TRATTAMENTI NEUROPROTETTIVI SU DANNO DA ISCHEMIA/RIPERFUSIONE IN MODELLI SPERIMENTALI DI ARRESTO CARDIACO ED EXTRA CORPOREAL LIFE SUPPORT

INTRODUZIONE: La sopravvivenza dopo un arresto cardiaco \ue8 strettamente correlata a una precoce riperfusione e ripresa della circolazione sanguigna spontanea (ROSC). Tuttavia, la sopravvivenza dipende anche dall\u2019instaurarsi di una serie di meccanismi fisiopatologici che caratterizzano la cosiddetta sindrome post arresto. Tra i pi\uf9 importanti meccanismi c\u2019\ue8 lo sviluppo di un danno cerebrale. Nonostante l\u2019avanzamento delle tecniche di riperfusione anche tramite aiuto di extracorporeal life support (ECLS) la sopravvivenza a 30 giorni dei pazienti che superano l\u2019evento acuto arresto cardiaco \ue8 inferiore al 30%. SCOPO: Scopo degli studi effettuati durante il Corso di Dottorato in Scienze Cardiovascolari \ue8 stato di studiare le capacit\ue0 neuro protettive di diversi approcci terapeutici applicati in 3 modelli sperimentali di arresto cardiaco trattato con ECLS. I trattamenti studiati sono stati: ipotermia terapeutica, ipotermia farmacologica, infusione della soluzione ALM (Adenosina-Lidocaina-Magnesio), perfusione cerebrale selettiva anterograda (SCP) durante arresto di circolo, infusione di Ossido Nitrico gassoso (NO), e riscaldamento lento dopo ipotermia accidentale. MATERIALI E METODI: Tutti gli esperimenti si sono svolti su ratti di sesso maschile, del ceppo Sprague Dawley, del peso di circa 500 g nelle strutture del CIRSAL (Centro Interdipartimentale di Servizio alla Ricerca Sperimentale) presso la Facolt\ue0 di Medicina e Chirurgia di Verona. In particolare, sono stati ideati, realizzati ed utilizzati 3 modelli sperimentali di arresto cardiaco e riperfusione con circuito miniaturizzato di Circolazione Extra Corporea (CPB): Modello di arresto cardiaco sottoposto ad ECLS, modello di arresto di circolo e SCP in CPB, modello di arresto cardiaco da ipotermia accidentale e riscaldamento tramite ECLS. RISULTATI: Nel modello di arresto cardiaco sottoposto a ECLS dall\u2019analisi western blot dei cervelli congelati si \ue8 evidenziato un ruolo protettivo, antinfiammatorio (IL-10, IL-6, MCP1, iNOS) e anti-apoptotico (RBM3) dell\u2019ipotermia topica e dell\u2019ipotermia farmacologica indotta con WIN55-212. Lo stesso modello di arresto \ue8 stato utilizzato sottoponendo un gruppo di ratti a trattamento con ALM: le analisi sui cervelli congelati hanno mostrato un certo grado di riduzione dell\u2019infiammazione (IL-6, Il-10) ma senza risultati definitivi certi. Molto pi\uf9 efficace si \ue8 invece dimostrato il trattamento con ossido nitrico inalatorio somministrato tramite ossigenatore con riduzione della risposta infiammatoria e dell\u2019ipossia visualizzata su preparati di cervelli in formalina con marker immunofluorescenti per microglia (Iba1) e per Tioli. Nel modello di arresto di circolo e SCP in CPB, durante gli esperimenti \ue8 stato possibile realizzare l\u2019obiettivo di dimostrare la validit\ue0 del nuovo modello attraverso l\u2019analisi EEG, i preparati istologici e l\u2019analisi dei marker di biologia molecolare (Caspasi 3, RBM3, VEGF, PARP) ma non \ue8 stata dimostrata l\u2019efficacia del trattamento con ALM infusa direttamente nel circolo cerebrale. Nel modello di arresto cardiaco da ipotermia accidentale e riscaldamento tramite ECLS \ue8 stato possibile evidenziare l\u2019importanza di un riscaldamento lento rispetto ad un riscaldamento veloce tramite riduzione dell\u2019attivazione della microglia (Iba1) riduzione, evidenziato alla analisi con Risonanza Magnetica, dell\u2019edema cerebrale e miglioramento della perfusione cerebrale a 24 ore dall\u2019arresto e riduzione dei marker infiammatori e di stress ossidativo evidenziati all\u2019analisi immunoistochimica (TNF\u3b1, IL-6, CCl5, ICAM1, Malondialdeide, Nitrotirosina). CONCLUSIONI: La frequenza in questi anni nel reparto di Cardiochirurgia ha fatto crescere in me la consapevolezza dell\u2019importanza di trovare metodi sempre migliori e pi\uf9 efficaci per ridurre i danni cerebrali dopo arresto cardiaco o arresto di circolo durante chirurgia dell\u2019arco aortico. Gli studi effettuati sui modelli sperimentali creati presso il laboratorio di ricerca cardiovascolare durante i 3 anni di Dottorato si sono evoluti portando alla creazione di modelli sperimentali sempre pi\uf9 realistici e complessi che riproducono ci\uf2 che avviene nella pratica clinica. Il lavoro effettuato pone delle basi scientifiche e metodologiche per la realizzazione di nuovi ulteriori studi sviluppando con ulteriori analisi i trattamenti sperimentali gi\ue0 proposti e utilizzando i nostri modelli sperimentali per studi su nuovi trattamenti nel campo della cardio e neuroprotezione nel danno da ischemia e riperfusione post arresto cardiaco accidentale o cardioplegico

