RESUMO
Molecular markers of dedifferentiation of dysfunctional liver sinusoidal endothelial cells (LSEC) have not been fully elucidated. We aimed at deciphering the molecular profile of dysfunctional LSEC in different pathological scenarios. Flow cytometry was used to sort CD11b-/CD32b+ and CD11b-/CD32b- LSEC from three rat models of liver disease (bile duct ligation-BDL; inhaled carbon tetrachloride-CCl4; and high fat glucose/fructose diet-HFGFD). A full proteomic profile was performed applying nano-scale liquid chromatography tandem mass spectrometry (nLC-MS) and analyzed with PEAKS software. The percentage of CD32b- LSEC varied across groups, suggesting different capillarization processes. Both CD32+ and CD32b- LSEC from models are different from control LSEC, but differently expressed proteins in CD32b- LSEC are significantly higher. Heatmaps evidenced specific protein expression patterns for each model. Analysis of biological significance comparing dysfunctional CD32b- LSEC with specialized CD32b+ LSEC from controls showed central similarities represented by 45 common down-regulated proteins involved in the suppression of the endocytic machinery and 63 common up-regulated proteins associated with the actin-dependent cytoskeleton reorganization. In summary; substantial differences but also similarities in dysfunctional LSEC from the three most common models of liver disease were found, supporting the idea that LSEC may harbor different protein expression profiles according to the etiology or disease stage.
Assuntos
Hepatopatias , Fígado , Ratos , Animais , Fígado/metabolismo , Células Endoteliais/metabolismo , Proteômica , Hepatopatias/metabolismo , Modelos TeóricosRESUMO
BACKGROUND & AIMS: Portal hypertension (PH) can be present in pre-cirrhotic stages, even in absence of fibrosis in non-alcoholic steatohepatitis (NASH) patients. Liver endothelial dysfunction (ED) has been shown as responsible for this effect in short-term dietary animal models. We evaluated the persistence of PH and underlying mechanisms in a long-term rat model of NASH. METHODS: Sprague-Dawley rats were fed 8 or 36 weeks with control diet or high-fat high-glucose/fructose diet. Metabolic parameters, histology, ED and haemodynamics were characterized. Structural characteristics of liver sections were analysed using image analysis. RESULTS: Both interventions reproduced NASH histological hallmarks (with steatosis being particularly increased at 36 weeks), but neither induced fibrosis. The 36-week intervention induced a significant increase in portal pressure (PP) compared to controls (12.1 vs 8.7 mmHg, P < .001) and the 8-week model (10.7 mmHg, P = .006), but all features of ED were normalized at 36 weeks. Image analysis revealed that the increased steatosis at 36-week was associated to an increase in hepatocyte area and a significant decrease in the sinusoidal area, which was inversely correlated with PP. The analysis provided a critical sinusoidal area above which animals were protected from developing PH and below which sinusoidal flux was compromised and PP started to increase. CONCLUSION: Liver steatosis per se (in absence of fibrosis) can induce PH through a decrease in the sinusoidal area secondary to the increase in hepatocyte area in a long-term diet-induced rat model of NASH. Image analysis of the sinusoidal area might predict the presence of PH.
Assuntos
Hipertensão Portal , Hepatopatia Gordurosa não Alcoólica , Animais , Dieta Hiperlipídica/efeitos adversos , Modelos Animais de Doenças , Humanos , Hipertensão Portal/etiologia , Fígado , Hepatopatia Gordurosa não Alcoólica/etiologia , Ratos , Ratos Sprague-DawleyRESUMO
Chronic liver disease (CLD) has no effective treatments apart from reducing its complications. Simvastatin has been tested as vasoprotective drug in experimental models of CLD showing promising results, but also limiting adverse effects. Two types of Pluronic® carriers loading simvastatin (PM108-simv and PM127-simv) as a drug delivery system were developed to avoid these toxicities while increasing the therapeutic window of simvastatin. PM127-simv showed the highest rates of cell internalization in rat liver sinusoidal endothelial cells (LSECs) and significantly lower toxicity than free simvastatin, improving cell phenotype. The in vivo biodistribution was mainly hepatic with 50% of the injected PM found in the liver. Remarkably, after one week of administration in a model of CLD, PM127-simv demonstrated superior effect than free simvastatin in reducing portal hypertension. Moreover, no signs of toxicity of PM127-simv were detected. Our results indicate that simvastatin targeted delivery to LSEC is a promising therapeutic approach for CLD.
