Sulfur Colloid ◽  
Ct System

AbstractRespiratory mucociliary clearance (MCC) is a key defense mechanism that functions to entrap and transport inhaled pollutants, particulates, and pathogens away from the lungs. Previous work has identified a number of anesthetics to have cilia depressive effects in vitro. Wild-type C57BL/6 J mice received intra-tracheal installation of 99mTc-Sulfur colloid, and were imaged using a dual-modality SPECT/CT system at 0 and 6 h to measure baseline MCC (n = 8). Mice were challenged for one hour with inhalational 1.5% isoflurane, or intraperitoneal ketamine (100 mg/kg)/xylazine (20 mg/kg), ketamine (0.5 mg/kg)/dexmedetomidine (50 mg/kg), fentanyl (0.2 mg/kg)/1.5% isoflurane, propofol (120 mg/Kg), or fentanyl/midazolam/dexmedetomidine (0.025 mg/kg/2.5 mg/kg/0.25 mg/kg) prior to MCC assessment. The baseline MCC was 6.4%, and was significantly reduced to 3.7% (p = 0.04) and 3.0% (p = 0.01) by ketamine/xylazine and ketamine/dexmedetomidine challenge respectively. Importantly, combinations of drugs containing fentanyl, and propofol in isolation did not significantly depress MCC. Although no change in cilia length or percent ciliation was expected, we tried to correlate ex-vivo tracheal cilia ciliary beat frequency and cilia-generated flow velocities with MCC and found no correlation. Our results indicate that anesthetics containing ketamine (ketamine/xylazine and ketamine/dexmedetomidine) significantly depress MCC, while combinations containing fentanyl (fentanyl/isoflurane, fentanyl/midazolam/dexmedetomidine) and propofol do not. Our method for assessing MCC is reproducible and has utility for studying the effects of other drug combinations.

Treatment of type 2 diabetic db/db mice with a novel PPARγ agonist improves cardiac metabolism but not contractile function

2004 ◽  
Vol 286 (3) ◽  
pp. E449-E455 ◽  
Author(s):  
Andrew N. Carley ◽  
Lisa M. Semeniuk ◽  
Yakhin Shimoni ◽  
Ellen Aasum ◽  
Terje S. Larsen ◽  
...  
Keyword(s):  
Contractile Function ◽  
Ex Vivo ◽  
Metabolic Changes ◽  
Pparγ Agonist ◽  
Perfused Hearts ◽  
Type 2 Diabetic ◽  
Contractile Performance ◽  
Exogenous Substrates

Hearts from insulin-resistant type 2 diabetic db/db mice exhibit features of a diabetic cardiomyopathy with altered metabolism of exogenous substrates and reduced contractile performance. Therefore, the effect of chronic oral administration of 2-(2-(4-phenoxy-2-propylphenoxy)ethyl)indole-5-acetic acid (COOH), a novel ligand for peroxisome proliferator-activated receptor-γ that produces insulin sensitization, to db/db mice (30 mg/kg for 6 wk) on cardiac function was assessed. COOH treatment reduced blood glucose from 27 mM in untreated db/db mice to a normal level of 10 mM. Insulin-stimulated glucose uptake was enhanced in cardiomyocytes from COOH-treated db/db hearts. Working perfused hearts from COOH-treated db/db mice demonstrated metabolic changes with enhanced glucose oxidation and decreased palmitate oxidation. However, COOH treatment did not improve contractile performance assessed with ex vivo perfused hearts and in vivo by echocardiography. The reduced outward K+ currents in diabetic cardiomyocytes were still attenuated after COOH. Metabolic changes in COOH-treated db/db hearts are most likely indirect, secondary to changes in supply of exogenous substrates in vivo and insulin sensitization.

Abstract P283: Parkin Mediates Mitophagy in Cardioprotection

2011 ◽  
Vol 109 (suppl_1) ◽  
Author(s):  
Allen M Andres ◽  
Chengqun Huang ◽  
Eric P Ratliff ◽  
Genaro Hernandez ◽  
Pamela Lee ◽  
...  
Keyword(s):  
Ex Vivo ◽  
Wild Type ◽  
Simulated Ischemia ◽  
Selective Targeting ◽  
Cardioprotective Effects ◽  
Mitochondrial Turnover ◽  
First Time