Role of calcium desensitization in the treatment of myocardial dysfunction after deep hypothermic circulatory arrest

Abstract Introduction Rewarming from deep hypothermic circulatory arrest (DHCA) produces calcium desensitization by troponin I (cTnI) phosphorylation which results in myocardial dysfunction. This study investigated the acute overall hemodynamic and metabolic effects of epinephrine and levosimendan, a calcium sensitizer, on myocardial function after rewarming from DHCA. Methods Forty male Wistar rats (400 to 500 g) underwent cardiopulmonary bypass (CPB) through central cannulation and were cooled to a core temperature of 13°C to 15°C within 30 minutes. After DHCA (20 minutes) and CPB-assisted rewarming (60 minutes) rats were randomly assigned to 60 minute intravenous infusion with levosimendan (0.2 μg/kg/min; n = 15), epinephrine (0.1 μg/kg/min; n = 15) or saline (control; n = 10). Systolic and diastolic functions were evaluated at different preloads with a conductance catheter. Results The slope of left ventricular end-systolic pressure volume relationship (Ees) and preload recruitable stroke work (PRSW) recovered significantly better with levosimendan compared to epinephrine (Ees: 85 ± 9% vs 51 ± 11%, P\u3c0.003 and PRSW: 78 ± 5% vs 48 ± 8%, P\u3c0.005; baseline: 100%). Levosimendan but not epinephrine reduced left ventricular stiffness shown by the end-diastolic pressure-volume relationship and improved ventricular relaxation (Tau). Levosimendan preserved ATP myocardial content as well as energy charge and reduced plasma lactate concentrations. In normothermia experiments epinephrine in contrast to Levosimendan increased cTnI phosphorylation 3.5-fold. After rewarming from DHCA, cTnI phosphorylation increased 4.5-fold in the saline and epinephrine group compared to normothermia but remained unchanged with levosimendan. Conclusions Levosimendan due to prevention of calcium desensitization by cTnI phosphorylation is more effective than epinephrine for treatment of myocardial dysfunction after rewarming from DHCA

Cardioprotective effects of sphingosine-1-phosphate receptor immunomodulator fty720 in a clinically relevant model of cardioplegic arrest and cardiopulmonary bypass

Objective: FTY720, an immunomodulator derived from sphingosine-1-phosphate, has recently demonstrated its immunomodulatory, anti-inflammatory, anti-oxidant, anti-apoptotic and anti-inflammatory properties. Furthermore, FTY720 might be a key pharmacological target for preconditioning. In this preclinical model, we have investigated the effects of FTY720 on myocardium during reperfusion in an experimental model of cardioplegic arrest (CPA) and cardiopulmonary bypass. Methods: 30 Sprague-Dawley rats (300-350 g) were randomized into two groups: Group-A, treated with FTY720 1 mg/kg via intravenous cannulation, and Group-B, as control. After 15 min of treatment, rats underwent CPA for 30 min followed by initiation of extracorporeal life support for 2 h. Support weaning was done, and blood and myocardial tissues were collected for analysis. Hemodynamic parameters, inflammatory mediators, nitro-oxidative stress, neutrophil infiltration, immunoblotting analysis, and immunohistochemical staining were analyzed and compared between groups. Results: FTY720 treatment activated the Akt/Erk1/2 signaling pathways, reduced the level of inflammatory mediators, activated antiapoptotic proteins, and inhibited proapoptotic proteins, leading to reduced nitro-oxidative stress and cardiomyocyte apoptosis. Moreover, significant preservation of high-energy phosphates were observed in the FTY720-treated group. This resulted in improved recovery of left ventricular systolic and diastolic functions. Conclusion: The cardioprotective mechanism in CPA is associated with activation of prosurvival cell signaling pathways that prevents myocardial damage. FTY720 preserves high-energy phosphates attenuates myocardial inflammation and oxidative stress, and improves cardiac function

