Allogeneic Mesenchymal Stromal Cells Overexpressing Mutant Human Hypoxia-Inducible Factor 1-α (HIF1-α) in an Ovine Model of Acute Myocardial Infarction.

BACKGROUND: Bone marrow mesenchymal stromal cells (BMMSCs) are cardioprotective in acute myocardial infarction (AMI) because of release of paracrine angiogenic and prosurvival factors. Hypoxia-inducible factor 1-α (HIF1-α), rapidly degraded during normoxia, is stabilized during ischemia and upregulates various cardioprotective genes. We hypothesized that BMMSCs engineered to overexpress mutant, oxygen-resistant HIF1-α would confer greater cardioprotection than nontransfected BMMSCs in sheep with AMI. METHODS AND RESULTS: Allogeneic BMMSCs transfected with a minicircle vector encoding mutant HIF1-α (BMMSC-HIF) were injected in the peri-infarct of sheep (n=6) undergoing coronary occlusion. Over 2 months, infarct volume measured by cardiac magnetic resonance (CMR) imaging decreased by 71.7±1.3% (P<0.001), and left ventricular (LV) percent ejection fraction (%EF) increased near 2-fold (P<0.001) in the presence of markedly decreased end-systolic volume. Sheep receiving nontransfected BMMSCs (BMMSC; n=6) displayed less infarct size limitation and percent LVEF improvement, whereas in placebo-treated animals (n=6), neither parameters changed over time. HIF1-α-transfected BMMSCs (BMMSC-HIF) induced angio-/arteriogenesis and decreased apoptosis by HIF1-mediated overexpression of erythropoietin, inducible nitrous oxide synthase, vascular endothelial growth factor, and angiopoietin-1. Cell tracking using paramagnetic iron nanoparticles in 12 additional sheep revealed enhanced long-term retention of BMMSC-HIF. CONCLUSIONS: Intramyocardial delivery of BMMSC-HIF reduced infarct size and improved LV systolic performance compared to BMMSC, attributed to increased neovascularization and cardioprotective effects induced by HIF1-mediated overexpression of paracrine factors and enhanced retention of injected cells. Given the safety of the minicircle vector and the feasibility of BMMSCs for allogeneic application, this treatment may be potentially useful in the clinic.

Activated macrophages as a feeder layer for growth of resident cardiac progenitor cells.

The adult heart contains a population of cardiac progenitor cells (CPCs). Growing and collecting an adequate number of CPCs demands complex culture media containing growth factors. Since activated macrophages secrete many growth factors, we investigated if activated isolated heart cells seeded on a feeder layer of activated peritoneal macrophages (PM) could result in CPCs and if these, in turn, could exert cardioprotection in rats with myocardial infarction (MI). Heart cells of inbred Wistar rats were isolated by collagenase digestion and cultured on PM obtained 72 h after intraperitoneal injection of 12 ml thioglycollate. Cells (1 × 10(6)) exhibiting CPC phenotype (immunohistochemistry) were injected in the periphery of rat MI 10 min after coronary artery occlusion. Control rats received vehicle. Three weeks later, left ventricular (LV) function (echocardiogram) was assessed, animals were euthanized and the hearts removed for histological studies. Five to six days after seeding heart cells on PM, spherical clusters composed of small bright and spherical cells expressing mostly c-Kit and Sca-1 antigens were apparent. After explant, those clusters developed cobblestone-like monolayers that expressed smooth muscle actin and sarcomeric actin and were successfully transferred for more than ten passages. When injected in the MI periphery, many of them survived at 21 days after coronary ligature, improved LV ejection fraction and decreased scar size as compared with control rats. CPC-derived cells with cardiocyte and smooth muscle phenotypes can be successfully grown on a feeder layer of activated syngeneic PM. These cells decreased scar size and improved heart function in rats with MI.

Vascular endothelial growth factor overexpression does not enhance adipose stromal cell-induced protection on muscle damage in critical limb ischemia.

OBJECTIVES: Critical limb ischemia complicates peripheral artery disease leading to tissue damage and amputation. We hypothesized that modifying adipose stromal cells (ASCs) to overexpress human vascular endothelial growth factor 165 (VEGF) would limit ischemic muscle damage to a larger extent than nonmodified ASCs. APPROACH AND RESULTS: Rabbits with critical hindlimb ischemia were injected with allogeneic abdominal fat-derived ASCs transfected with plasmid-VEGF165 (ASCs-VEGF; n=10). Additional rabbits received nontransfected ASCs (ASCs; n=10) or vehicle (placebo; n=10). One month later, ASCs-VEGF rabbits exhibited significantly higher density of angiographically visible collaterals and capillaries versus placebo (both P<0.05) but not versus ASCs (both P=NS). Arteriolar density, however, was increased in both ASCs and ASCs-VEGF groups (both P<0.05 versus placebo). ASCs-VEGF and ASCs showed comparable post-treatment improvements in Doppler-assessed peak systolic velocity, blood pressure ratio, and resistance index. Ischemic lesions were found in 40% of the muscle samples in the placebo group, 19% in the ASCs-VEGF group, and 17% in the ASCs groups (both P<0.05 versus placebo, Fisher test). CONCLUSIONS: In a rabbit model of critical limb ischemia, intramuscular injection of ASCs genetically modified to overexpress VEGF increase angiographically visible collaterals and capillary density. However, both modified and nonmodified ASCs increase arteriolar density to a similar extent and afford equal protection against ischemia-induced muscle lesions. These results indicate that modifying ASCs to overexpress VEGF does not enhance the protective effect of ASCs, and that arteriolar proliferation plays a pivotal role in limiting the irreversible tissue damage of critical limb ischemia.

