Effect of Colchicine on Myocardial Injury Induced by Trypanosoma cruzi in Experimental Chagas Disease

Background: The hallmark of Chagas disease (CD) is multifocal myocarditis and extensive fibrosis. We investigated the potential effect of colchicine on myocardial remodeling in experimental CD. Methods and Results: One hundred Syrian hamsters were randomly divided into noninfected untreated control (CG), noninfected control treated with colchicine (COLG 0.4 mg kg(-1) d(-1) by gavage), infected (IG), and infected treated with colchicine (ICOLG, 0.4 mg kg(-1) d(-1)) groups. The interstitial collagen volume fraction (ICVF) was evaluated by videomorphometry with picrosirius red staining. The gelatinolytic activities of matrix metalloproteinase (MMP) 2 were examined with the use of zymography. Myocarditis was described according to the Dallas criteria. Statistical comparisons were performed with parametric analysis of variance and Tukey test. ICVF (%) accumulation was attenuated in infected colchicine-treated animals in the left (CG 0.81 +/- 0.13, COLG 0.85 +/- 0.13, IG: 1.35 +/- 0.31,* ICOLG 1.06 +/- 0.19; *P < .05 compared with ICOLG) and right ventricles (CG 1.4 +/- 0.36, COLG 1.26 +/- 0.14, IG 1.97 +/- 0.058,* ICOLG: 1.52 +/- 0.23; *P < .05 compared with ICOLG). A significant increase in MMP-2 enzymatic activity (UA) was observed in ICOLG (17,432.8*) compared with GC (3731.6), COLG (2,792.6), and IG (4,286.3; *P < .001). In IG, 66% of animals had myocarditis compared with only 49% in ICOLG. Conclusions: Colchicine had a protective effect on myocardium, indicated by decreased interstitial myocardial fibrosis, increased intensity of MMP-2, and attenuated myocardial inflammation. (J Cardiac Fail 2012;18:654-659)

Parathyroid hormone and phosphorus overload in uremia: impact on cardiovascular system

Background. Cardiac remodeling in uremia is characterized by left ventricular hypertrophy, interstitial fibrosis and microvascular disease. Cardiovascular disease is the leading cause of death in uremic patients, but coronary events alone are not the prevalent cause, sudden death and heart failure are. We studied the cardiac remodeling in experimental uremia, evaluating the isolated effect of parathyroid hormone (PTH) and phosphorus. Methods. Wistar rats were submitted to parathyroidectomy (PTx) and 5/6 nephrectomy (Nx); they also received vehicle (V) and PTH at normal (nPTH) or high (hPTH) doses. They were fed with a poor-phosphorus (pP) or rich-phosphorus (rP) diet and were divided into the following groups: 'Sham': G1 (V + normal-phosphorus diet (np)) and 'Nx + PTx': G2 (nPTH + pP), G3 (nPTH + rP), G4 (hPTH + pP) and G5 (hPTH + rP). After 8 weeks, biochemical analysis, myocardium morphometry and arteriolar morphological analysis were performed. In addition, using immunohistochemical analysis, we evaluated angiotensin II, alpha-actin, transforming growth factor-beta (TGF-beta) and nitrotyrosine, as well as fibroblast growth factor-23 (FGF-23), fibroblast growth factor receptor-1 (FGFR-1) and runt-related transcription factor-2 (Runx-2) expression. Results. Nx animals presented higher serum creatinine levels as well as arterial hypertension. Higher PTH levels were associated with myocardial hypertrophy and fibrosis as well as a higher coronary lesion score. High PTH animals also presented a higher myocardial expression of TGF-beta, angiotensin II, FGF-23 and nitrotyrosine and a lower expression of alpha-actin. Phosphorus overload was associated with higher serum FGF-23 levels and Runx-2, as well as myocardial hypertrophy. FGFR-1 was positive in the cardiomyocytes of all groups as well as in calcified coronaries of G4 and G5 whereas Runx-2 was positive in G3, G4 and G5. Conclusion. In uremia, PTH and phosphorus overload are both independently associated with major changes related to the cardiac remodeling process, emphasizing the need for a better control of these factors in chronic kidney disease.