Sphingosine 1-Phosphate Receptor Modulator Fingolimod (FTY720) Attenuates Myocardial Fibrosis in Post-heterotopic Heart Transplantation

Background and Objective: Sphingosine 1-phosphate (S1P), and S1P receptor modulator fingolimod have been suggested to play important cardioprotective role in animal models of myocardial ischemia/reperfusion injuries. To understand the cardioprotective function of S1P and its mechanism in vivo, we analyzed apoptotic, inflammatory biomarkers, and myocardial fibrosis in an in vivo heterotopic rat heart transplantation model.Methods: Heterotopic heart transplantation is performed in 60 Sprague–Dawley (SD) rats (350–400 g). The heart transplant recipients (n = 60) are categorized into Group A (control) and Group B (fingolimod treated 1 mg/kg intravenous). At baseline with 24 h after heart transplantation, blood and myocardial tissue are collected for analysis of myocardial biomarkers, apoptosis, inflammatory markers, oxidative stress, and phosphorylation of Akt/Erk/STAT-3 signaling pathways. Myocardial fibrosis was investigated using Masson’s trichrome staining and L-hydroxyline.Results: Fingolimod treatment activates both Reperfusion Injury Salvage Kinase (RISK) and Survivor Activating Factor Enhancement (SAFE) pathways as evident from activation of anti-apoptotic and anti-inflammatory pathways. Fingolimod treatment caused a reduction in myocardial oxidative stress and hence cardiomyocyte apoptosis resulting in a decrease in myocardial reperfusion injury. Moreover, a significant (p < 0.001) reduction in collagen staining and hydroxyproline content was observed in fingolimod treated animals 30 days after transplantation demonstrating a reduction in cardiac fibrosis.Conclusion: S1P receptor activation with fingolimod activates anti-apoptotic and anti-inflammatory pathways, leading to improved myocardial salvage causing a reduction in cardiac fibrosis

Temperature Management During Circulatory Arrest in Cardiac Surgery

Surgery for complex aortic pathologies, such as acute dissections and aneurysms involving the aortic arch, remains one of the most technically and strategically challenging intervention in aortic surgery, requiring thorough understanding not only of cardiovascular physiology but also of neurophysiology (cerebral and spinal cord), and is still associated with significant mortality and morbidity. The introduction of deep hypothermia in the mid 1970s, allowing defined periods of circulatory arrest, has made possible the advent of modern aortic surgery requiring prolonged ischemic tolerance of central nervous system. In the late 1980s, when deep hypothermic circulatory arrest was the standard operative strategy for aortic surgery, selective cerebral perfusion, as an adjunct to deep hypothermia, made possible excellent neuroprotection and improved overall outcome. This encouraged the use of selective cerebral perfusion in combination with steadily increasing body core temperatures, a trend culminating in progressive promotion of moderate to mild hypothermia and even normothermia. The motivation for progressive temperature elevation was the limitation of adverse effects of deep hypothermia, in particular, reduction of systemic inflammatory response (and organ dysfunctions) and diminution of the risk of severe postoperative bleeding. However, adverse outcomes due to inappropriate temperature management (core temperatures too high for the required duration of circulatory arrest) are probably underreported. Indeed, complications historically associated with hypothermia are possibly overestimated

Elevated cardiac troponin in clinical scenarios beyond obstructive coronary artery disease

In this systematic review article, we aim to summarize the most up-to-date evidence regarding elevations of cardiac troponin, especially in clinical scenarios other than obstructive coronary artery disease. The accurate interpretation of raised cardiac troponin is challenging because it relies on unconfirmed postulations and dogmatic knowledge (e.g., the exclusive provenience of cardiac troponin from cardiac myocytes), based on which every troponin elevation is assumed to definitely indicate myocardial damage. Indeed, the investigation of the pathophysiologic mechanism leading to the release in the bloodstream of cardiac biomarkers should be the first step of the diagnostic process to fully understand the clinical significance of the elevated serum levels and identify the best management. A prominent effort should be put in place to identify the contribution of potential confounding factors, both cardiac and non-cardiac in etiology, with the ability to affect synthesis and clearance of cardiac biomarkers. Regardless of the underlying cause, it is well established that cardiovascular biomarkers are increasingly useful to further risk stratification and prognosticate patients. Accordingly, we sought to clarify the meaning and impact of elevated cardiac troponin in those frequently encountered real- world scenarios presenting clinicians with a diagnostic dilemma, with the final goal of facilitating the diagnosis and help optimize individually tailored treatment strategies

