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

Fingolimod plays role in attenuation of myocardial injury related to experimental model of cardiac arrest and extracorporeal life support resuscitation

Background: Sudden cardiac arrest is a major global health concern, and survival of patients with ischemia-reperfusion injury is a leading cause of myocardial dysfunction. The mechanism of this phenomenon is not well understood because of the complex pathophysiological nature of the disease. Aim of the study was to investigate the cardioprotective role of fingolimod in an in vivo model of cardiac arrest and resuscitation.Methods: In this study, an in vivo rat model of cardiac arrest using extracorporeal membrane oxygenation resuscitation monitored by invasive hemodynamic measurement was developed. At the beginning of extracorporeal life support (ECLS), animals were randomly treated with fingolimod (Group A, n = 30) or saline (Group B, n = 30). Half of the animals in each group (Group A1 and B1, n = 15 each) were sacrificed after 1 h, and the remaining animals (Group A2 and B2) after 24 h of reperfusion. Blood and myocardial tissues were collected for analysis of cardiac features, inflammatory biomarkers, and cell signaling pathways.Results: Treatment with fingolimod resulted in activation of survival pathways resulting into reduced inflammation, myocardial oxidative stress and apoptosis of cardiomyocytes. This led to significant improvement in systolic and diastolic functions of the left ventricle and improved contractility index.Conclusions: Sphingosine1phosphate receptor activation with fingolimod improved cardiac function after cardiac arrest supported with ECLS. Present study findings strongly support a cardioprotective role of fingolimod through sphingosine-1-phosphate receptor activation during reperfusion after circulatory arrest

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

S-Nitroso Human Serum Albumin Enhances Left Ventricle Hemodynamic Performance and Reduces Myocardial Damage after Local Ischemia–Reperfusion Injury

Endothelial nitric oxide (NO) production is crucial in maintaining vascular homeostasis. However, in the context of ischemia–reperfusion (I/R) injury, uncoupled endothelial nitric oxide synthase (eNOS) can exacerbate reactive oxygen species (ROS) generation. Supplementation with S-nitroso human serum albumin (S-NO-HSA) offers a potential solution by mitigating eNOS uncoupling, thereby enhancing NO bioavailability. In a study conducted at the University of Verona, male rats underwent thoracotomy followed by 30 min left anterior descendant coronary (LAD) occlusion and subsequent reperfusion. Hemodynamic parameters were meticulously assessed using a conductance catheter inserted via the carotid artery. The rats were stratified into two main groups based on reperfusion duration and the timing of drug infusion, with the effects of S-NO-HSA evaluated after 2 or 24 h. Remarkably, intravenous administration of S-NO-HSA, initiated before or during ischemia, exhibited notable benefits. It significantly improved left ventricular function, safeguarded energetic substrates such as phosphocreatine and ATP, and sustained glutathione levels akin to basal conditions, indicative of diminished oxidative stress. The data from this study strongly suggest a protective role for S-NO-HSA in mitigating I/R injury induced by LAD artery occlusion, a phenomenon observed at both 2 and 24 h post-reperfusion. These findings underscore the promising therapeutic potential of NO supplementation in alleviating myocardial damage subsequent to ischemic insult

Minimally invasive cardiopulmonary bypass in pediatrics

Minimally invasive cardiopulmonary bypass (MICPB) systems aim is to improve the biocompatibility of cardiopulmonary bypass components, minimize adverse effects, and improve organ protection. MICPB consists of a closed circuitry with a membrane oxygenator and a centrifugal pump, but without the venous reservoir with reduced priming volume, decreased surface area, and less blood–air interface. The advantages of using MICPB are lower systemic inflammatory response, reduced hemodilution, less activation of coagulation cascades, and lower requirement for blood transfusions. The use of MICPB brings clinical benefits, lower incidence of myocardial damage, atrial fibrillation, low cardiac output syndrome, neurological complications, and respiratory insufficiency with a reduced postoperative hospital stay. The clinical acceptance and application of MICPB in adult cardiac surgery are not widely spread. The risk of air embolism in closed MICPB systems limits the clinical application. The complex air-handling characteristics with increased risk of air embolism and the difficult volume handling in ventricular dilatation or excessive bleeding represent the most considerable objection regarding omitted venous reservoirs. Due to the very nature of congenital heart disease repair in newborns, infants, and children, a closed-circuit MICPB is hardly applicable with few exceptions. However, strategies to miniaturize the CPB circuitry and manage perfusion have been developed in pediatric cardiac surgery through the years, thereby representing a valid surrogate of a MICPB in children

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