Effects of aerobic exercise training on cardiac renin-angiotensin system in an obese Zucker rat strain

Objective: Obesity and renin angiotensin system (RAS) hyperactivity are profoundly involved in cardiovascular diseases, however aerobic exercise training (EXT) can prevent obesity and cardiac RAS activation. The study hypothesis was to investigate whether obesity and its association with EXT alter the systemic and cardiac RAS components in an obese Zucker rat strain. Methods: The rats were divided into the following groups: Lean Zucker rats (LZR); lean Zucker rats plus EXT (LZR+EXT); obese Zucker rats (OZR) and obese Zucker rats plus EXT (OZR+EXT). EXT consisted of 10 weeks of 60-min swimming sessions, 5 days/week. At the end of the training protocol heart rate (HR), systolic blood pressure (SBP), cardiac hypertrophy (CH) and function, local and systemic components of RAS were evaluated. Also, systemic glucose, triglycerides, total cholesterol and its LDL and HDL fractions were measured. Results: The resting HR decreased (, 12%) for both LZR+EXT and OZR+EXT. However, only the LZR+EXT reached significance (p, 0.05), while a tendency was found for OZR versus OZR+EXT (p = 0.07). In addition, exercise reduced (57%) triglycerides and (61%) LDL in the OZR+EXT. The systemic angiotensin I-converting enzyme (ACE) activity did not differ regardless of obesity and EXT, however, the OZR and OZR+EXT showed (66%) and (42%), respectively, less angiotensin II (Ang II) plasma concentration when compared with LZR. Furthermore, the results showed that EXT in the OZR prevented increase in CH, cardiac ACE activity, Ang II and AT2 receptor caused by obesity. In addition, exercise augmented cardiac ACE2 in both training groups. Conclusion: Despite the unchanged ACE and lower systemic Ang II levels in obesity, the cardiac RAS was increased in OZR and EXT in obese Zucker rats reduced some of the cardiac RAS components and prevented obesity-related CH. These results show that EXT prevented the heart RAS hyperactivity and cardiac maladaptive morphological alterations in obese Zucker rats.

Deficient Regulatory T Cell Activity and Low Frequency of IL-17-Producing T Cells Correlate with the Extent of Cardiomyopathy in Human Chagas' Disease

Background: Myocardium damage during Chagas' disease results from the immunological imbalance between pro-and production of anti-inflammatory cytokines and has been explained based on the Th1-Th2 dichotomy and regulatory T cell activity. Recently, we demonstrated that IL-17 produced during experimental T. cruzi infection regulates Th1 cells differentiation and parasite induced myocarditis. Here, we investigated the role of IL-17 and regulatory T cell during human Chagas' disease. Methodology/Principal Findings: First, we observed CD4(+)IL-17(+) T cells in culture of peripheral blood mononuclear cells (PBMC) from Chagas' disease patients and we evaluated Th1, Th2, Th17 cytokine profile production in the PBMC cells from Chagas' disease patients (cardiomyopathy-free, and with mild, moderate or severe cardiomyopathy) cultured with T. cruzi antigen. Cultures of PBMC from patients with moderate and severe cardiomyopathy produced high levels of TNF-alpha, IFN-gamma and low levels of IL-10, when compared to mild cardiomyopathy or cardiomyopathy-free patients. Flow cytometry analysis showed higher CD4(+)IL-17(+) cells in PBMC cultured from patients without or with mild cardiomyopathy, in comparison to patients with moderate or severe cardiomyopathy. We then analyzed the presence and function of regulatory T cells in all patients. All groups of Chagas' disease patients presented the same frequency of CD4(+)CD25(+) regulatory T cells. However, CD4(+)CD25(+) T cells from patients with mild cardiomyopathy or cardiomyopathy-free showed higher suppressive activity than those with moderate and severe cardiomyopathy. IFN-gamma levels during chronic Chagas' disease are inversely correlated to the LVEF (P = 0.007, r = -0.614), while regulatory T cell activity is directly correlated with LVEF (P = 0.022, r = 0.500). Conclusion/Significance: These results indicate that reduced production of the cytokines IL-10 and IL-17 in association with high levels of IFN-gamma and TNF-alpha is correlated with the severity of the Chagas' disease cardiomyopathy, and the immunological imbalance observed may be causally related with deficient suppressor activity of regulatory T cells that controls myocardial inflammation.

