RESUMO
Abstract Background: In view of the increased global prevalence of cardiovascular and hepatic diseases, the diet lipid content and its relationship with the accumulation of fat in hepatocytes have been investigated as key factors in preventing these diseases. Objective: To evaluate the metabolic effects of a high-lard diet supplemented or not with cholesterol on a modified dyslipidemia model. Methods: We divided 24 adult male Wistar rats into three groups: standard diet (STD - 4% lipids), high-lard diet (HLD - 21% lard), and high-lard and high-cholesterol diet (HL/HCD - 20% lard, 1% cholesterol, 0.1% cholic acid). After six weeks of treatment, blood and liver were collected for biochemical (serum lipid profile and liver enzymes) and morphological analyses. Statistical analysis included one-way analysis of variance (ANOVA), followed by Tukey test for mean comparisons, and a 5% probability was considered statistically significant. Results: Animals fed HL/HCD showed increased total cholesterol, triacylglycerol, LDL-c, non-HDL-c, alanine aminotransferase (ALT), and aspartate aminotransferase (AST) serum levels compared to those fed STD. In addition, the HL/HCD animals presented higher relative liver weight, with moderate macrovesicular hepatic steatosis and inflammatory infiltrate. Conclusion: A high-fat diet with lard (20%) and cholesterol (1%) triggered dyslipidemia with severe liver damage in rats in a shorter experimental time than the previously reported models. The high-lard diet without supplementation of cholesterol led to body weight gain, but not to dyslipidemia.
Resumo Fundamento: Tendo em vista o aumento da prevalência global de doenças cardiovasculares e hepáticas, o conteúdo lipídico da dieta e sua relação com o acúmulo de gordura nos hepatócitos têm sido investigados como fatores-chave na prevenção dessas doenças. Objetivo: Avaliar os efeitos metabólicos de uma dieta rica em banha suplementada com colesterol ou não, em um modelo modificado de dislipidemia. Métodos: Foram divididos 24 ratos Wistar machos adultos em três grupos: dieta padrão (DP - 4% de lipídios), dieta rica em banha (DRB - 21% de banha) e dieta rica em banha e colesterol (DRB/RC - 20% de banha, 1% de colesterol e 0,1% de ácido cólico). Após seis semanas de tratamento, o sangue e o fígado foram coletados para análises bioquímicas (perfil lipídico sérico e enzimas hepáticas) e morfológicas. A análise estatística incluiu análise de variância unidirecional (ANOVA), seguida do teste de Tukey para comparações de médias. Uma probabilidade de 5% foi considerada estatisticamente significativa. Resultados: Animais alimentados com DRB/RC apresentaram um aumento nos níveis séricos de colesterol total, triacilglicerol, LDL-c, não-HDL-c, alanina aminotransferase (ALT) e aspartato aminotransferase (AST) em comparação com aqueles alimentados com DP. Além disso, os animais tratados com DRB/RC apresentaram um peso relativo do fígado maior, com esteatose hepática macrovesicular moderada e infiltrado inflamatório. Conclusão: Uma dieta rica em gordura com banha (20%) e colesterol (1%) desencadeou dislipidemia com danos graves ao fígado em ratos em um tempo experimental menor do que os modelos previamente relatados. A dieta rica em banha sem suplementação de colesterol levou ao ganho de peso corporal, mas não à dislipidemia.
Assuntos
Animais , Masculino , Dislipidemias/induzido quimicamente , Dieta Hiperlipídica/efeitos adversos , Doenças Metabólicas/etiologia , Tamanho do Órgão , Aspartato Aminotransferases/sangue , Triglicerídeos/sangue , Peso Corporal , Gorduras na Dieta/efeitos adversos , Colesterol/efeitos adversos , Colesterol/sangue , Ratos Wistar , Alanina Transaminase/sangue , Modelos Animais de Doenças , Dislipidemias/metabolismo , Dislipidemias/sangue , Fígado Gorduroso/patologia , Inflamação , HDL-Colesterol/sangue , LDL-Colesterol/sangue , Fígado/metabolismo , Fígado/patologia , Doenças Metabólicas/metabolismo , Doenças Metabólicas/sangueRESUMO
BACKGROUND: In view of the increased global prevalence of cardiovascular and hepatic diseases, the diet lipid content and its relationship with the accumulation of fat in hepatocytes have been investigated as key factors in preventing these diseases. OBJECTIVE: To evaluate the metabolic effects of a high-lard diet supplemented or not with cholesterol on a modified dyslipidemia model. METHODS: We divided 24 adult male Wistar rats into three groups: standard diet (STD - 4% lipids), high-lard diet (HLD - 21% lard), and high-lard and high-cholesterol diet (HL/HCD - 20% lard, 1% cholesterol, 0.1% cholic acid). After six weeks of treatment, blood and liver were collected for biochemical (serum lipid profile and liver enzymes) and morphological analyses. Statistical analysis included one-way analysis of variance (ANOVA), followed by Tukey test for mean comparisons, and a 5% probability was considered statistically significant. RESULTS: Animals fed HL/HCD showed increased total cholesterol, triacylglycerol, LDL-c, non-HDL-c, alanine aminotransferase (ALT), and aspartate aminotransferase (AST) serum levels compared to those fed STD. In addition, the HL/HCD animals presented higher relative liver weight, with moderate macrovesicular hepatic steatosis and inflammatory infiltrate. CONCLUSION: A high-fat diet with lard (20%) and cholesterol (1%) triggered dyslipidemia with severe liver damage in rats in a shorter experimental time than the previously reported models. The high-lard diet without supplementation of cholesterol led to body weight gain, but not to dyslipidemia.
