A rodent model of NASH with cirrhosis, oval cell proliferation and hepatocellular carcinoma

Background/Aims: Hepatocellular carcinoma (HCC) is a well recognized complication of advanced NASH (non-alcoholic steatohepatitis). We sought to produce a rat model of NASH, cirrhosis and HCC. Methods: Adult Sprague-Dawley rats, weighing 250-300 g, were fed a choline-deficient, high trans-fat diet and exposed to DEN in drinking water. After 16 weeks, the animals underwent liver ultrasound (US), sacrifice and assessment by microscopy, immunohistochemistry and transmission electron microscopy (TEM). Results: US revealed steatosis and focal lesions in 6 of 7. All had steatohepatitis defined as inflammation, advanced fibrosis and ballooning with Mallory-Denk bodies (MDB) with frank cirrhosis in 6. Areas of more severe injury were associated with anti-CK19 positive ductular reaction. HCC, present in all, were macro-trabecullar or solid with polyhedral cells with foci of steatosis and ballooned cells. CK19 was positive in single or solid nests of oval cells and in neoplastic hepatocytes. TEM showed ballooning with small droplet fat, dilated endoplasmic reticulum and MDB in non-neoplastic hepatocytes and small droplet steatosis in some cancer cells. Conclusions: This model replicated many features of NASH including steatohepatitis with ballooning, fibrosis, cirrhosis and hepatocellular carcinoma. Oval cell proliferation was evident and the presence anti-CK 19 positivity in the cancer suggests oval cell origin of the malignancy. (C) 2008 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.

Implications of prevalent and incident diabetes mellitus on left ventricular geometry and function in the ageing heart: The MONICA/KORA Augsburg cohort study

Background and Aim: It is unclear to what extent diabetes modulates the ageing-related adaptations of cardiac geometry and function. Methods and Results: We examined 1005 adults, aged 25-74 years, from a population-based survey at baseline in 1994/5 and at follow-up in 2004/5. We compared persistently non-diabetic individuals (ND; no diabetes at baseline and at follow-up, n = 833) with incident (ID; non-diabetic at baseline and diabetic at follow-up, n = 36) and with prevalent diabetics (PD; diabetes at baseline and follow-up examination, n = 21). Left ventricular (LV) geometry and function were evaluated by echocardiography. Statistical analyses were performed with multivariate linear regression models. Over ten years the PD group displayed a significantly stronger relative increase of LV mass (+9.34% vs. +23.7%) that was mediated by a more pronounced increase of LV end-diastolic diameter (+0% vs. +6.95%) compared to the ND group. In parallel, LA diameter increased (+4.50% vs. +12.7%), whereas ejection fraction decreased (+3.02% vs. -4.92%) more significantly in the PD group. Moreover, at the follow-up examination the PD and ID groups showed a significantly worse diastolic function, indicated by a higher E/EM ratio compared with the ND group (11.6 and 11.8 vs. 9.79, respectively). Conclusions: Long-standing diabetes was associated with an acceleration of age-related changes of left ventricular geometry accumulating in an eccentric remodelling of the left ventricle. Likewise, echocardiographic measures of systolic and diastolic ventricular function deteriorated more rapidly in individuals with diabetes. (C) 2009 Elsevier B.V. All rights reserved.

Pkd1 Haploinsufficiency Increases Renal Damage and Induces Microcyst Formation following Ischemia/Reperfusion

Mutations in PKD1 cause the majority of cases of autosomal dominant polycystic kidney disease (ADPKD). Because polycystin 1 modulates cell proliferation, cell differentiation, and apoptosis, its lower biologic activity observed in ADPKD might influence the degree of injury after renal ischemia/reperfusion. We induced renal ischemia/reperfusion in 10- to 12-wk-old male noncystic Pkd1(+/-) and wild-type mice. Compared with wild-type mice, heterozygous mice had higher fractional excretions of sodium and potassium and higher serum creatinine after 48 h. In addition, in heterozygous mice, also cortical damage, rates of apoptosis, and inflammatory infiltration into the interstitium at time points out to 14 d after injury all increased, as well as cell proliferation at 48 h and 7 d. The mRNA and protein expression of p21 was lower in heterozygous mice than wild-type mice at 48 h. After 6 wk, we observed dilated tubules, microcysts, and increased renal fibrosis in heterozygotes. The early mortality of heterozygotes was significantly higher than that of wild-type mice when we extended the duration of ischemia from 32 to 35 min. In conclusion, ischemia/reperfusion induces a more severe injury in kidneys of Pkd1-haploin-sufficient mice, a process that apparently depends on a relative deficiency of p2l activity, tubular dilation, and microcyst formation. These data suggest the possibility that humans with ADPKD from PKD1 mutations may be at greater risk for damage from renal ischemia/reperfusion injury.