Assuntos
Sistemas de Liberação de Medicamentos , Cirrose Hepática/tratamento farmacológico , Fígado/efeitos dos fármacos , Sinvastatina/farmacologia , Animais , Modelos Animais de Doenças , Células Endoteliais/efeitos dos fármacos , Humanos , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacologia , Cirrose Hepática/patologia , Micelas , Polímeros/química , Polímeros/farmacologia , Ratos , Sinvastatina/química , Distribuição Tecidual/efeitos dos fármacosRESUMO
UNLABELLED: Endothelial dysfunction drives vascular derangement and organ failure associated with sepsis. However, the consequences of sepsis on liver sinusoidal endothelial function are largely unknown. Statins might improve microvascular dysfunction in sepsis. The present study explores liver vascular abnormalities and the effects of statins in a rat model of endotoxemia. For this purpose, lipopolysaccharide (LPS) or saline was given to: (1) rats treated with placebo; (2) rats treated with simvastatin (25 mg/kg, orally), given at 3 and 23 hours after LPS/saline challenge; (3) rats treated with simvastatin (25 mg/kg/24 h, orally) from 3 days before LPS/saline injection. Livers were isolated and perfused and sinusoidal endothelial function was explored by testing the vasodilation of the liver circulation to increasing concentrations of acetylcholine. The phosphorylated endothelial nitric oxide synthase (PeNOS)/endothelial nitric oxide synthase (eNOS) ratio was measured as a marker of eNOS activation. LPS administration induced an increase in baseline portal perfusion pressure and a decrease in vasodilation to acetylcholine (sinusoidal endothelial dysfunction). This was associated with reduced eNOS phosphorylation and liver inflammation. Simvastatin after LPS challenge did not prevent the increase in baseline portal perfusion pressure, but attenuated the development of sinusoidal endothelial dysfunction. Treatment with simvastatin from 3 days before LPS prevented the increase in baseline perfusion pressure and totally normalized the vasodilating response of the liver vasculature to acetylcholine and reduced liver inflammation. Both protocols of treatment restored a physiologic PeNOS/eNOS ratio. CONCLUSION: LPS administration induces intrahepatic endothelial dysfunction that might be prevented by simvastatin, suggesting that statins might have potential for liver protection during endotoxemia.
Assuntos
Doença Hepática Induzida por Substâncias e Drogas/tratamento farmacológico , Endotélio Vascular/efeitos dos fármacos , Endotoxemia/tratamento farmacológico , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacologia , Sinvastatina/farmacologia , Animais , Doença Hepática Induzida por Substâncias e Drogas/fisiopatologia , Modelos Animais de Doenças , Endotélio Vascular/fisiopatologia , Endotoxemia/induzido quimicamente , Endotoxemia/fisiopatologia , Lipopolissacarídeos/toxicidade , Fígado/irrigação sanguínea , Fígado/efeitos dos fármacos , Fígado/fisiopatologia , Circulação Hepática/efeitos dos fármacos , Circulação Hepática/fisiologia , Masculino , Microcirculação/efeitos dos fármacos , Microcirculação/fisiologia , Ratos , Ratos Wistar , Resistência Vascular/efeitos dos fármacos , Resistência Vascular/fisiologiaRESUMO
Hepatic microcirculatory dysfunction due to cold storage and warm reperfusion (CS+WR) injury during liver transplantation is partly mediated by oxidative stress and may lead to graft dysfunction. This is especially relevant when steatotic donors are considered. Using primary cultured liver sinusoidal endothelial cells (LSECs), liver grafts from healthy and steatotic rats, and human liver samples, we aimed to characterize the effects of a new recombinant form of human manganese superoxide dismutase (rMnSOD) on hepatic CS+WR injury. After CS+WR, the liver endothelium exhibited accumulation of superoxide anion (O2-) and diminished levels of nitric oxide (NO); these detrimental effects were prevented by rMnSOD. CS+WR control and steatotic rat livers exhibited markedly deteriorated microcirculation and acute endothelial dysfunction, together with liver damage, inflammation, oxidative stress, and low NO. rMnSOD markedly blunted oxidative stress, which was associated with a global improvement in liver damage and microcirculatory derangements. The addition of rMnSOD to CS solution maintained its antioxidant capability, protecting rat and human liver tissues. In conclusion, rMnSOD represents a new and highly effective therapy to significantly upgrade liver procurement for transplantation.