Presence of antigen-experienced T cells with low grade of differentiation and proliferative potential in chronic Chagas disease myocarditis.

BACKGROUND: The main consequence of chronic Trypanosoma cruzi infection is the development of myocarditis in approximately 20-30% of infected individuals but not until 10-20 years after the initial infection. We have previously shown that circulating interferon-γ-secreting T cells responsive to Trypanosoma cruzi antigens in chronic Chagas disease patients display a low grade of differentiation and the frequency of these T lymphocytes decreases along with the severity of heart disease. This study thought to explore the expression of inhibitory receptors, transcription factors of type 1 or regulatory T cells, and markers of T cell differentiation, immunosenescence or active cell cycle in cardiac explants from patients with advanced Chagas disease myocarditis. METHODOLOGY/PRINCIPAL FINDINGS: The expression of different markers for T and B cells as well as for macrophages was evaluated by immunohistochemistry and immunofluorescence techniques in cardiac explants from patients with advanced chronic Chagas disease submitted to heart transplantation. Most infiltrating cells displayed markers of antigen-experienced T cells (CD3(+), CD4(+), CD8(+), CD45RO(+)) with a low grade of differentiation (CD27(+), CD57(-), CD45RA(-), PD(-)1(-)). A skewed T helper1/T cytotoxic 1 profile was supported by the expression of T-bet; whereas FOXP3(+) cells were scarce and located only in areas of severe myocarditis. In addition, a significant proliferative capacity of CD3(+) T cells, assessed by Ki67 staining, was found. CONCLUSIONS/SIGNIFICANCE: The quality of T cell responses and immunoregulatory mechanisms might determine the pattern of the cellular response and the severity of disease in chronic Trypanosoma cruzi infection.

Severe mitral regurgitation after radiotherapy.

We present the case of a 69-year-old patient with a history of gynecological neoplasia and a pulmonary metastasis, who in 1996 underwent chemotherapy and mediastinal radiotherapy followed by cancer remission. Ten years later she presented with heart failure and her Doppler echocardiogram showed severe mitral regurgitation with pulmonary hypertension. In 2011, she underwent a mitral valve replacement with a biological prosthesis and the pathology exam revealed valve damage consistent with radiotherapy-induced changes. This unusual mechanism of mitral regurgitation can be demonstrated clearly by echocardiography and should be disseminated among cardiology physicians and in patients who have survived for long periods after radiotherapy, it is important to remember that cardiac complications may indeed occur, and the treating physician is responsible for detecting them.

Predictores de Mortalidad o trasplante cardíaco en la miocardiopatía periparto / Predictors of Mortality or Heart Transplantation in Peripartum Cardiomyopathy

Introducción La miocardiopatía periparto es una forma infrecuente de insuficiencia cardíaca congestiva con una evolución impredecible. Su verdadera incidencia y prevalencia no se conoce con certeza y su etiología aún no se ha aclarado, aunque se han involucrado varios mecanismos en los que se reconocen diversos factores de riesgo. Objetivo Analizar predictores pronósticos de mortalidad o de requerimiento de trasplante cardíaco. Material y métodos Entre 1992 y noviembre de 2011 se evaluaron retrospectivamente 23 pacientes. En aquellas con insuficiencia cardíaca descompensada se realizó monitorización hemodinámica. La mediana de seguimiento fue de 7,3 años (3,2-17,5). El análisis univariado se realizó por regresión de Cox y la supervivencia global se calculó con el método de Kaplan-Meier. Resultados La edad media fue de 28,7 ± 8,8 años, ocho pacientes eran multíparas. El 73% estaban en clase funcional III-IV. La presión arterial sistólica y diastólica fue de 103 ± 23 y 67 ± 11 mm Hg, respectivamente, y la frecuencia cardíaca, de 92 ± 19 lpm. El 100% se encontraba en ritmo sinusal. El índice cardiotorácico fue de 0,56 ± 0,07. El diámetro diastólico y sistólico del ventrículo izquierdo fue de 67,5 ± 10,2 y 56,7 ± 10,1 mm, respectivamente, el diámetro auricular izquierdo fue de 42,5 ± 6 mm y la fracción de eyección del ventrículo izquierdo, del 24,6% ± 10,8%. La presión media de la arteria pulmonar fue de 25 ± 9 mm Hg y la capilar pulmonar, de 18,4 ± 7,8 mm Hg; el índice cardíaco fue de 2,6 ± 0,6 L/min/m². Siete pacientes fallecieron y tres fueron sometidas a trasplante cardíaco. En el análisis univariado, la clase funcional, el índice cardíaco, la presión arterial sistólica y diastólica, la presión capilar y pulmonar media, el índice cardiotorácico y el diámetro auricular izquierdo se asociaron con mortalidad y trasplante cardíaco. La supervivencia a 1, 3 y 6 años fue del 91%, 82% y 64%, respectivamente. Conclusiones La mortalidad hospitalaria fue del 4,3% y el requerimiento de trasplante cardíaco o la muerte en el seguimiento fueron del 39%. Los parámetros hemodinámicos al ingreso fueron los principales predictores de mortalidad y de trasplante.