De novo copy number variants identify new genes and loci in isolated sporadic tetralogy of Fallot

Tetralogy of Fallot (TOF), the most common severe congenital heart malformation, occurs sporadically, without other anomaly, and from unknown cause in 70% of cases. Through a genome-wide survey of 114 subjects with TOF and their unaffected parents, we identified 11 de novo copy number variants (CNVs) that were absent or extremely rare (<0.1%) in 2,265 controls. We then examined a second, independent TOF cohort (n = 398) for additional CNVs at these loci. We identified CNVs at chromosome 1q21.1 in 1% (5/512, P = 0.0002, OR = 22.3) of nonsyndromic sporadic TOF cases. We also identified recurrent CNVs at 3p25.1, 7p21.3 and 22q11.2. CNVs in a single subject with TOF occurred at six loci, two that encode known (NOTCH1, JAG1) disease-associated genes. Our findings predict that at least 10% (4.5-15.5%, 95% confidence interval) of sporadic nonsyndromic TOF cases result from de novo CNVs and suggest that mutations within these loci might be etiologic in other cases of TOF.

Preliminary findings from a new animal model for Peyronie`s disease involving extracorporeal shock waves

To develop an experimental model in rabbits to analyse the efficiency of extracorporeal shock wave therapy (ESWT) for Peyronie`s disease. We used 15 adult male rabbits divided into three equal groups. In group 1 (no penile ESWT) rabbits had three sessions of ESWT with 2000 shocks each (15 kV), but a rubber mat was placed between the shock head and rabbit to protect the penis; the rabbits were killed at 7 days after the last session of ESWT. In group 2 the rabbits had three sessions of ESWT using the same parameters, and were killed immediately after the last session to analyse the penis. In group 3 the rabbits had three sessions of ESWT as before but were killed at 7 days after the last session, and the penile tissue analysed macroscopically and histologically. The results showed clearly that the model was efficient, creating a similar situation to that when applying ESWT in the human penis. All of the rabbits in groups 2 and 3 had haematomas and diffuse petechiae after ESWT, and only four had urethral and penile bleeding. Almost all macroscopic changes disappeared after 48 h and only one rabbit in group 3 after 7 days had a haematoma on the dorsal penile surface. The histology (assessed using haematoxylin and eosin staining) of the cavernous body of the penis showed: unchanged histology in group 1; in group 2 there was a dilated and congested vascular space in the cavernous body, with interstitial extensive bleeding in the dermis; and in group 3 there was an increase in interstitial fibrous tissue in the cavernous septum, with deposition of collagen fibres and thickening of the tunica albuginea. The present model was efficient in producing tissue injury in the normal penis when treated with ESWT, suggesting that this promising model should be considered for use future studies of Peyronie`s disease.

Cavopulmonary anastomosis improves left ventricular assist device support in acute biventricular failure