Assuntos
Fígado/efeitos dos fármacos , Traumatismo por Reperfusão/tratamento farmacológico , Superóxido Dismutase/farmacologia , Animais , Fígado Gorduroso/terapia , Humanos , Fígado/patologia , Transplante de Fígado , Masculino , Microcirculação/efeitos dos fármacos , Óxido Nítrico/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Ratos Wistar , Proteínas Recombinantes/farmacologiaRESUMO
BACKGROUND & AIMS: Statins improve hepatic endothelial function and liver fibrosis in experimental models of cirrhosis, thus they have been proposed as therapeutic options to ameliorate portal hypertension syndrome. The transcription factor Kruppel-like factor 2 (KLF2) may be induced by statins in liver sinusoidal endothelial cells (SEC), orchestrating an efficient vasoprotective response. The present study aimed at characterizing whether KLF2 mediates statins-derived hepatic protection. METHODS: Expression of KLF2 and its vasoprotective target genes was determined in SEC freshly isolated from control or CCl(4)-cirrhotic rats treated with four different statins (atorvastatin, mevastatin, simvastatin, and lovastatin), in the presence of mevalonate (or vehicle), under static or controlled shear stress conditions. KLF2-derived vasoprotective transcriptional programs were analyzed in SEC transfected with siRNA for KLF2 or siRNA-control, and incubated with simvastatin. Paracrine effects of SEC highly-expressing KLF2 on the activation status of rat and human hepatic stellate cells (HSC) were evaluated. RESULTS: Statins administration to SEC induced significant upregulation of KLF2 expression. KLF2 upregulation was observed after 6h of treatment and was accompanied by induction of its vasoprotective programs. Simvastatin vasoprotection was inhibited in the presence of mevalonate, and was magnified in cells cultured under physiological shear stress conditions. Statin-dependent induction of vasoprotective genes was not observed when KLF2 expression was muted with siRNA. SEC overexpressing KLF2 induced quiescence of HSC through a KLF2-nitric oxide-guanylate cyclase-mediated paracrine mechanism. CONCLUSIONS: Upregulation of hepatic endothelial KLF2-derived transcriptional programs by statins confers vasoprotection and stellate cells deactivation, reinforcing the therapeutic potential of these drugs for liver diseases that course with endothelial dysfunction.