Background Peripartum cardiomyopathy is an uncommon form of congestive heart failure with an unpredictable outcome. Very little is known about its real incidence and prevalence, and its etiology is still unknown, although a number of contributing factors, including diverse risk factors, have been proposed. Objective To analyze the predictors of mortality or need for heart transplantation. Methods Between 1992 and November 2011, 23 patients were retrospectively evaluated. Patients with decompensated heart failure were managed with hemodynamic monitoring. Me­dian follow-up was of 7.3 years (3.2-17.5). Univariate Cox regression analysis was performed and overall survival was calculated using the Kaplan-Meier method. Results Mean age was 28.7±8.8 years; eight patients were multipara. Seventy three percent were in functional class III-IV. Systolic blood pressure and diastolic blood pressure were 103±23 and 67±11 mm Hg, respectively, and heart rate was 92±19 bpm. All the patients were in sinus rhythm. The cardiothoracic index was 0.56±0.07. End-diastolic and end-systolic left ventricular dimensions were 67.5±10.2 and 56.7±10.1 mm, respectively; left atrial dimension was 42.5±6 mm and left ventricular ejection fraction was 24.6%±10.8%. Mean pulmonary artery pressure was 25±9 mm Hg and pulmonary wedge pressure, was 18.4±7.8 mm Hg; cardiac index was 2.6±0.6 L/min/m². Seven patients died and three patients underwent heart transplantation. Univariate analysis revealed that functional class, cardiac index, systolic and diastolic blood pressure, pulmonary wedge pressure and mean pulmonary artery pressure, cardiothoracic index and left atrial dimension were associated with mortality and heart transplantation. Survival at one, three and six years was of 91%, 82% y 64%, respectively. Conclusions In-hospital mortality was of 4.3% and the need for heart transplantation or mortality during follow-up was of 39%. The hemodynamic parameters at admission were the main predictors of mortality and transplant.

Predictores de Mortalidad o trasplante cardíaco en la miocardiopatía periparto / Predictors of Mortality or Heart Transplantation in Peripartum Cardiomyopathy

Introducción La miocardiopatía periparto es una forma infrecuente de insuficiencia cardíaca congestiva con una evolución impredecible. Su verdadera incidencia y prevalencia no se conoce con certeza y su etiología aún no se ha aclarado, aunque se han involucrado varios mecanismos en los que se reconocen diversos factores de riesgo. Objetivo Analizar predictores pronósticos de mortalidad o de requerimiento de trasplante cardíaco. Material y métodos Entre 1992 y noviembre de 2011 se evaluaron retrospectivamente 23 pacientes. En aquellas con insuficiencia cardíaca descompensada se realizó monitorización hemodinámica. La mediana de seguimiento fue de 7,3 años (3,2-17,5). El análisis univariado se realizó por regresión de Cox y la supervivencia global se calculó con el método de Kaplan-Meier. Resultados La edad media fue de 28,7 ± 8,8 años, ocho pacientes eran multíparas. El 73% estaban en clase funcional III-IV. La presión arterial sistólica y diastólica fue de 103 ± 23 y 67 ± 11 mm Hg, respectivamente, y la frecuencia cardíaca, de 92 ± 19 lpm. El 100% se encontraba en ritmo sinusal. El índice cardiotorácico fue de 0,56 ± 0,07. El diámetro diastólico y sistólico del ventrículo izquierdo fue de 67,5 ± 10,2 y 56,7 ± 10,1 mm, respectivamente, el diámetro auricular izquierdo fue de 42,5 ± 6 mm y la fracción de eyección del ventrículo izquierdo, del 24,6% ± 10,8%. La presión media de la arteria pulmonar fue de 25 ± 9 mm Hg y la capilar pulmonar, de 18,4 ± 7,8 mm Hg; el índice cardíaco fue de 2,6 ± 0,6 L/min/m². Siete pacientes fallecieron y tres fueron sometidas a trasplante cardíaco. En el análisis univariado, la clase funcional, el índice cardíaco, la presión arterial sistólica y diastólica, la presión capilar y pulmonar media, el índice cardiotorácico y el diámetro auricular izquierdo se asociaron con mortalidad y trasplante cardíaco. La supervivencia a 1, 3 y 6 años fue del 91%, 82% y 64%, respectivamente. Conclusiones La mortalidad hospitalaria fue del 4,3% y el requerimiento de trasplante cardíaco o la muerte en el seguimiento fueron del 39%. Los parámetros hemodinámicos al ingreso fueron los principales predictores de mortalidad y de trasplante.(AU)

