Journal of Molecular and Cellular Cardiology, Apr 1, 2021
Repolarization alternans, a periodic oscillation of long-short action potential duration, is an i... more Repolarization alternans, a periodic oscillation of long-short action potential duration, is an important source of arrhythmogenic substrate, although the mechanisms driving it are insufficiently understood. Despite its relevance as an arrhythmia precursor, there are no successful therapies able to target it specifically. We hypothesized that blockade of the sodium-calcium exchanger (NCX) could inhibit alternans. The effects of the selective NCX blocker ORM-10962 were evaluated on action potentials measured with microelectrodes from canine papillary muscle preparations, and calcium transients measured using Fluo4-AM from isolated ventricular myocytes paced to evoke alternans. Computer simulations were used to obtain insight into the drug's mechanisms of action. ORM-10962 attenuated cardiac alternans, both in action potential duration and calcium transient amplitude. Three morphological types of alternans were observed, with differential response to ORM-10962 with regards to APD alternans attenuation. Analysis of APD restitution indicates that calcium oscillations underlie alternans formation. Furthermore, ORM-10962 did not markedly alter APD restitution, but increased post-repolarization refractoriness, which may be mediated by indirectly reduced L-type calcium current. Computer simulations reproduced alternans attenuation via ORM-10962, suggesting that it is acts by reducing sarcoplasmic reticulum release refractoriness. This results from the ORM-10962-induced sodium-calcium exchanger block accompanied by an indirect reduction in L-type calcium current. Using a computer model of a heart failure cell, we furthermore demonstrate that the anti-alternans effect holds also for this disease, in which the risk of alternans is elevated. Targeting NCX may therefore be a useful anti-arrhythmic strategy to specifically prevent calcium driven alternans.
European Journal of Clinical Investigation, Oct 1, 2009
Background Acute heart failure is a potentially fatal manifestation of viral myocarditis. Develop... more Background Acute heart failure is a potentially fatal manifestation of viral myocarditis. Development of myocardial damage in myocarditis involves cardiomyocyte apoptosis. Levosimendan is a novel calcium sensitizing inotropic agent with anti-apoptotic properties. We studied the feasibility of inotropic treatment with levosimendan and its effects on apoptosis in experimental acute heart failure caused by coxsackievirus myocarditis. Materials and methods Adolescent BALB ⁄ c mice were infected with myocarditic Woodruff variant of coxsackievirus B3 (2 • 10 4 plaque-forming units). Mice were randomized into those receiving levosimendan 0AE33 mg kg)1 (total dose 1 mg kg)1 day)1) (n = 20) or vehicle (n = 19) given orally by gauge three times a day for 7 days after infection. Left ventricular function was evaluated by transthoracic echocardiography and the mice were euthanized on day 7. Histopathology, amount of virus in the heart (virus titration assay) and cardiomyocyte apoptosis (TUNEL assay) were studied. Uninfected untreated control mice were also studied. Results Infection resulted in histopathologically severe myocarditis and significant impairment of left ventricular function. Levosimendan treatment significantly improved ventricular function (fractional shortening 0AE32 ± 0AE04 vs. 0AE23 ± 0AE05, P = 0AE005; contractility 0AE60 ± 0AE12 vs. 0AE39 ± 0AE14, P = 0AE007 and myocardial performance index 0AE36 ± 0AE06 vs. 0AE62 ± 0AE15, P < 0AE0001) compared with vehicle. Levosimendan also reduced cardiomyocyte apoptosis (0AE26 ± 0AE08% vs. 0AE44 ± 0AE15% in vehicle, P = 0AE008), but did not have an effect on areas of myocardial necrosis or inflammation, or the amount of virus in the heart. Levosimendan treatment did not affect mortality (total mortality 63%). Conclusions Levosimendan improves ventricular function and inhibits cardiomyocyte apoptosis; therefore, it is suggested as a potentially feasible therapy in acute heart failure caused by viral myocarditis.
We investigated whether preconditioning with caloric restriction (CR) ameliorates kidney ischaemi... more We investigated whether preconditioning with caloric restriction (CR) ameliorates kidney ischaemia/reperfusion (I/R) injury and whether the salutary effects of CR are mediated through enhanced autophagy and/ or activation of key metabolic sensors SIRT1, AMP-kinase and PGC-1a. Methods: Six-to seven-week-old Wistar rats were divided into three groups: (i) sham-operated group; (ii) I/R group (40-min ischaemia followed by 24 h of reperfusion); and (iii) I/R group kept under CR (energy intake 70%) for 2 weeks before surgery. In additional experiments, sirtinol and 3-methyladenine (3-MA) were used as inhibitors of SIRT1 and autophagy respectively. Renal function was measured, and acute tubular damage and nitrotyrosine expression were scored. Kidney adenosine monophosphateactivated kinase (AMPK), SIRT1, eNOS, PGC-1a and LC-3B expressions were measured. Results: Caloric restriction improved renal function, protected against the development of acute tubular necrosis and attenuated I/R-induced nitrosative stress. Kidney I/R injury decreased eNOS and PGC-1a expression, inhibit autophagy and increased SIRT1 and AMPK expressions by 2.6-and fourfold respectively. However, phosphorylation level of AMPK was decreased. As compared with I/R injury group, CR further increased kidney SIRT1 expression by 1.8-fold, promoted autophagy and counteracted I/R-induced decreases in the expression of eNOS and PGC-1a. 3-MA abolished the renoprotective effects of CR, whereas sirtinol did not influence renal function in CR rats with I/R injury. Conclusions: Caloric restriction ameliorates acute kidney I/R injury through enhanced autophagy and counteraction of I/R-induced decreases in the renal expression of eNOS and PGC-1a.
Hypertension and persistent activation of the renin–angiotensin system (RAS) are predisposing fac... more Hypertension and persistent activation of the renin–angiotensin system (RAS) are predisposing factors for the development of acute kidney injury (AKI). Although bone-marrow-derived stromal cells (BMSCs) have shown therapeutic promise in treatment of AKI, the impact of pathological RAS on BMSC functionality has remained unresolved. RAS and its local components in the bone marrow are involved in several key steps of cell maturation processes. This may also render the BMSC population vulnerable to alterations even in the early phases of RAS pathology. We isolated transgenic BMSCs (TG-BMSCs) from young end-organ-disease-free rats with increased RAS activation [human angiotensinogen/renin double transgenic rats (dTGRs)] that eventually develop hypertension and die of end-organ damage and kidney failure at 8 weeks of age. Control cells (SD-BMSCs) were isolated from wild-type Sprague–Dawley rats. Cell phenotype, mitochondrial reactive oxygen species (ROS) production and respiration were assessed, and gene expression profiling was carried out using microarrays. Cells’ therapeutic efficacy was evaluated in a rat model of acute ischaemia/reperfusion-induced AKI. Serum urea and creatinine were measured at 24 h and 48 h. Acute tubular damage was scored and immunohistochemistry was used for evaluation for markers of inflammation [monocyte chemoattractant protein (MCP-1), ED-1], and kidney injury [kidney injury molecule-1 (KIM-1), neutrophil gelatinase-associated lipocalin (NGAL)]. TG-BMSCs showed distinct mitochondrial morphology, decreased cell respiration and increased production of ROS. Gene expression profiling revealed a pronounced pro-inflammatory phenotype. In contrast with the therapeutic effect of SD-BMSCs, administration of TG-BMSCs in the AKI model resulted in exacerbation of kidney injury and high mortality. Our results demonstrate that early persistent RAS activation can dramatically compromise therapeutic potential of BMSCs by causing a shift into a pro-inflammatory phenotype with mitochondrial dysfunction.