Objective: Right ventricular failure during left ventricular assist device (WAD) support can result in severe hemodynamic compromise with high mortality. This study investigated the acute effects of cavopulmonary anastomosis on right ventricular loading and WAD performance in a model of severe biventricular failure. Methods: LVAD support was performed by means of centrifugal pump implantation in 14 anesthetized dogs (20-30 kg) with severe biventricular failure obtained by ventricular fibrillation induction. Animals were randomized to be submitted to classical cavopulmonary anastomosis (Glenn shunt) or to control group and were maintained under WAD support for 2 h. Left and right atrial, right ventricular and systemic pressures were monitored, white total pulmonary flow was simultaneously recorded by transonic flowmeters located on the superior vena cava and pulmonary trunk. Blood gas and venous lactate determinations were also obtained. Results: Ventricular fibrillation maintenance resulted in acute WAD performance impairment after 90 min in the control group, while animals with Glenn circuit maintained normal WAD pump flow (55 +/- 13 ml kg(-1) min(-1) vs 21 +/- 4 ml kg(-1) min(-1), p < 0.001) and better peripheral perfusion (blood lactate of 29 +/- 10 pg/ml vs 46 +/- 9 pg/ml, p < 0.001). Left and right atrial pressures did not change significantly, while right ventricular pressure was tower in animals with Glenn circuit (13 +/- 3 mmHg vs 22 +/- 8 mmHg, p = 0.005). Right ventricular unloading with Glenn shunt also resulted in superior total pulmonary flow (59 +/- 13 ml kg(-1) min(-1) vs 17 +/- 3 ml kg(-1) min(-1), p < 0.001). Conclusion: The concomitant use of cavopulmonary anastomosis during LVAD support in a model of severe biventricular failure limited right ventricular overloading and resulted in better hemodynamic performance. (C) 2008 European Association for Cardio-Thoracic Surgery. Published by Elsevier B.V. All rights reserved.

Contrast-enhanced magnetic resonance imaging identifies focal regions of intramyocardial fibrosis, in patients with severe aortic valve disease: Correlation with quantitative histopathology

Background Chronic aortic valve disease (AVD) is characterized by progressive accumulation of interstitial myocardial fibrosis (MF). However, assessment of MF accumulation has only been possible through histologic analyses of endomyocardial biopsies. We sought to evaluate contrast-enhanced magnetic resonance imaging (ce-MRI) as a noninvasive method to identify the presence of increased MF in patients with severe AVD. Methods Seventy patients scheduled to undergo aortic valve replacement surgery were examined by cine and ce-MRI in a 1.5-T scanner. Cine images were used for the assessment of left ventricular (LV) volumes, mass, and function. Delayed-enhancement images were used to characterize the regions of MF. In addition, histologic analyses of myocardial samples obtained during aortic valve replacement surgery were used for direct quantification of interstitial MF. Ten additional subjects who died of noncardiac causes served as controls for the quantitative histologic analyses. Results Interstitial MF determined by histopathologic analysis was higher in patients with AVID than in controls (2.7% +/- 2.0% vs 0.6% +/- 0.2%, P =.001). When compared with histopathologic results, ce-MRI demonstrated a sensitivity of 74%, a specificity of 81%, and an accuracy of 76% to identify AVD patients with increased interstitial MF There was a significant inverse correlation between interstitial MF and LV ejection fraction (r = -0.67, P <.0001). Accordingly, patients with identifiable focal regions of MF by ce-MRI exhibited worse LV systolic function than those without MF (45% +/- 14% vs 65% +/- 14%, P <.0001). Conclusions Contrast-enhanced MRI allows for the noninvasive detection of focal regions of MF in patients with severe AVD. Moreover, patients with identifiable MF by ce-MRI exhibited worse LV functional parameters. (Am Heart J 2009; 157:361-8.)