Background Peripartum cardiomyopathy is an uncommon form of congestive heart failure with an unpredictable outcome. Very little is known about its real incidence and prevalence, and its etiology is still unknown, although a number of contributing factors, including diverse risk factors, have been proposed. Objective To analyze the predictors of mortality or need for heart transplantation. Methods Between 1992 and November 2011, 23 patients were retrospectively evaluated. Patients with decompensated heart failure were managed with hemodynamic monitoring. Me¡dian follow-up was of 7.3 years (3.2-17.5). Univariate Cox regression analysis was performed and overall survival was calculated using the Kaplan-Meier method. Results Mean age was 28.7±8.8 years; eight patients were multipara. Seventy three percent were in functional class III-IV. Systolic blood pressure and diastolic blood pressure were 103±23 and 67±11 mm Hg, respectively, and heart rate was 92±19 bpm. All the patients were in sinus rhythm. The cardiothoracic index was 0.56±0.07. End-diastolic and end-systolic left ventricular dimensions were 67.5±10.2 and 56.7±10.1 mm, respectively; left atrial dimension was 42.5±6 mm and left ventricular ejection fraction was 24.6%±10.8%. Mean pulmonary artery pressure was 25±9 mm Hg and pulmonary wedge pressure, was 18.4±7.8 mm Hg; cardiac index was 2.6±0.6 L/min/m². Seven patients died and three patients underwent heart transplantation. Univariate analysis revealed that functional class, cardiac index, systolic and diastolic blood pressure, pulmonary wedge pressure and mean pulmonary artery pressure, cardiothoracic index and left atrial dimension were associated with mortality and heart transplantation. Survival at one, three and six years was of 91%, 82% y 64%, respectively. Conclusions In-hospital mortality was of 4.3% and the need for heart transplantation or mortality during follow-up was of 39%. The hemodynamic parameters at admission were the main predictors of mortality and transplant.(AU)

Combined VEGF gene transfer and erythropoietin in ovine reperfused myocardial infarction.

BACKGROUND: In reperfused acute myocardial infarction (RAMI), cardioprotective treatments may enhance myocardial salvage and hence reduce the area of necrosis. Based on studies showing that plasmid-mediated vascular endothelial growth factor (pVEGF) gene transfer reduces infarct size by combining angio-arteriogenic and cardiomyogenic effects and that erythropoietin (EPO) exerts anti-apoptotic actions in animal models of AMI, we aimed to assess if their association would reduce infarct size to a larger extent than any of them individually in a large mammalian model of RAMI. METHODS: Adult sheep subjected to 90-minute coronary artery occlusion received upon reperfusion intramyocardial pVEGF 3.8 mg plus intravenous EPO 1000 IU/kg (n=8), pVEGF (n=8), EPO (n=8) or placebo (n=8). RESULTS: Fifteen days after treatment, infarct size was smaller in the 3 treatment groups (pVEGF+EPO: 8 ± 1 %; pVEGF: 16 ± 5 %; EPO: 13 ± 4 %) compared to placebo (25 ± 7 %, p<0.001). However, in the EPO+VEGF group infarct size was significantly smaller than in the groups receiving EPO or VEGF individually (p<0.05). DNA fragmentation, a hallmark of late apoptosis, was significantly lower in sheep receiving EPO. The combined treatment, while not affecting global left ventricular performance, improved regional peri-infarct function and prevented over-time expansion of the post-infarct perfusion defect. CONCLUSIONS: Combined pVEGF and EPO treatment might be clinically useful to enhance the benefits of early revascularization in patients with acute myocardial infarction.

Presencia de angiogénesis en placas vulnerables ateroscleróticas en corazones humanos aparentemente sanos / Angiogenesis in Vulnerable Atherosclerotic Plaques in Apparently Healthy Human Hearts / Angiogenesis in Vulnerable Atherosclerotic Plaques in Apparently Healthy Human Hearts

Introducción La angiogénesis o neovascularización involucra la formación de nuevos conductos en las adyacencias de vasos preexistentes. Esta proliferación vascular es frecuente en varias circunstancias clínicas, como es el caso de la aterosclerosis. Los microvasos de las placas ateroscleróticas coronarias pueden estar vinculados a la inestabilidad de la lesión. Objetivo Correlacionar la presencia de angiogénesis en placas ateroscleróticas con los criterios de vulnerabilidad de la clasificación de la American Heart Association (AHA). Material y métodos En 121 corazones de donantes no diabéticos aparentemente sanos y mayores de 40 años destinados para homoinjertos se examinaron las arterias coronarias y todas las áreas de estrechamiento luminal se sometieron a estudios histológicos, inmunohistoquímicos y morfométricos. Para el análisis de la angiogénesis se empleó un puntaje semicuantitativo (escala 0-3). Se realizó un análisis de regresión logística univariado y multivariado para identificar factores de riesgo relacionados con la angiogénesis. Resultados Se hallaron 143 lesiones de riesgo alto (AHA tipos IV, V y VI) en las arterias descendente anterior (46,3%), circunfleja (28,9%) y coronaria derecha (43%). La angiogénesis se asoció en forma estadísticamente significativa con el grado de oclusión vascular, la infiltración de células inflamatorias, la presencia de centro lipídico, la fibrosis, la periarteritis y, sólo en la descendente anterior, con el antecedente de hipertensión arterial (p < 0,006). Se detectó angiogénesis en 1 placa tipo II, en 5 tipo III, en 21 tipo IV, en 22 tipo V y en 7 placas tipo VI (AHA). Conclusiones La angiogénesis de placas vulnerables se asoció con el grado de oclusión vascular, la infiltración de células inflamatorias, la fibrosis, la presencia de núcleo lipídico y, sólo en la descendente anterior, con el antecedente de hipertensión arterial. No se encontró asociación con la hemorragia intraplaca o la calcificación, lo cual sugiere que la angiogénesis puede anticipar la rotura de las placas.