During ischaemia/reperfusion, the rise in [Na(+)](i), induced by simultaneous depression of the N... more During ischaemia/reperfusion, the rise in [Na(+)](i), induced by simultaneous depression of the Na(+)/K(+)-ATPase and activation of the Na(+)/H(+) exchanger (NHE), shifts the Na(+)/Ca(2+) exchanger (NCX) into reverse transport mode, resulting in Ca(2+)(i)overload, which is a critical factor in enhancing the liability to cardiac arrhythmias. The inhibition of NHE, and recently NCX has been suggested to effectively protect the heart from reperfusion-induced arrhythmias. In this study, we investigated and compared the efficacy of individual or the simultaneous inhibition of the NHE and NCX against reperfusion-induced arrhythmias in Langendorff-perfused rat hearts by applying a commonly used regional ischaemia-reperfusion protocol. The NHE and NCX were inhibited by cariporide and SEA0400 or the novel, more selective ORM-10103, respectively. Arrhythmia diagrams calculated for the reperfusion period were analysed for the incidence and duration of extrasystoles (ESs), ventricular tachycardia (VT) and ventricular fibrillation (VF). NHE inhibition by cariporide was highly efficient in reducing the recorded reperfusion-induced arrhythmias. Following the application of SEA0400 or ORM-10103, the number and duration of arrhythmic periods were efficiently or moderately decreased. While both NCX inhibitors effectively reduced ESs, the most frequently triggered arrhythmias, they exerted limited or no effect on VTs and VFs. Of the NCX inhibitors, ORM-10103 was more effective. Surprisingly, the simultaneous inhibition of the NCX and NHE failed to significantly improve the antiarrhythmic efficacy reached by NCX blockade alone. In conclusion, although principal simultaneous NHE+NCX inhibition should be highly effective against all types of the recorded reperfusion-induced arrhythmias, NCX inhibitors, alone or in combination with cariporide, seem to be moderately suitable to provide satisfactory cardioprotection - at least in the present arrhythmia model. Since ORM-10103 and SEA0400 are known to effectively inhibit after-depolarisations, it is suggested that their efficacy and that of other NCX inhibitors may be higher and more pronounced in the predominantly Ca(2+)(i)-dependent triggered arrhythmias.
Pharmacology Research & Perspectives, Apr 22, 2014
Kidney ischemia-reperfusion (I/R) injury is a common cause of acute kidney injury. We tested whet... more Kidney ischemia-reperfusion (I/R) injury is a common cause of acute kidney injury. We tested whether dexmedetomidine (Dex), an alpha2 adrenoceptor (a2-AR) agonist, protects against kidney I/R injury. Sprague-Dawley rats were divided into four groups: (1) Sham-operated group; (2) I/R group (40 min ischemia followed by 24 h reperfusion); (3) I/R group + Dex (1 lg/kg i.v. 60 min before the surgery), (4) I/R group + Dex (10 lg/kg). The effects of Dex postconditiong (Dex 1 or 10 lg/kg i.v. after reperfusion) as well as the effects of peripheral a2-AR agonism with fadolmidine were also examined. Hemodynamic effects were monitored, renal function measured, and acute tubular damage along with monocyte/macrophage infiltration scored. Kidney protein kinase B, toll like receptor 4, light chain 3B, p38 mitogen-activated protein kinase (p38 MAPK), sirtuin 1, adenosine monophosphate kinase (AMPK), and endothelial nitric oxide synthase (eNOS) expressions were measured, and kidney transciptome profiles analyzed. Dex preconditioning, but not postconditioning, attenuated I/R injury-induced renal dysfunction, acute tubular necrosis and inflammatory response. Neither pre-nor postconditioning with fadolmidine protected kidneys. Dex decreased blood pressure more than fadolmidine, ameliorated I/R-induced impairment of autophagy and increased renal p38 and eNOS expressions. Dex downregulated 245 and upregulated 61 genes representing 17 enriched Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways, in particular, integrin pathway and CD44. Ingenuity analysis revealed inhibition of Rac and nuclear factor (erythroid-derived 2)-like 2 pathways, whereas aryl hydrocarbon receptor (AHR) pathway was activated. Dex preconditioning ameliorates kidney I/R injury and inflammatory response, at least in part, through p38-CD44-pathway and possibly also through ischemic preconditioning.
Sinus node (SN) pacemaking is based on a coupling between surface membrane ion-channels and intra... more Sinus node (SN) pacemaking is based on a coupling between surface membrane ion-channels and intracellular Ca 2+-handling. The fundamental role of the inward Na + /Ca 2+ exchanger (NCX) is firmly established. However, little is known about the reverse mode exchange. A simulation study attributed important role to reverse NCX activity, however experimental evidence is still missing. Whole-cell and perforated patch-clamp experiments were performed on rabbit SN cells supplemented with fluorescent Ca 2+-tracking. We established 2 and 8 mM pipette NaCl groups to suppress and enable reverse NCX. NCX was assessed by specific block with 1 μM ORM-10962. Mechanistic simulations were performed by Maltsev-Lakatta minimal computational SN model. Active reverse NCX resulted in larger Ca 2+-transient amplitude with larger SR Ca 2+-content. Spontaneous action potential (AP) frequency increased with 8 mM NaCl. When reverse NCX was facilitated by 1 μM strophantin the Ca 2+ i and spontaneous rate increased. ORM-10962 applied prior to strophantin prevented Ca 2+ i and AP cycle change. Computational simulations indicated gradually increasing reverse NCX current, Ca 2+ i and heart rate with increasing Na + i. Our results provide further evidence for the role of reverse NCX in SN pacemaking. The reverse NCX activity may provide additional Ca 2+-influx that could increase SR Ca 2+content, which consequently leads to enhanced pacemaking activity. The SN AP is characterized by its slow diastolic depolarization (DD) phase starting from the maximal diastolic potential (MDP) and ending at the AP threshold of about − 40 mV. It was suggested that the decaying delayed rectifier potassium current (I X1 or later called I Kr) 1 and the cAMP-dependent hyperpolarization activated funnycurrent 2 has a major role forming the DD. This mechanism of pacemaking, driven by transmembrane ion channels with Hodgkin-Huxley kinetics was termed later as "membrane clock" (M clock). Further studies which identified rhythmic, spontaneous subsarcolemmal Ca 2+ releases (LCR) generated by the sarcoplasmic reticulum (SR) via ryanodine receptors during the DD ("Ca 2+ clock") have challenged the dominant role of membrane clock in the spontaneous automaticity of the SN 3-5. After intense debate regarding the underlying mechanism of spontaneous SN pacemaking, a large body of evidence proved that the M clock and Ca 2+ clock are functionally tightly coupled, so the two mechanisms form the latest concept of SN automaticity which is called the coupled clock system. This coupling is based on numerous time-, voltage-and Ca 2+-dependent mechanisms 6,7 including a major role of the L-type Ca 2+ current (I CaL) that "resets" and "refuels" the Ca 2+ clock 6. The crucial role of the (forward/inward) NCX in the SN pacemaking as part of the Ca 2+ clock now is firmly established, and represents a fundamental mechanism of spontaneous pacemaking, however little is known about the reverse NCX in SN. Several computational NCX models have not considered a significant reverse NCX in SN cells 8-13. To the best of our knowledge, the first reported model regarding SN cells which attributed a significant role to the reverse mode was proposed by Maltsev et al. in 2013, who claimed that Ca 2+ influx via reverse NCX could be functionally important since its contribution to refilling the sarcoplasmic reticulum is almost as large as the contribution of I CaL 14. This modelling prediction has important implications since it suggested yet unexplored and novel mechanisms contributing to the pacemaking system.
Calcium-sensitizing agents improve cardiac function in acute heart failure; however, their long-t... more Calcium-sensitizing agents improve cardiac function in acute heart failure; however, their long-term effects on cardiovascular mortality are unknown. We tested the hypothesis that levosimendan, an inodilator that acts through calcium sensitization, opening of ATP-dependent potassium channels and phosphodiesterase III inhibition, improves cardiac function and survival in double transgenic rats harboring human renin and angiotensinogen genes (dTGRs), a model of angiotensin II (Ang II)-induced hypertensive heart failure. Levosimendan (1 mg kg À1) was administered orally to 4-week-old dTGRs and normotensive Sprague-Dawley rats for 4 weeks. Untreated dTGRs developed severe hypertension, cardiac hypertrophy, heart failure with impaired diastolic relaxation, and exhibited a high mortality rate at the age of 8 weeks. Levosimendan did not decrease blood pressure and did not prevent cardiac hypertrophy. However, levosimendan improved systolic function, decreased cardiac atrial natriuretic peptide mRNA expression, ameliorated Ang II-induced cardiac damage and decreased mortality. Levosimendan did not correct Ang II-induced diastolic dysfunction and did not influence heart rate. In a separate survival study, levosimendan increased dTGR survival by 58% and median survival time by 27% (P¼0.004). Our findings suggest that levosimendan ameliorates Ang II-induced hypertensive heart failure and reduces mortality. The results also support the notion that the effects of levosimendan in dTGRs are mediated by blood pressure-independent mechanisms and include improved systolic function and amelioration of Ang II-induced coronary and cardiomyocyte damage.