Myocardial performance in conscious streptozotocin diabetic rats

Abstract Background In spite of a large amount of studies in anesthetized animals, isolated hearts, and in vitro cardiomyocytes, to our knowledge, myocardial function was never studied in conscious diabetic rats. Myocardial performance and the response to stress caused by dobutamine were examined in conscious rats, fifteen days after the onset of diabetes caused by streptozotocin (STZ). The protective effect of insulin was also investigated in STZ-diabetic rats. Methods Cardiac contractility and relaxation were evaluated by means of maximum positive (+dP/dtmax) and negative (-dP/dtmax) values of first derivative of left ventricular pressure over time. In addition, it was examined the myocardial response to stress caused by two dosages (1 and 15 μg/kg) of dobutamine. One-way analysis of variance (ANOVA) was used to compare differences among groups, and two-way ANOVA for repeated measure, followed by Tukey post hoc test, to compare the responses to dobutamine. Differences were considered significant if P < 0.05. Results Basal mean arterial pressure, heart rate, +dP/dtmax and -dP/dtmax were found decreased in STZ-diabetic rats, but unaltered in control rats treated with vehicle and STZ-diabetic rats treated with insulin. Therefore, insulin prevented the hemodynamic and myocardial function alterations observed in STZ-diabetic rats. Lower dosage of dobutamine increased heart rate, +dP/dtmax and -dP/dtmax only in STZ-diabetic rats, while the higher dosage promoted greater, but similar, responses in the three groups. In conclusion, the results indicate that myocardial function was remarkably attenuated in conscious STZ-diabetic rats. In addition, the lower dosage of dobutamine uncovered a greater responsiveness of the myocardium of STZ-diabetic rats. Insulin preserved myocardial function and the integrity of the response to dobutamine of STZ-diabetic rats. Conclusion The present study provides new data from conscious rats showing that the cardiomyopathy of this pathophysiological condition was expressed by low indices of contractility and relaxation. In addition, it was also demonstrated that these pathophysiological features were prevented by the treatment with insulin.

Left ventricular free wall impeding rupture in post-myocardial infarction period diagnosed by myocardial contrast echocardiography: Case report

Abstract Background: Left ventricular free wall rupture occurs in up to 10% of the in-hospital deaths following myocardial infarction. It is mainly associated with posterolateral myocardial infarction and its antemortem diagnosis is rarely made. Contrast echocardiography has been increasingly used for the evaluation of myocardial perfusion in patients with acute myocardial infarction, with important prognostic implications. In this case, we reported its use for the detection of a mechanical complication following myocardial infarction. Case presentation: A 50-year-old man with acute myocardial infarction in the lateral wall underwent myocardial contrast echocardiography for the evaluation of myocardial perfusion in the third day post-infarction. A perfusion defect was detected in lateral and inferior walls as well as the presence of contrast extrusion from the left ventricular cavity into the myocardium, forming a serpiginous duct extending from the endocardium to the epicardial region of the lateral wall, without communication with the pericardial space. Magnetic resonance imaging confirmed the diagnosis of impending rupture of the left ventricular free wall. While waiting for cardiac surgery, patient presented with cardiogenic shock and died. Anatomopathological findings were consistent with acute myocardial infarction in the lateral wall and a left ventricular free wall rupture at the infarct site. Conclusion: This case illustrates the early diagnosis of left ventricular free wall rupture by contrast echocardiography. Due to its ability to be performed at bedside this modality of imaging has the potential to identify this catastrophic condition in patients with acute myocardial infarction and help to treat these patients with emergent surgery.

Effects of vitamin C supplementation on acute phase Chagas disease in experimentally infected mice with Trypanosoma cruzi QM1 strain

The tissue changes that occur in Chagas disease are related to the degree of oxidative stress and antioxidant capacity of affected tissue. Studies with vitamin C supplementation did not develop oxidative damage caused by Chagas disease in the host, but other studies cite the use of peroxiredoxins ascorbate - dependent on T. cruzi to offer protection against immune reaction. Based on these propositions, thirty "Swiss" mice were infected with T. cruzi QM1 strain and treated with two different vitamin C doses in order to study the parasitemia evolution, histopathological changes and lipid peroxidation biomarkers during the acute phase of Chagas disease. The results showed that the parasite clearance was greater in animals fed with vitamin C overdose. There were no significant differences regarding the biomarkers of lipid peroxidation and inflammatory process or the increase of myocardium in animals treated with the recommended dosage. The largest amount of parasite growth towards the end of the acute phase suggests the benefit of high doses of vitamin C for trypomastigotes. The supplementation doesn't influence the production of free radicals or the number of amastigote nests in the acute phase of Chagas disease.