Background Angiogenesis or neovascularization involves the formation of new blood vessels adjacent to preexisting vessels. This vascular proliferation is prevalent in various clinical conditions, such as atherosclerosis. Microvessels in coronary artery atherosclerotic plaques may contribute to plaque instability. Objectives The aim of this study was to correlate the presence of angiogenesis in atherosclerotic plaques with the criteria of plaque vulnerability used by the American Heart Association (AHA). Methods One hundred and twenty one hearts from non-diabetic and apparently healthy transplant donors older than 40 years were selected. The coronary arteries were examined and all areas of cross-sectional luminal narrowing underwent histological, immunohistochemical and morphometric studies. A semi-quantitative score (scale 0-3) was used to identify of angiogenesis. Univariate and multivariate logistic regression analysis was performed to identify angiogenesisrelated risk factors. Results On hundred and forty three high-risk lesions (AHA type IV, V and VI) in the left anterior descending coronary artery (46.3%), the circumflex coronary artery (28.9%) and the right coronary artery (43%) were identified. Angiogenesis had a statistically significant association with the severity of vascular occlusion, inflammatory cell infiltration, presence of a lipid core, fibrosis and periarteritis. A history of hypertension (HT) was associated with angiogenesis only in lesions of the left anterior descending coronary artery (LAD). According to the AHA classification angiogenesis was detected in 1 Type II, 5 Type III, 21 Type IV, 22 Type V, and 7 Type VI plaques. Conclusions Angiogenesis in vulnerable plaques was associated with the severity of vascular occlusion, inflammatory cell infiltration, fibrosis and presence of a lipid core, and with a history of HT in LAD lesions. There was no association between angiogenesis and plaque hemorrhage or calcification, suggesting that angiogenesis may anticipate plaque rupture.

Presencia de angiogénesis en placas vulnerables ateroscleróticas en corazones humanos aparentemente sanos / Angiogenesis in Vulnerable Atherosclerotic Plaques in Apparently Healthy Human Hearts / Angiogenesis in Vulnerable Atherosclerotic Plaques in Apparently Healthy Human Hearts

Introducción La angiogénesis o neovascularización involucra la formación de nuevos conductos en las adyacencias de vasos preexistentes. Esta proliferación vascular es frecuente en varias circunstancias clínicas, como es el caso de la aterosclerosis. Los microvasos de las placas ateroscleróticas coronarias pueden estar vinculados a la inestabilidad de la lesión. Objetivo Correlacionar la presencia de angiogénesis en placas ateroscleróticas con los criterios de vulnerabilidad de la clasificación de la American Heart Association (AHA). Material y métodos En 121 corazones de donantes no diabéticos aparentemente sanos y mayores de 40 años destinados para homoinjertos se examinaron las arterias coronarias y todas las áreas de estrechamiento luminal se sometieron a estudios histológicos, inmunohistoquímicos y morfométricos. Para el análisis de la angiogénesis se empleó un puntaje semicuantitativo (escala 0-3). Se realizó un análisis de regresión logística univariado y multivariado para identificar factores de riesgo relacionados con la angiogénesis. Resultados Se hallaron 143 lesiones de riesgo alto (AHA tipos IV, V y VI) en las arterias descendente anterior (46,3%), circunfleja (28,9%) y coronaria derecha (43%). La angiogénesis se asoció en forma estadísticamente significativa con el grado de oclusión vascular, la infiltración de células inflamatorias, la presencia de centro lipídico, la fibrosis, la periarteritis y, sólo en la descendente anterior, con el antecedente de hipertensión arterial (p < 0,006). Se detectó angiogénesis en 1 placa tipo II, en 5 tipo III, en 21 tipo IV, en 22 tipo V y en 7 placas tipo VI (AHA). Conclusiones La angiogénesis de placas vulnerables se asoció con el grado de oclusión vascular, la infiltración de células inflamatorias, la fibrosis, la presencia de núcleo lipídico y, sólo en la descendente anterior, con el antecedente de hipertensión arterial. No se encontró asociación con la hemorragia intraplaca o la calcificación, lo cual sugiere que la angiogénesis puede anticipar la rotura de las placas.(AU)