Assuntos
Dieta Hiperlipídica/efeitos adversos , Dislipidemias/induzido quimicamente , Doenças Metabólicas/etiologia , Alanina Transaminase/sangue , Animais , Aspartato Aminotransferases/sangue , Peso Corporal , Colesterol/efeitos adversos , Colesterol/sangue , HDL-Colesterol/sangue , LDL-Colesterol/sangue , Gorduras na Dieta/efeitos adversos , Modelos Animais de Doenças , Dislipidemias/sangue , Dislipidemias/metabolismo , Fígado Gorduroso/patologia , Inflamação , Fígado/metabolismo , Fígado/patologia , Masculino , Doenças Metabólicas/sangue , Doenças Metabólicas/metabolismo , Tamanho do Órgão , Ratos Wistar , Triglicerídeos/sangueRESUMO
Dystrophin, an important protein of the dystrophin-glycoprotein complex, has been implicated in the pathogenesis of experimental Chagas disease. It is important for the maintenance of cell shape and contraction force transmission. Dystrophin loss has been related to end-stage cardiac myopathies and proposed as a common route for myocardial dysfunction and progression to advanced heart failure. Evidence suggests that calpains, calcium-dependent proteases, digest dystrophin when the calcium concentration is compatible with their activation. The objective of this in vitro study was to test the hypothesis that dantrolene, a calcium channel blocker, improves structural changes induced by serum from Trypanosoma cruzi-infected mice. Cultured neonatal cardiac myocytes were incubated with serum from T. cruzi-infected mice and treated with dantrolene for 24 h. Immunofluorescence and immunoblotting were performed to evaluate dystrophin and calpain-1 protein expression. The levels of dystrophin decreased 13 % and calpain increased 17 % after incubation of cultured neonatal cardiac myocytes with serum from T. cruzi-infected mice. The treatment with dantrolene restored the dystrophin and calpain levels near control levels. Our results demonstrate that alterations in calcium homeostasis in cardiac myocytes are responsible, in part, for cardiac structural changes in experimentally induced T. cruzi myocarditis and that calpain inhibitors may be beneficial in Chagasic heart disease.
Assuntos
Doença de Chagas/sangue , Dantroleno/farmacologia , Distrofina/química , Soro , Trypanosoma cruzi , Animais , Animais Recém-Nascidos , Células Cultivadas , Doença de Chagas/patologia , Imunofluorescência , Camundongos , Relaxantes Musculares Centrais/farmacologia , Miócitos CardíacosRESUMO
Cardiac dysfunction caused by the impairment of myocardial contractility has been recognized as an important factor contributing to the high mortality in sepsis. Calpain activation in the heart takes place in response to increased intracellular calcium influx resulting in proteolysis of structural and contractile proteins with subsequent myocardial dysfunction. The purpose of the present study was to test the hypothesis that increased levels of calpain in the septic heart leads to disruption of structural and contractile proteins and that administration of calpain inhibitor-1 (N-acetyl-leucinyl-leucinyl-norleucinal (ALLN)) after sepsis induced by cecal ligation and puncture prevents cardiac protein degradation. We also tested the hypothesis that calpain plays a role in the modulation of protein synthesis/degradation through the activation of proteasome-dependent proteolysis and inhibition of the mTOR pathway. Severe sepsis significantly increased heart calpain-1 levels and promoted ubiquitin and Pa28ß over-expression with a reduction in the mTOR levels. In addition, sepsis reduced the expression of structural proteins dystrophin and ß-dystroglycan as well as the contractile proteins actin and myosin. ALLN administration prevented sepsis-induced increases in calpain and ubiquitin levels in the heart, which resulted in decreased of structural and contractile proteins degradation and basal mTOR expression levels were re-established. Our results support the concept that increased calpain concentrations may be part of an important mechanism of sepsis-induced cardiac muscle proteolysis.