Background and purpose: Levosimendan is a novel, short half-life calcium sensitizer used as pharm... more Background and purpose: Levosimendan is a novel, short half-life calcium sensitizer used as pharmacological inotropic support in acute decompensated heart failure. After oral administration, levosimendan is metabolized to OR-1855, which, in rats, is further metabolized into OR-1896. OR-1896 is a long-lasting metabolite of levosimendan sharing the pharmacological properties of the parent compound. Experimental approach: Effects of oral OR-1896 treatment on post-infarct heart failure and cardiac remodelling were assessed in diabetic Goto-Kakizaki (GK) rats, an animal model of type II diabetes. Myocardial infarction (MI) was produced to GK rats by coronary ligation. Twenty-four hours after MI or sham operation, the rats were randomized into four groups: (i) MI; (ii) MI + OR-1896 treatment; (iii) sham; and (iv) sham + OR-1896. Cardiac function and markers of cardiac remodelling were assessed 1, 4 and 12 weeks after MI. Key results: OR-1896 increased ejection fraction and fractional shortening in GK rats with MI. OR-1896 ameliorated postinfarct cardiac hypertrophy, and prevented the MI-induced increase in cardiac mRNA for atrial natriuretic peptide, monocyte chemoattractant protein-1 and connective tissue growth factor, markers of pressure/volume overload, inflammation and fibrosis respectively. OR-1896 also suppressed mRNA for senescence-associated p16 INK4A and p19 ARF. The beneficial effects of OR-1896 were more prominent at week 12 than at week 4. OR-1896 did not influence systolic blood pressure, blood glucose level, myocardial infarct size or cardiovascular mortality. Conclusions and implications: Oral treatment with calcium sensitizer OR-1896 protects against post-infarct heart failure and cardiac remodelling in experimental model of type II diabetes.
Experimental and Molecular Therapeutics, Aug 13, 2020
Background: Nausea and vomiting are common side effects of cancer therapies thereby seriously aff... more Background: Nausea and vomiting are common side effects of cancer therapies thereby seriously affecting the quality of life of patients. Therefore, the prevention or treatment of chemotherapy-induced nausea and vomiting has become an important part of the comprehensive treatment of cancer. Several 5-HT3 receptor antagonists have been developed in the treatment of nausea and vomiting using mainly the oral route of administration (which is not convenient due to vomiting) or the intravenous route. In this study we tested whether ocular administration of the 5-HT3 receptor antagonist palonosetron (eye drops) could prevent nausea and vomiting associated with intravenous cisplatin injection in dogs. Methods: Six adult male Beagle dogs (10-13 kg) were included in a randomized crossover study. This study was carried out in AAALAC facilities in strict accordance with the European and French animal welfare regulations for animal use in experimentation. Dogs were administered 18 mg/m2 cisplatin intravenously over a 20-min period, followed 45 min later by ocular administration (eye drops, 200µL/eye) of either placebo (2% povidone in saline) or palonosetron (30 or 120 µg/kg). The number of vomits and nausea associated behaviours, scored on a visual analogue scale (VAS), were recorded every 15 min for 7 h following cisplatin administration. Blood samples were collected to measure plasma levels of palonosetron. Results: The placebo treated group vomited an average number of 10.2±2.1 times (range 5-17) over the 7-h period following cisplatin infusion. Ocular administration of palonosetron at 30 μg/kg in cisplatin-treated dogs very strongly and significantly decreased vomiting episodes, with only 1 vomiting episode in 2 dogs overall (p&lt;0.0001). After ocular administration of palonosetron at 120 μg/kg, no vomiting episodes were observed in any dog throughout the observation period (p&lt;0.0001). With regards to the VAS, in the placebo-treated group the nausea-associated behaviour started between 2.5 and 2.75 h after the end of the cisplatin infusion and peaked between 3.75 and 4.25 h after the end of the cisplatin infusion. The area under the curve (AUC) of VAS was 7344 ± 1050 mm*min in the placebo-treated group. Ocular administration of palonosetron at 30 or 120 µg/kg in cisplatin-treated dogs decreased the VAS score strongly. The VAS AUCs were 320 ± 268 (p&lt;0.0001) and 0 ± 0 mm*m (p&lt;0.0001) in 30 µg/kg and 120 µg/kg palonosetron groups, respectively. Plasma palonosetron concentration reached 10.3 ± 3.9 ng/mL, 5 min after ocular administration of palonosetron at 30 µg/kg and 2.8 ± 0.5 ng/mL and 0.9 ± 0.2 ng/mL at 2 h and 6 h post dosing, respectively. Conclusions: This study demonstrates for the first time that palonosetron administered via ocular route produces a rapid clear-cut anti-emetic and anti-nausea effect in conscious dogs exposed to cisplatin. This finding points out the tremendous interest of this novel and convenient route of administration which could help improving the quality of life of patients. Citation Format: Sandra Nourry, Minja Hyttila-Hopponen, Pierre Montagne, Lasse Saloranta, Sari Pappinen, Jouko Levijoki, Christophe Drieu La Rochelle. Ocular drops of palonosetron reduce chemotherapy-induced acute nausea and vomiting in the dog [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 593.
Background: Diabetes is associated with changes in myocardial stress-response pathways and is rec... more Background: Diabetes is associated with changes in myocardial stress-response pathways and is recognized as an independent risk factor for cardiac remodeling. Using spontaneously diabetic Goto Kakizaki rats as a model of type 2 DM we investigated whether post-translational modifications in the Akt-FOXO3a pathway, Sirt1-p53 pathway and the mitogen activated protein kinase p38 regulator are involved in post-infarct cardiac remodeling Methods: Experimental myocardial infarction (MI) was induced by left anterior descending coronary artery ligation in spontaneously diabetic Goto-Kakizaki rats and non-diabetic Wistar controls. Cardiac function was studied by echocardiography. Myocardial hypertrophy, cardiomyocyte apoptosis and cardiac fibrosis were determined histologically 12 weeks post MI or Sham operation. Western blotting was used to study Caspase-3, Bax, Sirt1, acetylation of p53 and phosphorylation of p38, Akt and FOXO3a. Electrophoretic mobility shift assay was used to assess FOXO3a activity and its nuclear localization. Results: Post-infarct heart failure in diabetic GK rats was associated with pronounced cardiomyocyte hypertrophy, increased interstitial fibrosis and sustained cardiomyocyte apoptosis as compared with their non-diabetic Wistar controls. In the GK rat myocardium, Akt-and FOXO3a-phosphorylation was decreased and nuclear localization of FOXO3a was increased concomitantly with increased PTEN protein expression. Furthermore, increased Sirt1 protein expression was associated with decreased p53 acetylation, and phosphorylation of p38 was increased in diabetic rats with MI. Conclusions: Post-infarct heart failure in diabetic GK rats was associated with more pronounced cardiac hypertrophy, interstitial fibrosis and sustained cardiomyocyte apoptosis as compared to their non-diabetic controls. The present study suggests important roles for Akt-FOXO3a, Sirt1-p53 and p38 MAPK in the regulation of postinfarct cardiac remodeling in type 2 diabetes.