Background Angiogenesis or neovascularization involves the formation of new blood vessels adjacent to preexisting vessels. This vascular proliferation is prevalent in various clinical conditions, such as atherosclerosis. Microvessels in coronary artery atherosclerotic plaques may contribute to plaque instability. Objectives The aim of this study was to correlate the presence of angiogenesis in atherosclerotic plaques with the criteria of plaque vulnerability used by the American Heart Association (AHA). Methods One hundred and twenty one hearts from non-diabetic and apparently healthy transplant donors older than 40 years were selected. The coronary arteries were examined and all areas of cross-sectional luminal narrowing underwent histological, immunohistochemical and morphometric studies. A semi-quantitative score (scale 0-3) was used to identify of angiogenesis. Univariate and multivariate logistic regression analysis was performed to identify angiogenesisrelated risk factors. Results On hundred and forty three high-risk lesions (AHA type IV, V and VI) in the left anterior descending coronary artery (46.3%), the circumflex coronary artery (28.9%) and the right coronary artery (43%) were identified. Angiogenesis had a statistically significant association with the severity of vascular occlusion, inflammatory cell infiltration, presence of a lipid core, fibrosis and periarteritis. A history of hypertension (HT) was associated with angiogenesis only in lesions of the left anterior descending coronary artery (LAD). According to the AHA classification angiogenesis was detected in 1 Type II, 5 Type III, 21 Type IV, 22 Type V, and 7 Type VI plaques. Conclusions Angiogenesis in vulnerable plaques was associated with the severity of vascular occlusion, inflammatory cell infiltration, fibrosis and presence of a lipid core, and with a history of HT in LAD lesions. There was no association between angiogenesis and plaque hemorrhage or calcification, suggesting that angiogenesis may anticipate plaque rupture.(AU)

Successful circumferential free tracheal transplantation in a large animal model.

BACKGROUND: Long segment tracheobronchial stenoses are associated with high morbi-mortality rates and difficult treatment. Transplantation hasn't proved to be useful yet. Currently, the successful results achieved in small animal models couldn't be satisfactorily accomplished or extrapolated in large mammals. We aimed to evaluate the viability of orthotopic tracheal autoimplantation in an ovine model. METHODS: All animals underwent tracheal transplantation of 4 cm (5-7 rings) of the cervical trachea and were divided randomly in two groups: isolated autoimplantation (Group A/6) and autoimplantation with omental wrapping (Group B/6). Clinical follow up and weekly bronchoscopical examinations were performed. The grafts were macroscopically, histologically, and bacteriologically analyzed. RESULTS: In group A, four animals achieved their planed survival and were sacrificed up to 60 days after transplantation with viable grafts. In group B, only two sheep had successful results. Graft failure with infection, necrosis and severe stenosis was observed in the rest of the animals from both groups. Pseudomonas aeruginose was isolated in all cases. The main complication of the omental pedicle was vascular congestion and peritracheal hemorrhage. CONCLUSIONS: Contrary to the data reported to date, we found that tracheal transplantation is viable in a large mammal like the sheep. The main complication observed in this animal model was graft infection. The use of an omental pedicle with the technique applied worsened the grafts survival. The encouraging results obtained in this investigation justify further research in order to manage graft infection, leading us to establish a suitable large animal model for allotransplantation.

Effect of vascular endothelial growth factor gene transfer on infarct size, left ventricular function and myocardial perfusion in sheep after 2 months of coronary artery occlusion.

BACKGROUND: In large mammalian models of acute myocardial infarction (AMI), plasmid-mediated vascular endothelial growth factor (pVEGF) gene transfer has been shown to induce angio-arteriogenesis, proliferation of myocyte precursors and adult cardiomyocyte mitosis, reducing infarct size at 15 days after coronary artery occlusion. However, it is unknown whether these effects persist at longer follow-up times, nor how they affect cardiac performance. We thus assessed infarct size, left ventricular (LV) function and perfusion in 2-month-old ovine AMI. METHODS: Adult sheep with coronary artery occlusion were randomized to blindly receive ten intramyocardial injections of 3.8 mg of pVEGF or empty plasmid distributed at the infarct border. Three and 60 days later, LV perfusion (single-photon emission computed tomography) and function (stress echocardiography) were assessed. Finally, hemodynamics (LV catheterization), scar size and peri-infarct histology were studied. RESULTS: Infarct size was 30% smaller in pVEGF-treated sheep (23.6 ± 1.9% versus 32.7 ± 2.7% of the LV; p < 0.02). Percentage fractional shortening and wall thickening at the infarct border improved after pVEGF, as did myocardial perfusion and LV wall motion under pharmacological stress. Global LV function did not differ between groups, although the force-frequency response was preserved in pVEGF group and lost in placebo animals. These effects were associated with angio-arteriogenesis and proliferation of cardiomyocyte precursors. CONCLUSIONS: In sheep with AMI, pVEGF gene transfer affords long-term infarct size reduction, yielding regional LV function and perfusion improvement and reducing remodeling progression. These results suggest the potential usefulness of this approach in the clinical setting.

Coronary microcirculation remodeling in patients with idiopathic dilated cardiomyopathy.

BACKGROUND: In patients with idiopathic dilated cardiomyopathy (IDCM), the myocardial blood flow is markedly depressed. The presence of alterations in the number and length of the coronary microcirculation has not been explored. METHODS: In explanted hearts of 6 patients with IDCM and in 6 normal control hearts, the arteriolar length density and capillary numerical density were determined in sections stained with orcein (vessels 50-200 µm in diameter), for smooth muscle actin (vessels 6-50 µm in diameter) and CD34 (capillaries). RESULTS: No difference with normal hearts was observed in capillary numerical density and in length density of vessels above 50 µm in diameter. In IDCM, more than 50% of arterioles below 50 µm in diameter displayed an incomplete smooth muscle wall, and length density, especially for arterioles between 6 and 20 µm in diameter, was significantly lower (42.84 ± 8.51 mm/mm(3)) than in controls (75.34 ± 3.05 mm/mm(3), p = 0.001). CONCLUSIONS: In IDCM, arterioles below 50 µm in diameter display an incomplete smooth muscle wall and a significant decrease in length density. These observations provide an anatomical basis for a decreased coronary flow reserve in IDCM.