Urgent Surgery for Pituitary Adenoma Bleeding After Coronary Bypass Surgery

Pituitary gland adenoma bleeding is an uncommon complication after coronary artery surgery. Clinical presentation may be variable. We report a case of hemorrhagic complication of a pituitary gland adenoma requiring urgent surgery in a 60-year-old male patient who underwent coronary artery bypass grafting operation

Ventricular and pulmonary vascular remodeling induced by pulmonary overflow in a chronic model of pretricuspid shunt

ObjectivesCurrent preclinical models of pulmonary arterial hypertension do not reproduce the clinical characteristics of congenital heart anomalies. Aortocaval shunt is relevant to a variety of clinical conditions. The pathophysiology and possible determination of pulmonary hypertension in this model are still undefined.MethodsA method to create a standardized and reproducible aortocaval shunt was developed in rats. After creation of the shunt, the animals were followed up for 20 weeks and a sham laparotomy was used as a control. The chronic effects of volume overload on the right and left ventricles and pulmonary hemodynamic modifications were evaluated by biventricular catheterization, echocardiography, and magnetic resonance. Pulmonary vascular changes were defined by histology.ResultsAn increased right ventricular end-diastolic area was confirmed by echocardiography. Left ventricular overload and decreased biventricular ejection fraction were demonstrated by magnetic resonance after 20 weeks in the shunt group compared with the controls (left ventricle, 50% ± 5% vs 62% ± 3%, P = .029; right ventricle, 53% ± 2% vs 65% ± 2%, P = .036). Preload recruitable stroke work of left and right ventricles decreased after 20 weeks in shunt rats (left ventricle: 36 ± 7 vs 98 ± 5, P = .004; right ventricle: 19 ± 2 vs 32 ± 9, P = .047). At the same time point, catheterization showed that effective pulmonary arterial elastance was increased only in the shunt group (1.29 ± 0.20 vs 0.14 ± 0.06 mm Hg/μL; P = .004). Histology showed medial hypertrophy, small artery luminal narrowing, and occlusion.ConclusionsThe aortocaval shunt model reliably produces right ventricular volume overload and secondary pulmonary hypertension. Due to a combination of left ventricular dysfunction and pulmonary overflow, the pulmonary hypertension produced shows features similar to those found in patients with chronic atrial-level shunt

Temperature Variation After Rewarming from Deep Hypothermic Circulatory Arrest Is Associated with Survival and Neurologic Outcome

Therapeutic hypothermia is recommended by international guidelines after cardio-circulatory arrest. However, the effects of different temperatures during the first 24 hours after deep hypothermic circulatory arrest (DHCA) for aortic arch surgery on survival and neurologic outcome are undefined. We hypothesize that temperature variation after aortic arch surgery is associated with survival and neurologic outcome. In the period 2010-2014, a total of 210 consecutive patients undergoing aortic arch surgery with DHCA were included. They were retrospectively divided into three groups by median nasopharyngeal temperature within 24 hours after rewarming: hypothermia (<36\ub0C; n\u2009=\u200965), normothermia (36-37\ub0C; n\u2009=\u2009110), and hyperthermia (>37\ub0C; n\u2009=\u200935). Multivariate stepwise logistic and linear regressions were performed to determine whether different temperature independently predicted 30-day mortality, stroke incidence, and neurologic outcome assessed by cerebral performance category (CPC) at hospital discharge. Compared with normothermia, hyperthermia was independently associated with a higher risk of 30-day mortality (28.6% vs. 10.9%; odds ratio [OR] 2.8; 95% confidence interval [CI], 1.1-8.6; p\u2009=\u20090.005), stroke incidence (64.3% vs. 9.1%; OR 9.1; 95% CI, 2.7-23.0; p\u2009=\u20090.001), and poor neurologic outcome (CPC 3-5) (68.8% vs. 39.6%; OR 4.8; 95% CI, 1.4-8.7; p\u2009=\u20090.01). No significant differences were demonstrated between hypothermia and normothermia. Postoperative hypothermia is not associated with a better outcome after aortic arch surgery with DHCA. However, postoperative hyperthermia (>37\ub0C) is associated with high stroke incidence, poor neurologic outcome, and increased 30-day mortality. Target temperature management in the first 24 hours after surgery should be evaluated in prospective randomized trials