Morfologia dos cardiomiócitos e quantificação do colágeno no miocárdio de ratas tratadas com isoflavonas ou estrogênios

OBJETIVOS: Avaliar a morfologia dos cardiomiócitos e quantificar o colágeno presente no miocárdio de ratas tratadas com extrato concentrado de soja ou 17β-estradiol (E2). MÉTODOS: Vinte e oito ratas foram divididas em quatro grupos: GCtrl - fase de estro; GOvx - ovariectomizadas (Ovx); GIso - Ovx tratadas com extrato de soja (150 mg/kg, por dia); GE2 - Ovx tratadas com E2 (10 µg/kg, por dia). As drogas e o veículo (0,2 mL de propilenoglicol) foram administrados após 30 dias da realização da ovariectomia, por 30 dias consecutivos. No último dia os animais foram anestesiados, o coração retirado, mergulhado em formaldeído a 10%, e fragmentos dos ventrículos submetidos a processamento histológico, sendo os cortes corados pela hematoxilina e eosina ou pelo picrosirius-red. As análises histomorfométricas (contagem, volume nuclear e quantificação do colágeno) foram realizadas em microscópio de luz e software AxioVision Rel. 4.2, sendo o colágeno determinado pelo programa Imagelab 2000. Os dados foram submetidos ao teste de ANOVA complementado pelo teste de Tukey (p<0,05). RESULTADOS: Notamos maior quantidade de núcleos de cardiomiócitos nos animais dos grupos Ovx e Iso do que no GE2 e GCtrl (GOvx=121,7±20,2=GIso=92,8±15,4>GE2=70,5±14,8=GCtrl=66,3±9,6; p<0,05), sendo o volume nuclear maior nos animais do grupo Ctrl e E2 (GE2=35,7±4,8=GCtrl=29,9±3,6>GIso=26,5±4,5=GOvx=22,4±2,9; p<0,05). Com relação ao colágeno notamos maior concentração no grupo Ovx (GOvx=5,4±0,1>GCtrl=4,0±0,1=GIso=4,4±0,08=GE2=4,3±0,5; p<0,05). CONCLUSÕES: Os estrogênios previnem a diminuição do volume nuclear dos cardiomiócitos e a deposição de colágeno entre as fibras musculares cardíacas. Já a administração de isoflavonas previne somente a deposição de colágeno, o que pode preservar as propriedades mecânicas das fibras cardíacas.

Angiotensin II type 2 receptor (AT2R) is associated with increased tolerance of the hyperthyroid heart to ischemia-reperfusion

Background Thyroid hormone induces cardiac hypertrophy and preconditions the myocardium against Ischemia/Reperfusion (I/R) injury. Type 2 Angiotensin II receptors (AT2R) are shown to be upregulated in cardiac hypertrophy observed in hyperthyroidism and this receptor has been reported to mediate cardioprotection against ischemic injury. Methods The aim of the present study was to evaluate the role of AT2R in the recovery of myocardium after I/R in isolated hearts from T3 treated rats. MaleWistar rats were treated with triiodothyronine (T3; 7 μg/100 gBW/day, i.p.) in the presence or not of a specific AT2R blocker (PD123,319; 10 mg/Kg) for 14 days, while normal rats served as control. After treatment, isolated hearts were perfused in Langendorff mode; after 30 min of stabilization, hearts were subjected to 20 min of zero-flow global ischemia followed by 25 min, 35 min and 45 min of reperfusion. Results T3 treatment induced cardiac hypertrophy, which was not changed by PD treatment. Post-ischemic recovery of cardiac function was increased in T3-treated hearts after 35 min and 45 min of reperfusion as compared to control and the ischemic contracture was accelerated and intensified. AT2R blockade was able to return the evaluated functional parameters of cardiac performance (LVDP, +dP/dtmáx and −dP/dtmin) to the control condition. Furthermore, AT2R blockade prevented the increase in AMPK expression levels induced by T3, suggesting its possible involvement in this process. Conclusion AT2R plays a significant role in T3-induced cardioprotection.