Comparison of prevalence, clinical course, and pathological findings of left ventricular systolic impairment versus normal systolic function in patients with hypertrophic cardiomyopathy.

Impaired left ventricular systolic function (ILVSF) in hypertrophic cardiomyopathy (HC) is a risk factor for sudden death and a determinant of high mortality. We determined its prevalence, clinical parameters, long-term outcome, and pathologic findings of explanted hearts. We retrospectively analyzed 382 patients with HC; ILVSF was characterized by LV ejection fraction <50% at rest and was identified in 24 patients (6.3%). Patients with ILVSF were younger than patients with normal SF (43.5 ± 14.1 vs 55.3 ± 20.4 years, p = 0.001) and had larger LV end-diastolic cavity diameter (53.2 ± 12.2 vs 43.8 ± 6.2 mm, p = 0.001), larger left atrium (51.2 ± 6.5 vs 44.3 ± 8 mm, p <0.001), and lower fractional shortening (30.7 ± 11.1% vs 45.5% ± 10.3%, p <0.001). A combined end point (heart failure death or heart transplantation) was considered. Median follow-up was 3 years (1.2 to 6.3). Fourteen patients with ILVSF (58.3%) had the end point compared to 3 (0.8%) with normal SF (p <0.001). In explanted hearts, fibrosis represented 30.5 ± 12.5% of the left ventricle; we observed a direct correlation between fibrosis and ventricular dilation (r = 0.794, p = 0.001) and an inverse correlation between fibrosis and ejection fraction (r = -0.623, p = 0.023). Number and length density of small arterioles (<50 µm in diameter) were significantly decreased. In conclusion, ILVSF in HC has a poor prognosis and is associated with fibrosis and selective decreased development of small arterioles.

Cardiomyocyte hypertrophy, oncosis, and autophagic vacuolization predict mortality in idiopathic dilated cardiomyopathy with advanced heart failure.

OBJECTIVES: The aim of this study was to identify the remodeling parameters cardiomyocyte (CM) damage or death, hypertrophy, and fibrosis that may be linked to outcomes in patients with advanced heart failure (HF) in an effort to understand the pathogenic mechanisms of HF that may support newer therapeutic modalities. BACKGROUND: There are controversial results on the influence of fibrosis, CM hypertrophy, and apoptosis on outcomes in patients with HF; other modalities of cell damage have been poorly investigated. METHODS: In endomyocardial biopsy specimens from 100 patients with idiopathic dilated cardiomyopathy and advanced HF, CM diameter and the extent of fibrosis were determined by morphometry. The proportion of CMs with evidence of apoptosis, autophagic vacuolization (AuV), and oncosis was investigated by immunohistochemical methods and by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling. Those parameters were correlated with mortality in 3 years of follow-up by univariate analysis and with multivariate models incorporating the clinical variables more relevant to the prediction of outcomes. RESULTS: CM AuV occurred in 28 patients (0.013 ± 0.012%) and oncosis in 41 (0.109 ± 0.139%). Nineteen patients showed both markers. Apoptotic CM nuclei were observed in 3 patients. In univariate analysis, CM diameter and AuV, either alone or associated with oncosis, were predictors of mortality. In multivariate analysis, CM diameter (hazard ratio: 1.37; 95% confidence interval: 1.12 to 1.68; p = 0.002) and simultaneous presence in the same endomyocardial biopsy specimen of AuV and oncosis (hazard ratio: 2.82; 95% confidence interval: 1.12 to 7.13; p = 0.028) were independent predictors of mortality. CONCLUSIONS: CM hypertrophy and AuV, especially in association with oncosis, are predictors of outcome in patients with idiopathic dilated cardiomyopathy and severe HF.

An ovine model of postinfarction dilated cardiomyopathy in animals with highly variable coronary anatomy.

Studies on cardiac regeneration require large mammalian models of dilated cardiomyopathy (DCM) after acute myocardial infarction (AMI), and pig and sheep models are increasingly used in this field of preclinical research. Given the large interindividual variability in ovine left anterior descending artery (LAD) anatomy, protocols based on the coronary arteries to be ligated often lead to significant variation in infarct sizes and hence to heterogeneous results, ranging from no ventricular remodeling to acute, lethal left ventricular (LV) failure. We designed an ovine model of postinfarction DCM based on estimated infarct size rather than on a predetermined menu of coronary artery ligatures. In seven adult sheep we induced an anterolateral AMI of approximately 25% of the LV mass by ligating the branches of the LAD that, by visual inspection, would lead to such an infarct size. In 10 to 12 weeks, LV end-diastolic volume more than doubled and LV end-systolic volume almost tripled. LV ejection fraction decreased dramatically, as did LV percent fractional shortening and LV percent wall thickening. Infarct size (planimetry) was approximately 25% of the LV endocardial surface. We conclude that in sheep, an anterolateral AMI of approximately 25% of the LV mass--regardless of the coronary branches ligated to attain that infarct size--results in a model of postinfarction DCM that may prove useful in preclinical research on myocardial regeneration.