Assuntos
Calpaína/metabolismo , Distrofina/metabolismo , Complexo de Endopeptidases do Proteassoma/metabolismo , Sepse/patologia , Serina-Treonina Quinases TOR/metabolismo , Ubiquitina/metabolismo , Actinas/metabolismo , Animais , Calpaína/antagonistas & inibidores , Calpaína/genética , Cardiomiopatias/complicações , Cardiomiopatias/patologia , Modelos Animais de Doenças , Expressão Gênica/efeitos dos fármacos , Leupeptinas/farmacologia , Leupeptinas/uso terapêutico , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Microscopia de Fluorescência , Miocárdio/metabolismo , Miocárdio/patologia , Miosinas/metabolismo , Proteólise/efeitos dos fármacos , Sepse/etiologia , Sepse/prevenção & controle , Serina-Treonina Quinases TOR/antagonistas & inibidoresRESUMO
The critical importance of dystrophin to cardiomyocyte contraction and sarcolemmal and myofibers integrity, led us to test the hypothesis that dystrophin reduction/loss could be involved in the pathogenesis of doxorubicin-induced cardiomyopathy, in order to determine a possible specific structural culprit behind heart failure. Rats received total cumulative doses of doxorubicin during 2 weeks: 3.75, 7.5, and 15 mg/kg. Controls rats received saline. Fourteen days after the last injection, hearts were collected for light and electron microscopy, immunofluorescence and western blot. The cardiac function was evaluated 7 and 14 days after drug or saline. Additionally, dantrolene (5 mg/kg), a calcium-blocking agent that binds to cardiac ryanodine receptors, was administered to controls and doxorubicin-treated rats (15 mg/kg). This study offers novel and mechanistic data to clarify molecular events that occur in the myocardium in doxorubicin-induced chronic cardiomyopathy. Doxorubicin led to a marked reduction/loss in dystrophin membrane localization in cardiomyocytes and left ventricular dysfunction, which might constitute, in association with sarcomeric actin/myosin proteins disruption, the structural basis of doxorubicin-induced cardiac depression. Moreover, increased sarcolemmal permeability suggests functional impairment of the dystrophin-glycoprotein complex in cardiac myofibers and/or oxidative damage. Increased expression of calpain, a calcium-dependent protease, was markedly increased in cardiomyocytes of doxorubicin-treated rats. Dantrolene improved survival rate and preserved myocardial dystrophin, calpain levels and cardiac function, which supports the opinion that calpain mediates dystrophin loss and myofibrils degradation in doxorubicin-treated rats. Studies are needed to further elucidate this mechanism, mainly regarding specific calpain inhibitors, which may provide new interventional pathways to prevent doxorubicin-induced cardiomyopathy.
Assuntos
Calpaína/metabolismo , Cardiomiopatias/induzido quimicamente , Cardiomiopatias/metabolismo , Doxorrubicina/efeitos adversos , Distrofina/metabolismo , Actinas/metabolismo , Animais , Peso Corporal/efeitos dos fármacos , Cardiomiopatias/patologia , Cardiomiopatias/fisiopatologia , Permeabilidade da Membrana Celular/efeitos dos fármacos , Dantroleno/farmacologia , Eletrocardiografia , Coração/efeitos dos fármacos , Coração/fisiopatologia , Pulmão/efeitos dos fármacos , Pulmão/patologia , Masculino , Miocárdio/metabolismo , Miocárdio/patologia , Miocárdio/ultraestrutura , Miosinas/metabolismo , Tamanho do Órgão/efeitos dos fármacos , Ratos , Ratos Wistar , Sarcolema/efeitos dos fármacos , Sarcolema/metabolismo , Análise de Sobrevida , Fatores de TempoRESUMO
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-alpha2 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.
Assuntos
Cardiomiopatias/fisiopatologia , Distroglicanas/fisiologia , Distrofina/fisiologia , Miócitos Cardíacos/fisiologia , Sarcolema/fisiologia , Sepse/fisiopatologia , Animais , Cardiomiopatias/etiologia , Modelos Animais de Doenças , Masculino , Camundongos , Sepse/complicações , Sepse/terapiaRESUMO
OBJECTIVE: The present study describes intercalated disc remodeling under both protein expression and structural features in experimental severe sepsis induced by cecal ligation and puncture in mice. DESIGN: Controlled animal study. SETTING: University research laboratory. SUBJECTS: Male C57BL/6 mice. INTERVENTIONS: Mice were submitted to moderate and severe septic injury by cecal ligation and puncture. MEASUREMENT AND MAIN RESULTS: Severe septic injury was accompanied by a large number of bacteria in the peritoneal cavity and blood, high levels of tumor necrosis factor-alpha, and monocyte inflammatory protein-1alpha in the septic focus and serum, marked hypotension, and a high mortality rate. Western blot analysis and immunofluorescence showed a marked decrease of key gap and adherens junction proteins (connexin43 and N-cadherin, respectively) in mice submitted to severe septic injury. These changes may result in the loss of intercalated disc structural integrity, characterized in the electron microscopic study by partial separation or dehiscence of gap junctions and adherens junctions. CONCLUSIONS: Our data provide important insight regarding the alterations in intercalated disc components resulting from severe septic injury. The intercalated disc remodeling under both protein expression and structural features in experimental severe sepsis induced by cecal ligation and puncture may be partly responsible for myocardial depression in sepsis/septic shock. Although further electrophysiological studies in animals and humans are needed to determine the effect of these alterations on myocardial conduction velocity, the abnormal variables may emerge as therapeutic targets, and their modulation might provide beneficial effects on future cardiovascular outcomes and mortality in sepsis.