Journal of Pharmacology and Experimental Therapeutics
Cancer treatments are frequently associated with nausea and vomiting despite greatly improved pre... more Cancer treatments are frequently associated with nausea and vomiting despite greatly improved preventive medication. Administration of anti-nausea agents as eye drops might provide an easy and rapid access to the systemic circulation for prevention of nausea and vomiting and for the treatment of breakthrough nausea, but the ocular administration route has rarely been evaluated. Palonosetron is a second-generation 5-HT 3 receptor antagonist approved for prevention and treatment of chemotherapy-induced nausea and vomiting. We compared ocular administration of palonosetron to non-active vehicle eye drops and to intravenous palonosetron in the prevention of cisplatin-induced nausea and vomiting in beagle dogs. Palonosetron ocular drops at the dose of 30 µg/kg reduced cumulative nausea over time as measured with the area under the visual analog scale curve (VAS-AUC) by 98% compared with the vehicle, and reduced nausea-associated dog behavior by 95%. Vomiting was completely prevented with repeated palonosetron ocular dosing. Hydroxypropyl-βcyclodextrin (HP-β-CD) palonosetron formulation was well tolerated locally at the palonosetron concentration of 3 mg/mL. Absorption of palonosetron from eye drops was fast. Ten minutes after ocular administration, palonosetron plasma concentrations were similar compared to intravenous administration, and remained similar for six hours. We conclude that palonosetron is rapidly absorbed into the systemic circulation from eye drops. Ocularly administered palonosetron was well tolerated in the HP-β-CD formulation and was highly effective in the prevention of cisplatin-induced nausea and vomiting. Evaluation of the safety and efficacy of ocular administration of palonosetron is warranted in the prevention and treatment of chemotherapy-induced nausea and vomiting in clinical trials.
Sinus node (SN) pacemaking is based on a coupling between surface membrane ion-channels and intra... more Sinus node (SN) pacemaking is based on a coupling between surface membrane ion-channels and intracellular Ca2+-handling. The fundamental role of the inward Na+/Ca2+ exchanger (NCX) is firmly established. However, little is known about the reverse mode exchange. A simulation study attributed important role to reverse NCX activity, however experimental evidence is still missing. Whole-cell and perforated patch-clamp experiments were performed on rabbit SN cells supplemented with fluorescent Ca2+-tracking. We established 2 and 8 mM pipette NaCl groups to suppress and enable reverse NCX. NCX was assessed by specific block with 1 μM ORM-10962. Mechanistic simulations were performed by Maltsev–Lakatta minimal computational SN model. Active reverse NCX resulted in larger Ca2+-transient amplitude with larger SR Ca2+-content. Spontaneous action potential (AP) frequency increased with 8 mM NaCl. When reverse NCX was facilitated by 1 μM strophantin the Ca2+i and spontaneous rate increased. ORM...
: OR-1855 and OR-1896 are 2 hemodynamically active metabolites of the inodilator levosimendan, wi... more : OR-1855 and OR-1896 are 2 hemodynamically active metabolites of the inodilator levosimendan, with calcium sensitizing activity, but their mechanism of action is still not fully understood. It has been previously reported that the positive inotropic effect of levosimendan is not potentiated by the adenylate cyclase activator forskolin, whereas forskolin does potentiate the effects of the phosphodiesterase (PDE) inhibitor milrinone. To ascertain whether the active metabolites follow the same pattern of levosimendan, the positive inotropic effects of OR- 1855 and OR-1896 were studied in guinea-pig-isolated papillary muscle in the presence and absence of forskolin. OR-1855 and OR-1896 were also tested as inhibitors of PDE-III and PDE-IV. Our results show that 0.1 µM forskolin did not potentiate the positive inotropic effect of OR-1855 or OR-1896, as in the case of the parent compound levosimendan. As in previous studies, the positive inotropic effect of milrinone was markedly potentiated in the presence of forskolin. From these data, we propose an explanation for the divergent behavior of the calcium sensitizing drugs and PDE inhibitors.
Journal of Molecular and Cellular Cardiology, 2021
Repolarization alternans, a periodic oscillation of long-short action potential duration, is an i... more Repolarization alternans, a periodic oscillation of long-short action potential duration, is an important source of arrhythmogenic substrate, although the mechanisms driving it are insufficiently understood. Despite its relevance as an arrhythmia precursor, there are no successful therapies able to target it specifically. We hypothesized that blockade of the sodium-calcium exchanger (NCX) could inhibit alternans. The effects of the selective NCX blocker ORM-10962 were evaluated on action potentials measured with microelectrodes from canine papillary muscle preparations, and calcium transients measured using Fluo4-AM from isolated ventricular myocytes paced to evoke alternans. Computer simulations were used to obtain insight into the drug's mechanisms of action. ORM-10962 attenuated cardiac alternans, both in action potential duration and calcium transient amplitude. Three morphological types of alternans were observed, with differential response to ORM-10962 with regards to APD alternans attenuation. Analysis of APD restitution indicates that calcium oscillations underlie alternans formation. Furthermore, ORM-10962 did not markedly alter APD restitution, but increased post-repolarization refractoriness, which may be mediated by indirectly reduced L-type calcium current. Computer simulations reproduced alternans attenuation via ORM-10962, suggesting that it is acts by reducing sarcoplasmic reticulum release refractoriness. This results from the ORM-10962-induced sodium-calcium exchanger block accompanied by an indirect reduction in L-type calcium current. Using a computer model of a heart failure cell, we furthermore demonstrate that the anti-alternans effect holds also for this disease, in which the risk of alternans is elevated. Targeting NCX may therefore be a useful anti-arrhythmic strategy to specifically prevent calcium driven alternans.
Kidney ischemia-reperfusion (I/R) injury is a common cause of acute kidney injury. We tested whet... more Kidney ischemia-reperfusion (I/R) injury is a common cause of acute kidney injury. We tested whether dexmedetomidine (Dex), an alpha2 adrenoceptor (a2-AR) agonist, protects against kidney I/R injury. Sprague–Dawley rats were divided into four groups: (1) Sham-operated group; (2) I/R group (40 min ischemia followed by 24 h reperfusion); (3) I/R group + Dex (1 lg/kg i.v. 60 min before the surgery), (4) I/R group + Dex (10 lg/kg). The effects of Dex postconditiong (Dex 1 or 10 lg/kg i.v. after reperfusion) as well as the effects of peripheral a2-AR agonism with fadolmidine were also examined. Hemody-namic effects were monitored, renal function measured, and acute tubular dam-age along with monocyte/macrophage infiltration scored. Kidney protein kinase B, toll like receptor 4, light chain 3B, p38 mitogen-activated protein kinase (p38 MAPK), sirtuin 1, adenosine monophosphate kinase (AMPK), and endo-thelial nitric oxide synthase (eNOS) expressions were measured, and kidney transciptome p...
Background and PurposeThe lack of selective sodium–calcium exchanger (NCX) inhibitors has hampere... more Background and PurposeThe lack of selective sodium–calcium exchanger (NCX) inhibitors has hampered the exploration of physiological and pathophysiological roles of cardiac NCX 1.1. We aimed to discover more potent and selective drug like NCX 1.1 inhibitor.Experimental ApproachA flavan series‐based pharmacophore model was constructed. Virtual screening helped us identify a novel scaffold for NCX inhibition. A distinctively different NCX 1.1 inhibitor, ORM‐11372, was discovered after lead optimization. Its potency against human and rat NCX 1.1 and selectivity against other ion channels was assessed. The cardiovascular effects of ORM‐11372 were studied in normal and infarcted rats and rabbits. Human cardiac safety was studied ex vivo using human ventricular trabeculae.Key ResultsORM‐11372 inhibited human NCX 1.1 reverse and forward currents; IC50 values were 5 and 6 nM respectively. ORM‐11372 inhibited human cardiac sodium 1.5 (INa) and hERG KV11.1 currents (IhERG) in a concentration‐d...
Journal of physiology and pharmacology : an official journal of the Polish Physiological Society, 2009
Calcium-sensitizing agents have been shown to improve cardiac function in patients suffering from... more Calcium-sensitizing agents have been shown to improve cardiac function in patients suffering from acute decompensated heart failure, however, their long-term effects on cardiac remodeling and cardiovascular mortality are still largely unknown. In the present study we tested the hypothesis whether OR-1896, an active and long-lasting metabolite of calcium sensitizer levosimendan, prevents cardiovascular mortality and hypertension-induced myocardial remodelling in salt-sensitive Dahl/Rapp rats. OR-1896 was given orally to Dahl/Rapp SS rats on high-salt diet (NaCl 7% w/w) for 7 weeks at two different doses (0.5 and 0.05 mg/kg). OR-1896 prevented salt-induced cardiovascular mortality (survival rate 75 % in OR-1896 treated groups vs 38 % in untreated controls, p<0.01), ameliorated cardiac hypertrophy and improved systolic functions of the heart without major influence on systemic blood pressure. OR-1896 also ameliorated salt-induced increase in cardiac ANP mRNA expression and plasma BN...