Assuntos
Células Endoteliais/metabolismo , Células Estreladas do Fígado/metabolismo , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacologia , Fatores de Transcrição Kruppel-Like/genética , Fatores de Transcrição Kruppel-Like/metabolismo , Animais , Atorvastatina , Comunicação Celular/efeitos dos fármacos , Comunicação Celular/fisiologia , Modelos Animais de Doenças , Células Endoteliais/citologia , Expressão Gênica/efeitos dos fármacos , Expressão Gênica/fisiologia , Células Estreladas do Fígado/citologia , Ácidos Heptanoicos/farmacologia , Hipertensão Portal/tratamento farmacológico , Hipertensão Portal/metabolismo , Hipertensão Portal/patologia , Fígado/citologia , Fígado/metabolismo , Circulação Hepática/efeitos dos fármacos , Circulação Hepática/fisiologia , Cirrose Hepática Experimental/tratamento farmacológico , Cirrose Hepática Experimental/metabolismo , Cirrose Hepática Experimental/patologia , Lovastatina/análogos & derivados , Lovastatina/farmacologia , Masculino , Comunicação Parácrina/efeitos dos fármacos , Comunicação Parácrina/fisiologia , Pirróis/farmacologia , Ratos , Ratos Wistar , Sinvastatina/farmacologia , Estresse MecânicoRESUMO
BACKGROUND & AIMS: Liver grafts obtained from healthy rat donors develop acute microcirculatory dysfunction due to cold-storage and warm-reperfusion injuries. These detrimental effects are avoided adding simvastatin to the cold-storage solution. Considering the importance of increasing organ donor pool for transplantation, we characterized whether simvastatin pretreatment can protect steatotic grafts from cold-storage and warm-reperfusion injuries. METHODS: Rats fed with high-fat diet received a single dose of simvastatin, or its vehicle, 30 min before liver procurement. Grafts were then cold stored for 0 h (control group) or 16 h and warm reperfused. At the end of the reperfusion period, hepatic vascular resistance, endothelial function, nitric oxide pathway, cell death, oxidative stress, autophagy, and liver injury were evaluated. Hepatic vascular resistance and endothelial function were determined in a group of simvastatin-treated livers in the presence of the nitric oxide synthase inhibitor L-NNA. RESULTS: Cold-stored rat steatotic livers exhibit increased hepatic vascular resistance and marked endothelial dysfunction, together with liver damage, oxidative stress, and low nitric oxide. Simvastatin markedly improved liver injury and prevented hepatic endothelial dysfunction. The beneficial effects of simvastatin were associated with cell death diminution, autophagy induction, and nitric oxide release. Statin-derived liver microcirculation protection was not observed when nitric oxide production was blunted. CONCLUSIONS: Pretreatment of steatotic liver donors with simvastatin shortly before procurement of the liver graft strongly protects both parenchymal and endothelial components of the liver after warm reperfusion. Our data reinforce the use of statins to protect liver grafts undergoing transplantation.
Assuntos
Fígado Gorduroso/fisiopatologia , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacologia , Transplante de Fígado , Fígado/efeitos dos fármacos , Sinvastatina/farmacologia , Animais , Criopreservação , Circulação Hepática/efeitos dos fármacos , Masculino , Preservação de Órgãos , Ratos , Ratos Sprague-DawleyRESUMO
In this study, we developed functionalized polymeric micelles (FPMs) loaded with simvastatin (FPM-Sim) as a drug delivery system to target liver sinusoidal endothelial cells (LSECs) for preserving liver function in chronic liver disease (CLD). Polymeric micelles (PMs) were functionalized by coupling peptide ligands of LSEC membrane receptors CD32b, CD36 and ITGB3. Functionalization was confirmed via spectroscopy and electron microscopy. In vitro and in vivo FPM-Sim internalization was assessed by means of flow cytometry in LSECs, hepatocytes, Kupffer and hepatic stellate cells from healthy rats. Maximum tolerated dose assays were performed in healthy mice and efficacy studies of FPM-Sim were carried out in bile duct ligation (BDL) and thioacetamide (TAA) induction rat models of cirrhosis. Functionalization with the three peptide ligands resulted in stable formulations with a greater degree of in vivo internalization in LSECs than non-functionalized PMs. Administration of FPM-Sim in BDL rats reduced toxicity relative to free simvastatin, albeit with a moderate portal-pressure-lowering effect. In a less severe model of TAA-induced cirrhosis, treatment with FPM-CD32b-Sim nanoparticles for two weeks significantly decreased portal pressure, which was associated with a reduction in liver fibrosis, lower collagen expression as well as the stimulation of nitric oxide synthesis. In conclusion, CD32b-FPM stands out as a good nanotransporter for drug delivery, targeting LSECs, key inducers of liver injury.