[Accuracy of multislice computed tomography in the diagnosis of hepatocellular carcinoma in patients with cirrhosis evaluated for liver transplantation]. / Precisión de la tomografía multidetector en el diagnóstico de hepatocarcinoma en pacientes con cirrosis evaluados para trasplante hepático.

BACKGROUND: The presence of hepatocellular carcinoma (HCC) within Milan criteria provides additional points in the MELD (Model for end stage liver disease) system and benefits in the order of organ allocation. The imaging methods play a key role in this process and represent an essential tool in the diagnosis and staging of HCC. OBJECTIVES: 1) To assess the accuracy of dynamic multidetector computed tomography (MDCT) in the diagnosis of HCC in patients with cirrhosis who are listed for liver transplantation. 2) To evaluate the diagnostic performance of TCMD in relation to tumor size. MATERIAL AND METHODS: We retrospectively reviewed the reports of MDCT performed in our institution to 62 patients who were then transplanted The histological analysis of explants was considered as the reference method in the diagnosis of HCC. MDCT studies were performed with dynamic protocol in arterial, portal and late phases. RESULTS: Dynamic MDCT showed a sensitivity of 87.5% and correctly characterized 28 of 35 patients with pathology proved hepatocellular carcinoma. MDCT was negative in 25 of 30 patients without hepatocellular carcinoma in the explanted liver, with a specificity of 83.3%. Nodule by nodule evaluation revealed a sensitivity of 80.3% and a specificity of 72.2%. CONCLUSION: In our center MDCT presented high accuracy in the correct diagnosis of HCC, showing its reliability when requesting additional points for organ allocation.

Molecular identification of Trypanosoma cruzi discrete typing units in end-stage chronic Chagas heart disease and reactivation after heart transplantation.

BACKGROUND: One hundred years after the discovery of Chagas disease, it remains a major neglected tropical disease. Chronic Chagas heart disease (cChHD) is the most severe manifestation. Heart transplantation is the proper treatment for end-stage heart failure, although reactivation of disease may result after receipt of immunosuppressive therapy. T. cruzi strains cluster into 6 discrete typing units (DTUs; I-VI) associated with different geographical distribution, transmission cycles and varying disease symptoms. In the southern cone of South America, T. cruzi II, V, and VI populations appear to be associated with Chagas disease and T. cruzi I with sylvatic cycles. METHODS: Molecular characterization of DTUs, T. cruzi I genotypes (on the basis of spliced-leader gene polymorphisms), and minicircle signatures was conducted using cardiac explant specimens and blood samples obtained from a cohort of 16 Argentinean patients with cChHD who underwent heart transplantation and from lesion samples obtained from 6 of these patients who presented with clinical reactivation of Chagas disease. RESULTS: Parasite persistence was associated with myocarditis progression, revealing T. cruzi I (genotype Id) in 3 explant samples and T. cruzi II, V, or VI in 5 explant samples. Post-heart transplantation follow-up examination of bloodstream DTUs identified T. cruzi I in 5 patients (genotypes Ia or Id) and T. cruzi II, V, or VI in 7 patients. T. cruzi I, V, and VI were detected in skin chagoma specimens, and T. cruzi V and VI were detected in samples obtained from patients with myocarditis reactivations. Multiple DTUs or genotypes at diverse body sites and polymorphic minicircle signatures at different cardiac regions revealed parasite histotropism. T. cruzi I infections clustered in northern Argentina (latitude, 23 degrees S-27 degrees S), whereas T. cruzi II, V, or VI DTUs were more ubiquitous. CONCLUSIONS: Multiple DTUs coexist in patients with Chagas disease. The frequent finding of T. cruzi I associated with cardiac damage was astounding, revealing its pathogenic role in cChHD at the southern cone.

Long-term effects of growth hormone on infarct size and left ventricular function in sheep with coronary artery occlusion.

The effects of growth hormone (GH) on infarct size and left ventricular (LV) function in experimental acute myocardial infarction (AMI) have been controversial. Moreover, little, if any, information exists regarding long-term evaluation of therapeutic doses of GH in large mammalian models of AMI. We therefore aimed to assess the effect of therapeutic doses of GH over 3.5 months on infarct size and heart function in sheep with AMI. After coronary artery ligation, sheep received subcutaneous human GH 8 IU/d (n = 8) or vehicle (n = 8) over 100 days. Infarct area was similar in GH (16.9% +/- 3% of LV area) and placebo (16.5% +/- 3.7%, P = not significant) sheep. At 3 days of treatment onset, but not at later times, GH sheep had higher LV shortening fraction (30.7% +/- 3.5% vs. 24.8% +/- 6.1%, P < 0.04), systolic anterior wall thickness (10.1 +/- 0.8 vs. 8.6 +/- 1.2 mm, P < 0.02), and cardiac index (3.8 +/- 0.6 vs. 2.8 +/- 0.7 L x min x m, P < 0.01). This evolution of function parameters paralleled that of serum insulin-like growth factor 1 levels, which differed significantly only during the first week, suggesting a direct effect of GH on LV contractility. These results may suggest the usefulness of therapeutic doses of GH at the early phases of AMI but do not support maintaining the treatment for longer time.