Stem Cells as a therapy for myocardial infarction in animal models

Advances in stem cell biology have challenged the notion that infarcted myocardium is irreparable. The pluripotent ability of stem cells to differentiate into specialized cell lines began to garner intense interest within cardiology when it was shown in animal models that intramyocardial injection of bone marrow stem cells (MSCs), or the mobilization of bone marrow stem cells with spontaneous homing to myocardium, could improve cardiac function and survival after induced myocardial infarction (MI) [1, 2]. Furthermore, the existence of stem cells in myocardium has been identified in animal heart [3, 4], and intense research is under way in an attempt to clarify their potential clinical application for patients with myocardial infarction. To date, in order to identify the best one, different kinds of stem cells have been studied; these have been derived from embryo or adult tissues (i.e. bone marrow, heart, peripheral blood etc.). Currently, three different biologic therapies for cardiovascular diseases are under investigation: cell therapy, gene therapy and the more recent “tissue-engineering” therapy . During my Ph.D. course, first I focalised my study on the isolation and characterization of Cardiac Stem Cells (CSCs) in wild-type and transgenic mice and for this purpose I attended, for more than one year, the Cardiovascular Research Institute of the New York Medical College, in Valhalla (NY, USA) under the direction of Doctor Piero Anversa. During this period I learnt different Immunohistochemical and Biomolecular techniques, useful for investigating the regenerative potential of stem cells. Then, during the next two years, I studied the new approach of cardiac regenerative medicine based on “tissue-engineering” in order to investigate a new strategy to regenerate the infracted myocardium. Tissue-engineering is a promising approach that makes possible the creation of new functional tissue to replace lost or failing tissue. This new discipline combines isolated functioning cells and biodegradable 3-dimensional (3D) polymeric scaffolds. The scaffold temporarily provides the biomechanical support for the cells until they produce their own extracellular matrix. Because tissue-engineering constructs contain living cells, they may have the potential for growth and cellular self-repair and remodeling. In the present study, I examined whether the tissue-engineering strategy within hyaluron-based scaffolds would result in the formation of alternative cardiac tissue that could replace the scar and improve cardiac function after MI in syngeneic heterotopic rat hearts. Rat hearts were explanted, subjected to left coronary descending artery occlusion, and then grafted into the abdomen (aorta-aorta anastomosis) of receiving syngeneic rat. After 2 weeks, a pouch of 3 mm2 was made in the thickness of the ventricular wall at the level of the post-infarction scar. The hyaluronic scaffold, previously engineered for 3 weeks with rat MSCs, was introduced into the pouch and the myocardial edges sutured with few stitches. Two weeks later we evaluated the cardiac function by M-Mode echocardiography and the myocardial morphology by microscope analysis. We chose bone marrow-derived mensenchymal stem cells (MSCs) because they have shown great signaling and regenerative properties when delivered to heart tissue following a myocardial infarction (MI). However, while the object of cell transplantation is to improve ventricular function, cardiac cell transplantation has had limited success because of poor graft viability and low cell retention, that’s why we decided to combine MSCs with a biopolimeric scaffold. At the end of the experiments we observed that the hyaluronan fibres had not been substantially degraded 2 weeks after heart-transplantation. Most MSCs had migrated to the surrounding infarcted area where they were especially found close to small-sized vessels. Scar tissue was moderated in the engrafted region and the thickness of the corresponding ventricular wall was comparable to that of the non-infarcted remote area. Also, the left ventricular shortening fraction, evaluated by M-Mode echocardiography, was found a little bit increased when compared to that measured just before construct transplantation. Therefore, this study suggests that post-infarction myocardial remodelling can be favourably affected by the grafting of MSCs delivered through a hyaluron-based scaffold

Statistical methods for analysis and processing of medical ultrasound: applications to segmentation and restoration

In this thesis two major topics inherent with medical ultrasound images are addressed: deconvolution and segmentation. In the first case a deconvolution algorithm is described allowing statistically consistent maximum a posteriori estimates of the tissue reflectivity to be restored. These estimates are proven to provide a reliable source of information for achieving an accurate characterization of biological tissues through the ultrasound echo. The second topic involves the definition of a semi automatic algorithm for myocardium segmentation in 2D echocardiographic images. The results show that the proposed method can reduce inter- and intra observer variability in myocardial contours delineation and is feasible and accurate even on clinical data.