Autophagy-dependent mitochondrial turnover in response to cellular stress is necessary for maintaining cellular homeostasis. However, the mechanisms that govern the selective targeting of damaged mitochondria are poorly understood. Parkin, an E3 ubiquitin ligase, has been shown to be essential for the selective clearance of damaged mitochondria. Parkin is expressed in the heart, yet its function has not been investigated in the context of cardioprotection. We previously reported that autophagy is required for cardioprotection by ischemic preconditioning (IPC). In the present study, we used simulated ischemia in vitro and IPC in hearts (in vivo and ex vivo) to investigate the role of Parkin in mediating cardioprotection. In HL-1 cells, simulated ischemia induced Parkin translocation to mitochondria and mitochondrial elimination. Mitochondrial loss was blunted in Atg5-deficient cells, revealing the requirement for autophagy in mitochondrial elimination. Consistent with previous reports implicating p62/SQSTM1 in mitophagy, we found that downregulation of p62 attenuated mitophagy and exacerbated cell death in HL-1 cardiomyocytes subjected to simulated ischemia. While wild type mice showed p62 translocation to mitochondria after IPC, Parkin knockout mice exhibited attenuated translocation of p62 to mitochondria. Importantly, ablation of Parkin in mice abolished the cardioprotective effects of IPC. These results reveal for the first time the crucial role of Parkin and mitophagy in cardioprotection.

scholarly journals Influence of genetic background on ex vivo and in vivo cardiac function in several commonly used inbred mouse strains

2010 ◽  
Vol 42A (2) ◽  
pp. 103-113 ◽  
Author(s):  
Matthew S. Barnabei ◽  
Nathan J. Palpant ◽  
Joseph M. Metzger
Keyword(s):  
Cardiac Function ◽  
Genetic Divergence ◽  
Ex Vivo ◽  
Inbred Mouse ◽  
Critical Role ◽  
Inbred Strains ◽  
Mouse Strains ◽  
Inbred Mouse Strains ◽  
Cardiac Decompensation

Inbred mouse strains play a critical role in biomedical research. Genetic homogeneity within inbred strains and their general amenability to genetic manipulation have made them an ideal resource for dissecting the physiological function(s) of individual genes. However, the inbreeding that makes inbred mice so useful also results in genetic divergence between them. This genetic divergence is often unaccounted for but may be a confounding factor when comparing studies that have utilized distinct inbred strains. Here, we compared the cardiac function of C57BL/6J mice to seven other commonly used inbred mouse strains: FVB/NJ, DBA/2J, C3H/HeJ, BALB/cJ, 129X1/SvJ, C57BL/10SnJ, and 129S1/SvImJ. The assays used to compare cardiac function were the ex vivo isolated Langendorff heart preparation and in vivo real-time hemodynamic analysis using conductance micromanometry. We report significant strain-dependent differences in cardiac function between C57BL/6J and other commonly used inbred strains. C57BL/6J maintained better cardiac function than most inbred strains after ex vivo ischemia, particularly compared with 129S1/SvImJ, 129X1/SvJ, and C57BL/10SnJ strains. However, during in vivo acute hypoxia 129X1/SvJ and 129S1/SvImJ maintained relatively normal cardiac function, whereas C57BL/6J animals showed dramatic cardiac decompensation. Additionally, C3H/HeJ showed rapid and marked cardiac decompensation in response to esmolol infusion compared with effects of other strains. These findings demonstrate the complex effects of genetic divergence between inbred strains on cardiac function. These results may help inform analysis of gene ablation or transgenic studies and further demonstrate specific quantitative traits that could be useful in discovery of genetic modifiers relevant to cardiac health and disease.

scholarly journals Loss of Sirt3 accelerates arterial thrombosis by increasing formation of neutrophil extracellular traps and plasma tissue factor activity

2018 ◽  
Vol 114 (8) ◽  
pp. 1178-1188 ◽  
Author(s):  
Daniel S Gaul ◽  
Julien Weber ◽  
Lambertus J van Tits ◽  
Susanna Sluka ◽  
Lisa Pasterk ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Myocardial Infarction ◽  
Bone Marrow ◽  
Arterial Thrombosis ◽  
Ex Vivo ◽  
Neutrophil Extracellular Traps ◽  
Wild Type ◽  
Rotational Thromboelastometry ◽  
Extracellular Traps