Journal of Molecular and Cellular Cardiology, Apr 1, 2021
Repolarization alternans, a periodic oscillation of long-short action potential duration, is an i... more Repolarization alternans, a periodic oscillation of long-short action potential duration, is an important source of arrhythmogenic substrate, although the mechanisms driving it are insufficiently understood. Despite its relevance as an arrhythmia precursor, there are no successful therapies able to target it specifically. We hypothesized that blockade of the sodium-calcium exchanger (NCX) could inhibit alternans. The effects of the selective NCX blocker ORM-10962 were evaluated on action potentials measured with microelectrodes from canine papillary muscle preparations, and calcium transients measured using Fluo4-AM from isolated ventricular myocytes paced to evoke alternans. Computer simulations were used to obtain insight into the drug's mechanisms of action. ORM-10962 attenuated cardiac alternans, both in action potential duration and calcium transient amplitude. Three morphological types of alternans were observed, with differential response to ORM-10962 with regards to APD alternans attenuation. Analysis of APD restitution indicates that calcium oscillations underlie alternans formation. Furthermore, ORM-10962 did not markedly alter APD restitution, but increased post-repolarization refractoriness, which may be mediated by indirectly reduced L-type calcium current. Computer simulations reproduced alternans attenuation via ORM-10962, suggesting that it is acts by reducing sarcoplasmic reticulum release refractoriness. This results from the ORM-10962-induced sodium-calcium exchanger block accompanied by an indirect reduction in L-type calcium current. Using a computer model of a heart failure cell, we furthermore demonstrate that the anti-alternans effect holds also for this disease, in which the risk of alternans is elevated. Targeting NCX may therefore be a useful anti-arrhythmic strategy to specifically prevent calcium driven alternans.
European Journal of Clinical Investigation, Oct 1, 2009
Background Acute heart failure is a potentially fatal manifestation of viral myocarditis. Develop... more Background Acute heart failure is a potentially fatal manifestation of viral myocarditis. Development of myocardial damage in myocarditis involves cardiomyocyte apoptosis. Levosimendan is a novel calcium sensitizing inotropic agent with anti-apoptotic properties. We studied the feasibility of inotropic treatment with levosimendan and its effects on apoptosis in experimental acute heart failure caused by coxsackievirus myocarditis. Materials and methods Adolescent BALB ⁄ c mice were infected with myocarditic Woodruff variant of coxsackievirus B3 (2 • 10 4 plaque-forming units). Mice were randomized into those receiving levosimendan 0AE33 mg kg)1 (total dose 1 mg kg)1 day)1) (n = 20) or vehicle (n = 19) given orally by gauge three times a day for 7 days after infection. Left ventricular function was evaluated by transthoracic echocardiography and the mice were euthanized on day 7. Histopathology, amount of virus in the heart (virus titration assay) and cardiomyocyte apoptosis (TUNEL assay) were studied. Uninfected untreated control mice were also studied. Results Infection resulted in histopathologically severe myocarditis and significant impairment of left ventricular function. Levosimendan treatment significantly improved ventricular function (fractional shortening 0AE32 ± 0AE04 vs. 0AE23 ± 0AE05, P = 0AE005; contractility 0AE60 ± 0AE12 vs. 0AE39 ± 0AE14, P = 0AE007 and myocardial performance index 0AE36 ± 0AE06 vs. 0AE62 ± 0AE15, P < 0AE0001) compared with vehicle. Levosimendan also reduced cardiomyocyte apoptosis (0AE26 ± 0AE08% vs. 0AE44 ± 0AE15% in vehicle, P = 0AE008), but did not have an effect on areas of myocardial necrosis or inflammation, or the amount of virus in the heart. Levosimendan treatment did not affect mortality (total mortality 63%). Conclusions Levosimendan improves ventricular function and inhibits cardiomyocyte apoptosis; therefore, it is suggested as a potentially feasible therapy in acute heart failure caused by viral myocarditis.
We investigated whether preconditioning with caloric restriction (CR) ameliorates kidney ischaemi... more We investigated whether preconditioning with caloric restriction (CR) ameliorates kidney ischaemia/reperfusion (I/R) injury and whether the salutary effects of CR are mediated through enhanced autophagy and/ or activation of key metabolic sensors SIRT1, AMP-kinase and PGC-1a. Methods: Six-to seven-week-old Wistar rats were divided into three groups: (i) sham-operated group; (ii) I/R group (40-min ischaemia followed by 24 h of reperfusion); and (iii) I/R group kept under CR (energy intake 70%) for 2 weeks before surgery. In additional experiments, sirtinol and 3-methyladenine (3-MA) were used as inhibitors of SIRT1 and autophagy respectively. Renal function was measured, and acute tubular damage and nitrotyrosine expression were scored. Kidney adenosine monophosphateactivated kinase (AMPK), SIRT1, eNOS, PGC-1a and LC-3B expressions were measured. Results: Caloric restriction improved renal function, protected against the development of acute tubular necrosis and attenuated I/R-induced nitrosative stress. Kidney I/R injury decreased eNOS and PGC-1a expression, inhibit autophagy and increased SIRT1 and AMPK expressions by 2.6-and fourfold respectively. However, phosphorylation level of AMPK was decreased. As compared with I/R injury group, CR further increased kidney SIRT1 expression by 1.8-fold, promoted autophagy and counteracted I/R-induced decreases in the expression of eNOS and PGC-1a. 3-MA abolished the renoprotective effects of CR, whereas sirtinol did not influence renal function in CR rats with I/R injury. Conclusions: Caloric restriction ameliorates acute kidney I/R injury through enhanced autophagy and counteraction of I/R-induced decreases in the renal expression of eNOS and PGC-1a.
Hypertension and persistent activation of the renin–angiotensin system (RAS) are predisposing fac... more Hypertension and persistent activation of the renin–angiotensin system (RAS) are predisposing factors for the development of acute kidney injury (AKI). Although bone-marrow-derived stromal cells (BMSCs) have shown therapeutic promise in treatment of AKI, the impact of pathological RAS on BMSC functionality has remained unresolved. RAS and its local components in the bone marrow are involved in several key steps of cell maturation processes. This may also render the BMSC population vulnerable to alterations even in the early phases of RAS pathology. We isolated transgenic BMSCs (TG-BMSCs) from young end-organ-disease-free rats with increased RAS activation [human angiotensinogen/renin double transgenic rats (dTGRs)] that eventually develop hypertension and die of end-organ damage and kidney failure at 8 weeks of age. Control cells (SD-BMSCs) were isolated from wild-type Sprague–Dawley rats. Cell phenotype, mitochondrial reactive oxygen species (ROS) production and respiration were assessed, and gene expression profiling was carried out using microarrays. Cells’ therapeutic efficacy was evaluated in a rat model of acute ischaemia/reperfusion-induced AKI. Serum urea and creatinine were measured at 24 h and 48 h. Acute tubular damage was scored and immunohistochemistry was used for evaluation for markers of inflammation [monocyte chemoattractant protein (MCP-1), ED-1], and kidney injury [kidney injury molecule-1 (KIM-1), neutrophil gelatinase-associated lipocalin (NGAL)]. TG-BMSCs showed distinct mitochondrial morphology, decreased cell respiration and increased production of ROS. Gene expression profiling revealed a pronounced pro-inflammatory phenotype. In contrast with the therapeutic effect of SD-BMSCs, administration of TG-BMSCs in the AKI model resulted in exacerbation of kidney injury and high mortality. Our results demonstrate that early persistent RAS activation can dramatically compromise therapeutic potential of BMSCs by causing a shift into a pro-inflammatory phenotype with mitochondrial dysfunction.