Automated Segmentation and Non-Rigid Registration for the Quantitative Evaluation of Myocardial Perfusion from Magnetic Resonance Images

Myocardial perfusion quantification by means of Contrast-Enhanced Cardiac Magnetic Resonance images relies on time consuming frame-by-frame manual tracing of regions of interest. In this Thesis, a novel automated technique for myocardial segmentation and non-rigid registration as a basis for perfusion quantification is presented. The proposed technique is based on three steps: reference frame selection, myocardial segmentation and non-rigid registration. In the first step, the reference frame in which both endo- and epicardial segmentation will be performed is chosen. Endocardial segmentation is achieved by means of a statistical region-based level-set technique followed by a curvature-based regularization motion. Epicardial segmentation is achieved by means of an edge-based level-set technique followed again by a regularization motion. To take into account the changes in position, size and shape of myocardium throughout the sequence due to out of plane respiratory motion, a non-rigid registration algorithm is required. The proposed non-rigid registration scheme consists in a novel multiscale extension of the normalized cross-correlation algorithm in combination with level-set methods. The myocardium is then divided into standard segments. Contrast enhancement curves are computed measuring the mean pixel intensity of each segment over time, and perfusion indices are extracted from each curve. The overall approach has been tested on synthetic and real datasets. For validation purposes, the sequences have been manually traced by an experienced interpreter, and contrast enhancement curves as well as perfusion indices have been computed. Comparisons between automatically extracted and manually obtained contours and enhancement curves showed high inter-technique agreement. Comparisons of perfusion indices computed using both approaches against quantitative coronary angiography and visual interpretation demonstrated that the two technique have similar diagnostic accuracy. In conclusion, the proposed technique allows fast, automated and accurate measurement of intra-myocardial contrast dynamics, and may thus address the strong clinical need for quantitative evaluation of myocardial perfusion.

The effects of hypocalcemia on spatial alternans and ventricular fibrillation studied with the optical mapping technique

The heart is a wonderful but complex organ: it uses electrochemical mechanisms in order to produce mechanical energy to pump the blood throughout the body and allow the life of humans and animals. This organ can be subject to several diseases and sudden cardiac death (SCD) is the most catastrophic manifestation of these diseases, responsible for the death of a large number of people throughout the world. It is estimated that 325000 Americans annually die for SCD. SCD most commonly occurs as a result of reentrant tachyarrhythmias (ventricular tachycardia (VT) and ventricular fibrillation (VF)) and the identification of those patients at higher risk for the development of SCD has been a difficult clinical challenge. Nowadays, a particular electrocardiogram (ECG) abnormality, “T-wave alternans” (TWA), is considered a precursor of lethal cardiac arrhythmias and sudden death, a sensitive indicator of risk for SCD. TWA is defined as a beat-to-beat alternation in the shape, amplitude, or timing of the T-wave on the ECG, indicative of the underlying repolarization of cardiac cells [5]. In other words TWA is the macroscopic effect of subcellular and celluar mechanisms involving ionic kinetics and the consequent depolarization and repolarization of the myocytes. Experimental activities have shown that TWA on the ECG is a manifestation of an underlying alternation of long and short action potential durations (APDs), the so called APD-alternans, of cardiac myocytes in the myocardium. Understanding the mechanism of APDs-alternans is the first step for preventing them to occur. In order to investigate these mechanisms it’s very important to understand that the biological systems are complex systems and their macroscopic properties arise from the nonlinear interactions among the parts. The whole is greater than the sum of the parts, and it cannot be understood only by studying the single parts. In this sense the heart is a complex nonlinear system and its way of working follows nonlinear dynamics; alternans also, they are a manifestation of a phenomenon typical in nonlinear dynamical systems, called “period-dubling bifurcation”. Over the past decade, it has been demonstrated that electrical alternans in cardiac tissue is an important marker for the development of ventricular fibrillation and a significant predictor for mortality. It has been observed that acute exposure to low concentration of calcium does not decrease the magnitude of alternans and sustained ventricular Fibrillation (VF) is still easily induced under these condition. However with prolonged exposure to low concentration of calcium, alternans disappears, but VF is still inducible. This work is based on this observation and tries to make it clearer. The aim of this thesis is investigate the effect of hypocalcemia spatial alternans and VF doing experiments with canine hearts and perfusing them with a solution with physiological ionic concentration and with a solution with low calcium concentration (hypocalcemia); in order to investigate the so called memory effect, the experimental activity was modified during the way. The experiments were performed with the optical mapping technique, using voltage-sensitive dye, and a custom made Java code was used in post-processing. Finding the Nolasco and Dahlen’s criterion [8] inadequate for the prediction of alternans, and takin into account the experimental results, another criterion, which consider the memory effect, has been implemented. The implementation of this criterion could be the first step in the creation of a method, AP-based, discriminating who is at risk if developing VF. This work is divided into four chapters: the first is a brief presentation of the physiology of the heart; the second is a review of the major theories and discovers in the study of cardiac dynamics; the third chapter presents an overview on the experimental activity and the optical mapping technique; the forth chapter contains the presentation of the results and the conclusions.