Cardiovascular Risk and Clinical Factors in Athletes: 10 Years of Evaluation

DE MATOS, L. D. N. J., N. D. A. O. CALDEIRA, P. D. S. PERLINGEIRO, I. L. G. DOS SANTOS, C. E. NEGRAO and L. F. AZEVEDO. Cardiovascular Risk and Clinical Factors in Athletes: 10 Years of Evaluation. Med. Sci. Sports Exerc., Vol. 43, No. 6, pp. 943-950, 2011. Purpose: Preparticipation screening in athletes is a very current but controversial theme. Part of this controversy is due to the cost benefit, especially when the screening is merely used as a prevention of sudden cardiac death caused by rare and hereditary diseases. The purpose of this study was to describe the prevalence of preexisting diseases, cardiovascular risk factor for cardiovascular diseases development, and hematological profile in a population of amateur and professional athletes. Methods: Data of 623 athletes (529 men and 94 women), aged 13-77 yr, were analyzed to detect preexisting diseases. The variables total cholesterol, LDL, HDL, triglycerides, fasting glucose, body mass index, hemoglobin, hematocrit, and ferritin were analyzed in two groups according to age, that is, younger and older 35 yr old, and their prevalence (%) and distribution in quartiles were presented. chi(2) test and Pearson product-moment correlation coefficients between variables were applied, and P < 0.05 was adopted for significance. Results: Hypertension was the most prevalent preexisting diseases, although the data showed low prevalence of cardiomyopathy. Cardiovascular risk factors were prevalent in both genders. There were positive correlations between cardiovascular risk factors and age and between body mass index and lipid levels in male athletes. Also, there was a high prevalence of low ferritin levels for women, with positive correlation between the levels of hemoglobin and ferritin. Conclusions: In the present study, hypertension was the most prevalent diagnosed disease, and cardiovascular risk factors showed important prevalence, especially in athletes older than 35 yr. Although physical training represents a cardioprotective factor to the onset of cardiovascular disease, it does not exclude the prevalence of risk factors and diseases in athletes.

Atrogin-1 and MuRF1 regulate cardiac MyBP-C levels via different mechanisms

Familial hypertrophic cardiomyopathy (FHC) is frequently caused by cardiac myosin-binding protein C (cMyBP-C) gene mutations, which should result in C-terminal truncated mutants. However, truncated mutants were not detected in myocardial tissue of FHC patients and were rapidly degraded by the ubiquitin-proteasome system (UPS) after gene transfer in cardiac myocytes. Since the diversity and specificity of UPS regulation lie in E3 ubiquitin ligases, we investigated whether the muscle-specific E3 ligases atrogin-1 or muscle ring finger protein-1 (MuRF1) mediate degradation of truncated cMyBP-C. Human wild-type (WT) and truncated (M7t, resulting from a human mutation) cMyBP-C species were co-immunoprecipitated with atrogin-1 after adenoviral overexpression in cardiac myocytes, and WT-cMyBP-C was identified as an interaction partner of MuRF1 by yeast two-hybrid screens. Overexpression of atrogin-1 in cardiac myocytes decreased the protein level of M7t-cMyBP-C by 80% and left WT-cMyBP-C level unaffected. This was rescued by proteasome inhibition. In contrast, overexpression of MuRF1 in cardiac myocytes not only reduced the protein level of WT- and M7t-cMyBP-C by > 60%, but also the level of myosin heavy chains (MHCs) by > 40%, which were not rescued by proteasome inhibition. Both exogenous cMyBP-C and endogenous MHC mRNA levels were markedly reduced by MuRF1 overexpression. Similar to cardiac myocytes, MuRF1-overexpressing (TG) mice exhibited 40% lower levels of MHC mRNAs and proteins. Protein levels of cMyBP-C were 29% higher in MuRF1 knockout and 34% lower in TG than in WT, without a corresponding change in mRNA levels. These data suggest that atrogin-1 specifically targets truncated M7t-cMyBP-C, but not WT-cMyBP-C, for proteasomal degradation and that MuRF1 indirectly reduces cMyBP-C levels by regulating the transcription of MHC.

Alterations in myocardial gene expression associated with experimental Trypanosoma cruzi infection

Chagas disease, characterized by acute myocarditis and chronic cardiomyopathy, is caused by infection with the protozoan parasite Trypanosoma cruzi. We sought to identify genes altered during the development of parasite-induced cardiomyopathy. Microarrays containing 27,400 sequence-verified mouse cDNAs were used to analyze global gene expression changes in the myocardium of a murine model of chagasic cardiomyopathy. Changes in gene expression were determined as the acute stage of infection developed into the chronic stage. This analysis was performed on the hearts of male CD-1 mice infected with trypomastigotes of T. cruzi (Brazil strain). At each interval we compared infected and uninfected mice and confirmed the microarray data with dye reversal. We identified eight distinct categories of mRNAs that were differentially regulated during infection and identified dysregulation of several key genes. These data may provide insight into the pathogenesis of chagasic cardiomyopathy and provide new targets for intervention. (c) 2008 Elsevier Inc. All rights reserved.