AbstractAimsSirtuin 3 (Sirt3) is a mitochondrial, nicotinamide adenine dinucleotide (NAD+)-dependent deacetylase that reduces oxidative stress by activation of superoxide dismutase 2 (SOD2). Oxidative stress enhances arterial thrombosis. This study investigated the effects of genetic Sirt3 deletion on arterial thrombosis in mice in an inflammatory setting and assessed the clinical relevance of these findings in patients with ST-elevation myocardial infarction (STEMI).Methods and resultsUsing a laser-induced carotid thrombosis model with lipopolysaccharide (LPS) challenge, in vivo time to thrombotic occlusion in Sirt3−/− mice (n = 6) was reduced by half compared to Sirt3+/+ wild-type (n = 8, P < 0.01) controls. Ex vivo analyses of whole blood using rotational thromboelastometry revealed accelerated clot formation and increased clot stability in Sirt3−/− compared to wild-type blood. rotational thromboelastometry of cell-depleted plasma showed accelerated clotting initiation in Sirt3−/− mice, whereas overall clot formation and firmness remained unaffected. Ex vivo LPS-induced neutrophil extracellular trap formation was increased in Sirt3−/− bone marrow-derived neutrophils. Plasma tissue factor (TF) levels and activity were elevated in Sirt3−/− mice, whereas plasma levels of other coagulation factors and TF expression in arterial walls remained unchanged. SOD2 expression in bone marrow -derived Sirt3−/− neutrophils was reduced. In STEMI patients, transcriptional levels of Sirt3 and its target SOD2 were lower in CD14+ leukocytes compared with healthy donors (n = 10 each, P < 0.01).ConclusionsSirt3 loss-of-function enhances experimental thrombosis in vivo via an increase of neutrophil extracellular traps and elevation of TF suggesting thrombo-protective effects of endogenous Sirt3. Acute coronary thrombosis in STEMI patients is associated with lower expression levels of SIRT3 and SOD2 in CD14+ leukocytes. Therefore, enhancing SIRT3 activity by pan-sirtuin activating NAD+-boosters may provide a novel therapeutic target to prevent or treat thrombotic arterial occlusion in myocardial infarction or stroke.

Modulation of mouse cardiac function in vivo by eNOS and ANP

2000 ◽  
Vol 278 (3) ◽  
pp. H971-H981 ◽  
Author(s):  
Robert Gyurko ◽  
Peter Kuhlencordt ◽  
Mark C. Fishman ◽  
Paul L. Huang
Keyword(s):  
Cardiac Function ◽  
Isolated Heart ◽  
Cardiac Contractility ◽  
Enos Gene ◽  
Wild Type ◽  
Adrenergic Stimulation ◽  
Microvascular Endothelium ◽  
Ventricular Relaxation ◽  
Nos Inhibitors

To study the role of endothelial nitric oxide synthase (eNOS) in cardiac function, we compared eNOS expression, contractility, and relaxation in the left ventricles of wild-type and eNOS-deficient mice. eNOS immunostaining is localized to the macro- and microvascular endothelium throughout the myocardium in wild-type mice and is absent in eNOS−/− mice. Whereas blood pressure is elevated in eNOS−/− mice, baseline cardiac contractility (dP/d t max) is similar in wild-type and eNOS−/− mice (9,673 ± 2,447 and 9,928 ± 1,566 mmHg/s, respectively). The β-adrenergic agonist isoproterenol (Iso) at doses of ≥1 ng causes enhanced increases in dP/d t max in eNOS−/− mice compared with wild-type controls in vivo ( P < 0.01) as well as in Langendorff isolated heart preparations ( P < 0.02). β-Adrenergic receptor binding (Bmax) is not significantly different in the two groups of animals (Bmax = 41.4 ± 9.4 and 36.1 ± 5.1 fmol/mg for wild-type and eNOS−/−). Iso-stimulated ventricular relaxation is also enhanced in the eNOS−/− mice, as measured by dP/d t min in the isolated heart. However, baseline ventricular relaxation is normal in eNOS−/− mice (τ = 5.2 ± 1.0 and 5.6 ± 1.5 ms for wild-type and eNOS−/−, respectively), whereas it is impaired in wild-type mice after NOS inhibition (τ = 8.3 ± 2.4 ms). cGMP levels in the left ventricle are unaffected by eNOS gene deletion (wild-type: 3.1 ± 0.8 pmol/mg, eNOS−/−: 3.1 ± 0.6 pmol/mg), leading us to examine the level of another physiological regulator of cGMP. Atrial natriuretic peptide (ANP) expression is markedly upregulated in the eNOS−/− mice, and exogenous ANP restores ventricular relaxation in wild-type mice treated with NOS inhibitors. These results suggest that eNOS attenuates both inotropic and lusitropic responses to β-adrenergic stimulation, and it also appears to regulate baseline ventricular relaxation in conjunction with ANP.

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