Efficient ultrasonic signal processing techniques for aided medical diagnostics

Ultrasound imaging is widely used in medical diagnostics as it is the fastest, least invasive, and least expensive imaging modality. However, ultrasound images are intrinsically difficult to be interpreted. In this scenario, Computer Aided Detection (CAD) systems can be used to support physicians during diagnosis providing them a second opinion. This thesis discusses efficient ultrasound processing techniques for computer aided medical diagnostics, focusing on two major topics: (i) Ultrasound Tissue Characterization (UTC), aimed at characterizing and differentiating between healthy and diseased tissue; (ii) Ultrasound Image Segmentation (UIS), aimed at detecting the boundaries of anatomical structures to automatically measure organ dimensions and compute clinically relevant functional indices. Research on UTC produced a CAD tool for Prostate Cancer detection to improve the biopsy protocol. In particular, this thesis contributes with: (i) the development of a robust classification system; (ii) the exploitation of parallel computing on GPU for real-time performance; (iii) the introduction of both an innovative Semi-Supervised Learning algorithm and a novel supervised/semi-supervised learning scheme for CAD system training that improve system performance reducing data collection effort and avoiding collected data wasting. The tool provides physicians a risk map highlighting suspect tissue areas, allowing them to perform a lesion-directed biopsy. Clinical validation demonstrated the system validity as a diagnostic support tool and its effectiveness at reducing the number of biopsy cores requested for an accurate diagnosis. For UIS the research developed a heart disease diagnostic tool based on Real-Time 3D Echocardiography. Thesis contributions to this application are: (i) the development of an automated GPU based level-set segmentation framework for 3D images; (ii) the application of this framework to the myocardium segmentation. Experimental results showed the high efficiency and flexibility of the proposed framework. Its effectiveness as a tool for quantitative analysis of 3D cardiac morphology and function was demonstrated through clinical validation.

Sistemi endorfinergici e modulazione di segnali molecolari e profili trascrizionali coinvolti in processi di protezione e autoriparazione del miocardio danneggiato

Con il termine IPC (precondizionamento ischemico) si indica un fenomeno per il quale, esponendo il cuore a brevi cicli di ischemie subletali prima di un danno ischemico prolungato, si conferisce una profonda resistenza all’infarto, una delle principali cause di invalidità e mortalità a livello mondiale. Studi recenti hanno suggerito che l’IPC sia in grado di migliorare la sopravvivenza, la mobilizzazione e l’integrazione di cellule staminali in aree ischemiche e che possa fornire una nuova strategia per potenziare l’efficacia della terapia cellulare cardiaca, un’area della ricerca in continuo sviluppo. L’IPC è difficilmente trasferibile nella pratica clinica ma, da anni, è ben documentato che gli oppioidi e i loro recettori hanno un ruolo cardioprotettivo e che attivano le vie di segnale coinvolte nell’IPC: sono quindi candidati ideali per una possibile terapia farmacologica alternativa all’IPC. Il trattamento di cardiomiociti con gli agonisti dei recettori oppioidi Dinorfina B, DADLE e Met-Encefalina potrebbe proteggere, quindi, le cellule dall’apoptosi causata da un ambiente ischemico ma potrebbe anche indurle a produrre fattori che richiamino elementi staminali. Per testare quest’ipotesi è stato messo a punto un modello di “microambiente ischemico” in vitro sui cardiomioblasti di ratto H9c2 ed è stato dimostrato che precondizionando le cellule in modo “continuativo” (ventiquattro ore di precondizionamento con oppioidi e successivamente ventiquattro ore di induzione del danno, continuando a somministrare i peptidi oppioidi) con Dinorfina B e DADLE si verifica una protezione diretta dall’apoptosi. Successivamente, saggi di migrazione e adesione hanno mostrato che DADLE agisce sulle H9c2 “ischemiche” spronandole a creare un microambiente capace di attirare cellule staminali mesenchimali umane (FMhMSC) e di potenziare le capacità adesive delle FMhMSC. I dati ottenuti suggeriscono, inoltre, che la capacità del microambiente ischemico trattato con DADLE di attirare le cellule staminali possa essere imputabile alla maggiore espressione di chemochine da parte delle H9c2.