Structural evaluation of the effects of chronic ethanol ingestion on the testis of Calomys callosus

Chronic exposure to ethanol may results in pathophysiologic changes in cellular function. The present work was designed to investigate the morphology of testis submitted to experimental ethanol ingestion. Experimental animals were divided into two groups. The control group (n = 23) received a solid diet and tap water and the alcoholic group (n = 23) received the same solid diet and ethanol P.A. diluted 20% in water (v/v). After 120 days of treatment, all animals were anesthetized, weighed and sacrificed. Testosterone and luteinizing hormone levels in serum were lower in the alcoholic group than in the control group. Histological and ultrastructural alterations were observed in the testicular alcoholic germinative cells like enormous spaces, lipid droplets accumulation, digestive vacuoles, irregular diameter of the seminiferous tubules and interstitial dilated blood vessels. It was concluded that 20% ethanol provokes lesions on the testis germinative epithelium probably inducing gonadal dysfunction. (C) 2008 Elsevier Ltd. All rights reserved.

Acute chagasic myocardiopathy after orthotopic liver transplantation with donor and recipient serologically negative for Trypanosoma cruzi: A case report

Chagas disease (American trypanosomiasis) is caused by the protozoan parasite Trypanosoma cruzi. Chagas disease following solid-organ transplantation has occurred in Latin America. This report presents the occurrence of Chagas disease despite negative serological tests in both the donor and the recipient, as well as the effectiveness of treatment. A 21-year-old woman from the state of Sao Paulo (Brazil) underwent cadaveric donor liver transplantation in November 2005, due to cirrhosis of autoimmune etiology. Ten months after liver transplantation, she developed signs and symptoms of congestive heart failure (New York Heart Association functional class IV). The echocardiogram, which was normal preoperatively, showed dilated cardiac chambers, depressed left ventricular systolic function (ejection fraction = 35%) and moderate pulmonary hypertension. Clinical investigation discarded ischemic heart disease and autoimmune and other causes for heart failure. Immuno fluorescence (immunoglobulin M and immunoglobulin G) and hemagglutination tests for T cruzi were positive, and abundant T cruzi amastigotes were readily identified in myocardial biopsy specimens. Treatment with benznidazole for 2 months yielded an excellent clinical response. At the moment of submission, the patient remains in functional class I. This case highlighted that more appropriate screening for T cruzi infection is mandatory in potential donors and recipients of solid-organ transplants in regions where Chagas disease is prevalent. Moreover, it stressed that this diagnosis should always be considered in recipients who develop cardiac complications, since negative serological tests do not completely discard the possibility of disease transmission and since good results can be achieved with prompt trypanocidal therapy.

Chagas disease

Chagas disease is a chronic, systemic, parasitic infection caused by the protozoan Trypanosoma cruzi, and was discovered in 1909. The disease affects about 8 million people in Latin America, of whom 30-40% either have or will develop cardiomyopathy, digestive megasyndromes, or both. In the past three decades, the control and management of Chagas disease has undergone several improvements. Large-scale vector control programmes and screening of blood donors have reduced disease incidence and prevalence. Although more effective trypanocidal drugs are needed, treatment with benznidazole (or nifurtimox) is reasonably safe and effective, and is now recommended for a widened range of patients. Improved models for risk stratification are available, and certain guided treatments could halt or reverse disease progression. By contrast, some challenges remain: Chagas disease is becoming an emerging health problem in non-endemic areas because of growing population movements; early detection and treatment of asymptomatic individuals are underused; and the potential benefits of novel therapies (eg, implantable cardioverter defibrillators) need assessment in prospective randomised trials.