During ischaemia/reperfusion, the rise in [Na(+)](i), induced by simultaneous depression of the N... more During ischaemia/reperfusion, the rise in [Na(+)](i), induced by simultaneous depression of the Na(+)/K(+)-ATPase and activation of the Na(+)/H(+) exchanger (NHE), shifts the Na(+)/Ca(2+) exchanger (NCX) into reverse transport mode, resulting in Ca(2+)(i)overload, which is a critical factor in enhancing the liability to cardiac arrhythmias. The inhibition of NHE, and recently NCX has been suggested to effectively protect the heart from reperfusion-induced arrhythmias. In this study, we investigated and compared the efficacy of individual or the simultaneous inhibition of the NHE and NCX against reperfusion-induced arrhythmias in Langendorff-perfused rat hearts by applying a commonly used regional ischaemia-reperfusion protocol. The NHE and NCX were inhibited by cariporide and SEA0400 or the novel, more selective ORM-10103, respectively. Arrhythmia diagrams calculated for the reperfusion period were analysed for the incidence and duration of extrasystoles (ESs), ventricular tachycardia (VT) and ventricular fibrillation (VF). NHE inhibition by cariporide was highly efficient in reducing the recorded reperfusion-induced arrhythmias. Following the application of SEA0400 or ORM-10103, the number and duration of arrhythmic periods were efficiently or moderately decreased. While both NCX inhibitors effectively reduced ESs, the most frequently triggered arrhythmias, they exerted limited or no effect on VTs and VFs. Of the NCX inhibitors, ORM-10103 was more effective. Surprisingly, the simultaneous inhibition of the NCX and NHE failed to significantly improve the antiarrhythmic efficacy reached by NCX blockade alone. In conclusion, although principal simultaneous NHE+NCX inhibition should be highly effective against all types of the recorded reperfusion-induced arrhythmias, NCX inhibitors, alone or in combination with cariporide, seem to be moderately suitable to provide satisfactory cardioprotection - at least in the present arrhythmia model. Since ORM-10103 and SEA0400 are known to effectively inhibit after-depolarisations, it is suggested that their efficacy and that of other NCX inhibitors may be higher and more pronounced in the predominantly Ca(2+)(i)-dependent triggered arrhythmias.
Pharmacology Research & Perspectives, Apr 22, 2014
Kidney ischemia-reperfusion (I/R) injury is a common cause of acute kidney injury. We tested whet... more Kidney ischemia-reperfusion (I/R) injury is a common cause of acute kidney injury. We tested whether dexmedetomidine (Dex), an alpha2 adrenoceptor (a2-AR) agonist, protects against kidney I/R injury. Sprague-Dawley rats were divided into four groups: (1) Sham-operated group; (2) I/R group (40 min ischemia followed by 24 h reperfusion); (3) I/R group + Dex (1 lg/kg i.v. 60 min before the surgery), (4) I/R group + Dex (10 lg/kg). The effects of Dex postconditiong (Dex 1 or 10 lg/kg i.v. after reperfusion) as well as the effects of peripheral a2-AR agonism with fadolmidine were also examined. Hemodynamic effects were monitored, renal function measured, and acute tubular damage along with monocyte/macrophage infiltration scored. Kidney protein kinase B, toll like receptor 4, light chain 3B, p38 mitogen-activated protein kinase (p38 MAPK), sirtuin 1, adenosine monophosphate kinase (AMPK), and endothelial nitric oxide synthase (eNOS) expressions were measured, and kidney transciptome profiles analyzed. Dex preconditioning, but not postconditioning, attenuated I/R injury-induced renal dysfunction, acute tubular necrosis and inflammatory response. Neither pre-nor postconditioning with fadolmidine protected kidneys. Dex decreased blood pressure more than fadolmidine, ameliorated I/R-induced impairment of autophagy and increased renal p38 and eNOS expressions. Dex downregulated 245 and upregulated 61 genes representing 17 enriched Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways, in particular, integrin pathway and CD44. Ingenuity analysis revealed inhibition of Rac and nuclear factor (erythroid-derived 2)-like 2 pathways, whereas aryl hydrocarbon receptor (AHR) pathway was activated. Dex preconditioning ameliorates kidney I/R injury and inflammatory response, at least in part, through p38-CD44-pathway and possibly also through ischemic preconditioning.
Sinus node (SN) pacemaking is based on a coupling between surface membrane ion-channels and intra... more Sinus node (SN) pacemaking is based on a coupling between surface membrane ion-channels and intracellular Ca 2+-handling. The fundamental role of the inward Na + /Ca 2+ exchanger (NCX) is firmly established. However, little is known about the reverse mode exchange. A simulation study attributed important role to reverse NCX activity, however experimental evidence is still missing. Whole-cell and perforated patch-clamp experiments were performed on rabbit SN cells supplemented with fluorescent Ca 2+-tracking. We established 2 and 8 mM pipette NaCl groups to suppress and enable reverse NCX. NCX was assessed by specific block with 1 μM ORM-10962. Mechanistic simulations were performed by Maltsev-Lakatta minimal computational SN model. Active reverse NCX resulted in larger Ca 2+-transient amplitude with larger SR Ca 2+-content. Spontaneous action potential (AP) frequency increased with 8 mM NaCl. When reverse NCX was facilitated by 1 μM strophantin the Ca 2+ i and spontaneous rate increased. ORM-10962 applied prior to strophantin prevented Ca 2+ i and AP cycle change. Computational simulations indicated gradually increasing reverse NCX current, Ca 2+ i and heart rate with increasing Na + i. Our results provide further evidence for the role of reverse NCX in SN pacemaking. The reverse NCX activity may provide additional Ca 2+-influx that could increase SR Ca 2+content, which consequently leads to enhanced pacemaking activity. The SN AP is characterized by its slow diastolic depolarization (DD) phase starting from the maximal diastolic potential (MDP) and ending at the AP threshold of about − 40 mV. It was suggested that the decaying delayed rectifier potassium current (I X1 or later called I Kr) 1 and the cAMP-dependent hyperpolarization activated funnycurrent 2 has a major role forming the DD. This mechanism of pacemaking, driven by transmembrane ion channels with Hodgkin-Huxley kinetics was termed later as "membrane clock" (M clock). Further studies which identified rhythmic, spontaneous subsarcolemmal Ca 2+ releases (LCR) generated by the sarcoplasmic reticulum (SR) via ryanodine receptors during the DD ("Ca 2+ clock") have challenged the dominant role of membrane clock in the spontaneous automaticity of the SN 3-5. After intense debate regarding the underlying mechanism of spontaneous SN pacemaking, a large body of evidence proved that the M clock and Ca 2+ clock are functionally tightly coupled, so the two mechanisms form the latest concept of SN automaticity which is called the coupled clock system. This coupling is based on numerous time-, voltage-and Ca 2+-dependent mechanisms 6,7 including a major role of the L-type Ca 2+ current (I CaL) that "resets" and "refuels" the Ca 2+ clock 6. The crucial role of the (forward/inward) NCX in the SN pacemaking as part of the Ca 2+ clock now is firmly established, and represents a fundamental mechanism of spontaneous pacemaking, however little is known about the reverse NCX in SN. Several computational NCX models have not considered a significant reverse NCX in SN cells 8-13. To the best of our knowledge, the first reported model regarding SN cells which attributed a significant role to the reverse mode was proposed by Maltsev et al. in 2013, who claimed that Ca 2+ influx via reverse NCX could be functionally important since its contribution to refilling the sarcoplasmic reticulum is almost as large as the contribution of I CaL 14. This modelling prediction has important implications since it suggested yet unexplored and novel mechanisms contributing to the pacemaking system.
Calcium-sensitizing agents improve cardiac function in acute heart failure; however, their long-t... more Calcium-sensitizing agents improve cardiac function in acute heart failure; however, their long-term effects on cardiovascular mortality are unknown. We tested the hypothesis that levosimendan, an inodilator that acts through calcium sensitization, opening of ATP-dependent potassium channels and phosphodiesterase III inhibition, improves cardiac function and survival in double transgenic rats harboring human renin and angiotensinogen genes (dTGRs), a model of angiotensin II (Ang II)-induced hypertensive heart failure. Levosimendan (1 mg kg À1) was administered orally to 4-week-old dTGRs and normotensive Sprague-Dawley rats for 4 weeks. Untreated dTGRs developed severe hypertension, cardiac hypertrophy, heart failure with impaired diastolic relaxation, and exhibited a high mortality rate at the age of 8 weeks. Levosimendan did not decrease blood pressure and did not prevent cardiac hypertrophy. However, levosimendan improved systolic function, decreased cardiac atrial natriuretic peptide mRNA expression, ameliorated Ang II-induced cardiac damage and decreased mortality. Levosimendan did not correct Ang II-induced diastolic dysfunction and did not influence heart rate. In a separate survival study, levosimendan increased dTGR survival by 58% and median survival time by 27% (P¼0.004). Our findings suggest that levosimendan ameliorates Ang II-induced hypertensive heart failure and reduces mortality. The results also support the notion that the effects of levosimendan in dTGRs are mediated by blood pressure-independent mechanisms and include improved systolic function and amelioration of Ang II-induced coronary and cardiomyocyte damage.
Background and purpose: Levosimendan is a novel, short half-life calcium sensitizer used as pharm... more Background and purpose: Levosimendan is a novel, short half-life calcium sensitizer used as pharmacological inotropic support in acute decompensated heart failure. After oral administration, levosimendan is metabolized to OR-1855, which, in rats, is further metabolized into OR-1896. OR-1896 is a long-lasting metabolite of levosimendan sharing the pharmacological properties of the parent compound. Experimental approach: Effects of oral OR-1896 treatment on post-infarct heart failure and cardiac remodelling were assessed in diabetic Goto-Kakizaki (GK) rats, an animal model of type II diabetes. Myocardial infarction (MI) was produced to GK rats by coronary ligation. Twenty-four hours after MI or sham operation, the rats were randomized into four groups: (i) MI; (ii) MI + OR-1896 treatment; (iii) sham; and (iv) sham + OR-1896. Cardiac function and markers of cardiac remodelling were assessed 1, 4 and 12 weeks after MI. Key results: OR-1896 increased ejection fraction and fractional shortening in GK rats with MI. OR-1896 ameliorated postinfarct cardiac hypertrophy, and prevented the MI-induced increase in cardiac mRNA for atrial natriuretic peptide, monocyte chemoattractant protein-1 and connective tissue growth factor, markers of pressure/volume overload, inflammation and fibrosis respectively. OR-1896 also suppressed mRNA for senescence-associated p16 INK4A and p19 ARF. The beneficial effects of OR-1896 were more prominent at week 12 than at week 4. OR-1896 did not influence systolic blood pressure, blood glucose level, myocardial infarct size or cardiovascular mortality. Conclusions and implications: Oral treatment with calcium sensitizer OR-1896 protects against post-infarct heart failure and cardiac remodelling in experimental model of type II diabetes.
Experimental and Molecular Therapeutics, Aug 13, 2020
Background: Nausea and vomiting are common side effects of cancer therapies thereby seriously aff... more Background: Nausea and vomiting are common side effects of cancer therapies thereby seriously affecting the quality of life of patients. Therefore, the prevention or treatment of chemotherapy-induced nausea and vomiting has become an important part of the comprehensive treatment of cancer. Several 5-HT3 receptor antagonists have been developed in the treatment of nausea and vomiting using mainly the oral route of administration (which is not convenient due to vomiting) or the intravenous route. In this study we tested whether ocular administration of the 5-HT3 receptor antagonist palonosetron (eye drops) could prevent nausea and vomiting associated with intravenous cisplatin injection in dogs. Methods: Six adult male Beagle dogs (10-13 kg) were included in a randomized crossover study. This study was carried out in AAALAC facilities in strict accordance with the European and French animal welfare regulations for animal use in experimentation. Dogs were administered 18 mg/m2 cisplatin intravenously over a 20-min period, followed 45 min later by ocular administration (eye drops, 200µL/eye) of either placebo (2% povidone in saline) or palonosetron (30 or 120 µg/kg). The number of vomits and nausea associated behaviours, scored on a visual analogue scale (VAS), were recorded every 15 min for 7 h following cisplatin administration. Blood samples were collected to measure plasma levels of palonosetron. Results: The placebo treated group vomited an average number of 10.2±2.1 times (range 5-17) over the 7-h period following cisplatin infusion. Ocular administration of palonosetron at 30 μg/kg in cisplatin-treated dogs very strongly and significantly decreased vomiting episodes, with only 1 vomiting episode in 2 dogs overall (p&lt;0.0001). After ocular administration of palonosetron at 120 μg/kg, no vomiting episodes were observed in any dog throughout the observation period (p&lt;0.0001). With regards to the VAS, in the placebo-treated group the nausea-associated behaviour started between 2.5 and 2.75 h after the end of the cisplatin infusion and peaked between 3.75 and 4.25 h after the end of the cisplatin infusion. The area under the curve (AUC) of VAS was 7344 ± 1050 mm*min in the placebo-treated group. Ocular administration of palonosetron at 30 or 120 µg/kg in cisplatin-treated dogs decreased the VAS score strongly. The VAS AUCs were 320 ± 268 (p&lt;0.0001) and 0 ± 0 mm*m (p&lt;0.0001) in 30 µg/kg and 120 µg/kg palonosetron groups, respectively. Plasma palonosetron concentration reached 10.3 ± 3.9 ng/mL, 5 min after ocular administration of palonosetron at 30 µg/kg and 2.8 ± 0.5 ng/mL and 0.9 ± 0.2 ng/mL at 2 h and 6 h post dosing, respectively. Conclusions: This study demonstrates for the first time that palonosetron administered via ocular route produces a rapid clear-cut anti-emetic and anti-nausea effect in conscious dogs exposed to cisplatin. This finding points out the tremendous interest of this novel and convenient route of administration which could help improving the quality of life of patients. Citation Format: Sandra Nourry, Minja Hyttila-Hopponen, Pierre Montagne, Lasse Saloranta, Sari Pappinen, Jouko Levijoki, Christophe Drieu La Rochelle. Ocular drops of palonosetron reduce chemotherapy-induced acute nausea and vomiting in the dog [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 593.
Background: Diabetes is associated with changes in myocardial stress-response pathways and is rec... more Background: Diabetes is associated with changes in myocardial stress-response pathways and is recognized as an independent risk factor for cardiac remodeling. Using spontaneously diabetic Goto Kakizaki rats as a model of type 2 DM we investigated whether post-translational modifications in the Akt-FOXO3a pathway, Sirt1-p53 pathway and the mitogen activated protein kinase p38 regulator are involved in post-infarct cardiac remodeling Methods: Experimental myocardial infarction (MI) was induced by left anterior descending coronary artery ligation in spontaneously diabetic Goto-Kakizaki rats and non-diabetic Wistar controls. Cardiac function was studied by echocardiography. Myocardial hypertrophy, cardiomyocyte apoptosis and cardiac fibrosis were determined histologically 12 weeks post MI or Sham operation. Western blotting was used to study Caspase-3, Bax, Sirt1, acetylation of p53 and phosphorylation of p38, Akt and FOXO3a. Electrophoretic mobility shift assay was used to assess FOXO3a activity and its nuclear localization. Results: Post-infarct heart failure in diabetic GK rats was associated with pronounced cardiomyocyte hypertrophy, increased interstitial fibrosis and sustained cardiomyocyte apoptosis as compared with their non-diabetic Wistar controls. In the GK rat myocardium, Akt-and FOXO3a-phosphorylation was decreased and nuclear localization of FOXO3a was increased concomitantly with increased PTEN protein expression. Furthermore, increased Sirt1 protein expression was associated with decreased p53 acetylation, and phosphorylation of p38 was increased in diabetic rats with MI. Conclusions: Post-infarct heart failure in diabetic GK rats was associated with more pronounced cardiac hypertrophy, interstitial fibrosis and sustained cardiomyocyte apoptosis as compared to their non-diabetic controls. The present study suggests important roles for Akt-FOXO3a, Sirt1-p53 and p38 MAPK in the regulation of postinfarct cardiac remodeling in type 2 diabetes.