Genital Manifestation of Graft-vs.-Host Disease: A Series of Case Reports

Introduction. After hematopoietic stem cell transplantation (HSCT), many patients present genital graft-vs.-host disease (GVHD) that can culminate with sexual problems, which are poorly dimensioned. Aim. We hope to draw attention to the need to perform genital biopsy to diagnose genital GVHD, and thus to call attention to the need to incorporate careful attention to sexual health in the treatment of these patients. Methods. Five allogeneic stem cell transplant recipients complaining of coital pain after HSCT were clinically diagnosed for genital GVHD. Genital biopsies were given for histological analysis, and microphotographs of the corresponding marked field in the slide were taken. Specimens were evaluated by the site pathologist and then sent to a reference pathologist, each blinded to the histological findings. A literature search was performed in PubMed/MEDLINE (1966-2009) for cross-sectional and cohort studies or trials related to genital GVHD. Expert opinions peer reviews and case reports were also considered. Main Outcome Measures. HSCT, genital GVHD, genital biopsy. Results. The biopsy showed evidence of dilated apoptotic cells in the basal layer and detachment of the epithelial lining of the mucosa, hyalinization and thickening of collagen fibers, capillary ectasia, and mononuclear inflammatory infiltrate of the submucosa. Three patients presented vulval lesion such as leucoplasia and ulcer on the large lip. Histological analyses showed evidence of epithelial hyperplasia and influx of inflammatory cells to the epithelial surface, intercellular edema and spongiosis, apoptotic bodies on the basal layer of the epithelium, spongiosis, and nuclear vacuolization. A common treatment based on corticotherapy resulted in complete remission of coetaneous or mucous genital lesions in all five patients. Conclusion. Genital biopsy is important to differentially diagnose GVHD and secondary symptoms due to hypoestrogenism. Prevention is the most important step in controlling the evolution GVHD in the vagina to prevent vaginal obstruction and sexual dysfunction. da Silva Lara LA, de Andrade JM, Mauad LMQ, Ferrarese SR, Marana HRC, Tiezzi DG, and de Sa Rosa e Silva ACJ. Genital manifestation of graft-vs.-host disease: A series of case reports. J Sex Med 2010;7:3216-3225.

Disruption of sarcolemmal dystrophin and beta-dystroglycan may be a potential mechanism for myocardial dysfunction in severe sepsis

Evidence from our laboratory has shown alterations in myocardial structure in severe sepsis/septic shock. The morphological alterations are heralded by sarcolemmal damage, characterized by increased plasma membrane permeability caused by oxidative damage to lipids and proteins. The critical importance of the dystrophin-glycoprotein complex (DGC) in maintaining sarcolemmal stability led us to hypothesize that loss of dystrophin and associated glycoproteins could be involved in early increased sarcolemmal permeability in experimentally induced septic cardiomyopathy. Male C57Bl/6 mice were subjected to sham operation and moderate (MSI) or severe (SSI) septic injury induced by cecal ligation and puncture (CLP). Using western blot and immunofluorescence, a downregulation of dystrophin and beta-dystroglycan expression in both severe and moderate injury could be observed in septic hearts. The immunofluorescent and protein amount expressions of laminin-alpha 2 were similar in SSI and sham-operated hearts. Consonantly, the evaluation of plasma membrane permeability by intracellular albumin staining provided evidence of severe injury of the sarcolemma in SSI hearts, whereas antioxidant treatment significantly attenuated the loss of sarcolemmal dystrophin expression and the increased membrane permeability. This study offers novel and mechanistic data to clarify subcellular events in the pathogenesis of cardiac dysfunction in severe sepsis. The main finding was that severe sepsis leads to a marked reduction in membrane localization of dystrophin and beta-dystroglycan in septic cardiomyocytes, a process that may constitute a structural basis of sepsis-induced cardiac depression. In addition, increased sarcolemmal permeability suggests functional impairment of the DGC complex in cardiac myofibers. In vivo observation that antioxidant treatment significantly abrogated the loss of dystrophin expression and plasma membrane increased permeability supports the hypothesis that oxidative damage may mediate the loss of dystrophin and beta-dystroglycan in septic mice. These abnormal parameters emerge as therapeutic targets and their modulation may provide beneficial effects on future cardiovascular outcomes and mortality in sepsis. Laboratory Investigation (2010) 90, 531-542; doi: 10.1038/labinvest.2010.3; published online 8 February 2010