Journal of Pharmacology and Experimental Therapeutics
Cancer treatments are frequently associated with nausea and vomiting despite greatly improved pre... more Cancer treatments are frequently associated with nausea and vomiting despite greatly improved preventive medication. Administration of anti-nausea agents as eye drops might provide an easy and rapid access to the systemic circulation for prevention of nausea and vomiting and for the treatment of breakthrough nausea, but the ocular administration route has rarely been evaluated. Palonosetron is a second-generation 5-HT 3 receptor antagonist approved for prevention and treatment of chemotherapy-induced nausea and vomiting. We compared ocular administration of palonosetron to non-active vehicle eye drops and to intravenous palonosetron in the prevention of cisplatin-induced nausea and vomiting in beagle dogs. Palonosetron ocular drops at the dose of 30 µg/kg reduced cumulative nausea over time as measured with the area under the visual analog scale curve (VAS-AUC) by 98% compared with the vehicle, and reduced nausea-associated dog behavior by 95%. Vomiting was completely prevented with repeated palonosetron ocular dosing. Hydroxypropyl-βcyclodextrin (HP-β-CD) palonosetron formulation was well tolerated locally at the palonosetron concentration of 3 mg/mL. Absorption of palonosetron from eye drops was fast. Ten minutes after ocular administration, palonosetron plasma concentrations were similar compared to intravenous administration, and remained similar for six hours. We conclude that palonosetron is rapidly absorbed into the systemic circulation from eye drops. Ocularly administered palonosetron was well tolerated in the HP-β-CD formulation and was highly effective in the prevention of cisplatin-induced nausea and vomiting. Evaluation of the safety and efficacy of ocular administration of palonosetron is warranted in the prevention and treatment of chemotherapy-induced nausea and vomiting in clinical trials.
Sinus node (SN) pacemaking is based on a coupling between surface membrane ion-channels and intra... more Sinus node (SN) pacemaking is based on a coupling between surface membrane ion-channels and intracellular Ca2+-handling. The fundamental role of the inward Na+/Ca2+ exchanger (NCX) is firmly established. However, little is known about the reverse mode exchange. A simulation study attributed important role to reverse NCX activity, however experimental evidence is still missing. Whole-cell and perforated patch-clamp experiments were performed on rabbit SN cells supplemented with fluorescent Ca2+-tracking. We established 2 and 8 mM pipette NaCl groups to suppress and enable reverse NCX. NCX was assessed by specific block with 1 μM ORM-10962. Mechanistic simulations were performed by Maltsev–Lakatta minimal computational SN model. Active reverse NCX resulted in larger Ca2+-transient amplitude with larger SR Ca2+-content. Spontaneous action potential (AP) frequency increased with 8 mM NaCl. When reverse NCX was facilitated by 1 μM strophantin the Ca2+i and spontaneous rate increased. ORM...
: OR-1855 and OR-1896 are 2 hemodynamically active metabolites of the inodilator levosimendan, wi... more : OR-1855 and OR-1896 are 2 hemodynamically active metabolites of the inodilator levosimendan, with calcium sensitizing activity, but their mechanism of action is still not fully understood. It has been previously reported that the positive inotropic effect of levosimendan is not potentiated by the adenylate cyclase activator forskolin, whereas forskolin does potentiate the effects of the phosphodiesterase (PDE) inhibitor milrinone. To ascertain whether the active metabolites follow the same pattern of levosimendan, the positive inotropic effects of OR- 1855 and OR-1896 were studied in guinea-pig-isolated papillary muscle in the presence and absence of forskolin. OR-1855 and OR-1896 were also tested as inhibitors of PDE-III and PDE-IV. Our results show that 0.1 µM forskolin did not potentiate the positive inotropic effect of OR-1855 or OR-1896, as in the case of the parent compound levosimendan. As in previous studies, the positive inotropic effect of milrinone was markedly potentiated in the presence of forskolin. From these data, we propose an explanation for the divergent behavior of the calcium sensitizing drugs and PDE inhibitors.
Journal of Molecular and Cellular Cardiology, 2021
Repolarization alternans, a periodic oscillation of long-short action potential duration, is an i... more Repolarization alternans, a periodic oscillation of long-short action potential duration, is an important source of arrhythmogenic substrate, although the mechanisms driving it are insufficiently understood. Despite its relevance as an arrhythmia precursor, there are no successful therapies able to target it specifically. We hypothesized that blockade of the sodium-calcium exchanger (NCX) could inhibit alternans. The effects of the selective NCX blocker ORM-10962 were evaluated on action potentials measured with microelectrodes from canine papillary muscle preparations, and calcium transients measured using Fluo4-AM from isolated ventricular myocytes paced to evoke alternans. Computer simulations were used to obtain insight into the drug's mechanisms of action. ORM-10962 attenuated cardiac alternans, both in action potential duration and calcium transient amplitude. Three morphological types of alternans were observed, with differential response to ORM-10962 with regards to APD alternans attenuation. Analysis of APD restitution indicates that calcium oscillations underlie alternans formation. Furthermore, ORM-10962 did not markedly alter APD restitution, but increased post-repolarization refractoriness, which may be mediated by indirectly reduced L-type calcium current. Computer simulations reproduced alternans attenuation via ORM-10962, suggesting that it is acts by reducing sarcoplasmic reticulum release refractoriness. This results from the ORM-10962-induced sodium-calcium exchanger block accompanied by an indirect reduction in L-type calcium current. Using a computer model of a heart failure cell, we furthermore demonstrate that the anti-alternans effect holds also for this disease, in which the risk of alternans is elevated. Targeting NCX may therefore be a useful anti-arrhythmic strategy to specifically prevent calcium driven alternans.
Kidney ischemia-reperfusion (I/R) injury is a common cause of acute kidney injury. We tested whet... more Kidney ischemia-reperfusion (I/R) injury is a common cause of acute kidney injury. We tested whether dexmedetomidine (Dex), an alpha2 adrenoceptor (a2-AR) agonist, protects against kidney I/R injury. Sprague–Dawley rats were divided into four groups: (1) Sham-operated group; (2) I/R group (40 min ischemia followed by 24 h reperfusion); (3) I/R group + Dex (1 lg/kg i.v. 60 min before the surgery), (4) I/R group + Dex (10 lg/kg). The effects of Dex postconditiong (Dex 1 or 10 lg/kg i.v. after reperfusion) as well as the effects of peripheral a2-AR agonism with fadolmidine were also examined. Hemody-namic effects were monitored, renal function measured, and acute tubular dam-age along with monocyte/macrophage infiltration scored. Kidney protein kinase B, toll like receptor 4, light chain 3B, p38 mitogen-activated protein kinase (p38 MAPK), sirtuin 1, adenosine monophosphate kinase (AMPK), and endo-thelial nitric oxide synthase (eNOS) expressions were measured, and kidney transciptome p...
Background and PurposeThe lack of selective sodium–calcium exchanger (NCX) inhibitors has hampere... more Background and PurposeThe lack of selective sodium–calcium exchanger (NCX) inhibitors has hampered the exploration of physiological and pathophysiological roles of cardiac NCX 1.1. We aimed to discover more potent and selective drug like NCX 1.1 inhibitor.Experimental ApproachA flavan series‐based pharmacophore model was constructed. Virtual screening helped us identify a novel scaffold for NCX inhibition. A distinctively different NCX 1.1 inhibitor, ORM‐11372, was discovered after lead optimization. Its potency against human and rat NCX 1.1 and selectivity against other ion channels was assessed. The cardiovascular effects of ORM‐11372 were studied in normal and infarcted rats and rabbits. Human cardiac safety was studied ex vivo using human ventricular trabeculae.Key ResultsORM‐11372 inhibited human NCX 1.1 reverse and forward currents; IC50 values were 5 and 6 nM respectively. ORM‐11372 inhibited human cardiac sodium 1.5 (INa) and hERG KV11.1 currents (IhERG) in a concentration‐d...
Journal of physiology and pharmacology : an official journal of the Polish Physiological Society, 2009
Calcium-sensitizing agents have been shown to improve cardiac function in patients suffering from... more Calcium-sensitizing agents have been shown to improve cardiac function in patients suffering from acute decompensated heart failure, however, their long-term effects on cardiac remodeling and cardiovascular mortality are still largely unknown. In the present study we tested the hypothesis whether OR-1896, an active and long-lasting metabolite of calcium sensitizer levosimendan, prevents cardiovascular mortality and hypertension-induced myocardial remodelling in salt-sensitive Dahl/Rapp rats. OR-1896 was given orally to Dahl/Rapp SS rats on high-salt diet (NaCl 7% w/w) for 7 weeks at two different doses (0.5 and 0.05 mg/kg). OR-1896 prevented salt-induced cardiovascular mortality (survival rate 75 % in OR-1896 treated groups vs 38 % in untreated controls, p<0.01), ameliorated cardiac hypertrophy and improved systolic functions of the heart without major influence on systemic blood pressure. OR-1896 also ameliorated salt-induced increase in cardiac ANP mRNA expression and plasma BN...
Uploads
Papers by Jouko Levijoki