RESUMO
Local cancer treatment by in situ injections of thermo-responsive hydrogels (HG) offers several advantages over conventional systemic anti-cancer treatments. In this work, a biodegradable and multicompartmental HG composed of N-isopropylacrylamide, cellulose, citric acid, and ceric ammonium nitrate was developed for the controlled release of hydrophilic (doxorubicin) and hydrophobic (niclosamide) drugs. The formulation presented ideal properties regarding thermo-responsiveness, rheological behavior, drug release profile, biocompatibility, and biological activity in colon and ovarian cancer cells. Cellulose was found to retard drugs release rate, being only 4 % of doxorubicin and 30 % of niclosamide released after 1 week. This low release was sufficient to cause cell death in both cell lines. Moreover, HG demonstrated a proper injectability, in situ prevalence, and safety profile in vivo. Overall, the HG properties, together with its natural and eco-friendly composition, create a safe and efficient platform for the local treatment of non-resectable tumors or tumors requiring pre-surgical adjuvant therapy.
Assuntos
Hidrogéis , Neoplasias , Acrilamidas , Celulose/farmacologia , Doxorrubicina/química , Doxorrubicina/farmacologia , Humanos , Hidrogéis/química , Niclosamida , TemperaturaRESUMO
In non-alcoholic steatohepatitis (NASH), decreased nitric oxide and increased endothelin-1 (ET-1, also known as EDN1) released by sinusoidal endothelial cells (LSEC) induce hepatic stellate cell (HSC) contraction and contribute to portal hypertension (PH). Statins improve LSEC function, and ambrisentan is a selective endothelin-receptor-A antagonist. We aimed to analyse the combined effects of atorvastatin and ambrisentan on liver histopathology and hemodynamics, together with assessing the underlying mechanism in a rat NASH model. Diet-induced NASH rats were treated with atorvastatin (10â mg/kg/day), ambrisentan (30â mg/kg/day or 2â mg/kg/day) or a combination of both for 2â weeks. Hemodynamic parameters were registered and liver histology and serum biochemical determinations analysed. Expression of proteins were studied by immunoblotting. Conditioned media experiments were performed with LSEC. HSCs were characterized by RT-PCR, and a collagen lattice contraction assay was performed. Atorvastatin and ambrisentan act synergistically in combination to completely normalize liver hemodynamics and reverse histological NASH by 75%. Atorvastatin reversed the sinusoidal contractile phenotype, thus improving endothelial function, whereas ambrisentan prevented the contractile response in HSCs by blocking ET-1 response. Additionally, ambrisentan also increased eNOS (also known as Nos3) phosphorylation levels in LSEC, via facilitating the stimulation of endothelin-receptor-B in these cells. Furthermore, the serum alanine aminotransferase of the combined treatment group decreased to normal levels, and this group exhibited a restoration of the HSC quiescent phenotype. The combination of atorvastatin and ambrisentan remarkably improves liver histology and PH in a diet-induced NASH model. By recovering LSEC function, together with inhibiting the activation and contraction of HSC, this combined treatment may be an effective treatment for NASH patients.
Assuntos
Atorvastatina/farmacologia , Hemodinâmica , Hepatopatia Gordurosa não Alcoólica/fisiopatologia , Fenilpropionatos/farmacologia , Piridazinas/farmacologia , Alanina Transaminase/metabolismo , Animais , Biomarcadores/metabolismo , Colágeno/metabolismo , Modelos Animais de Doenças , Sinergismo Farmacológico , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/enzimologia , Células Endoteliais/patologia , Endotelina-1/farmacologia , Ativação Enzimática/efeitos dos fármacos , Hemodinâmica/efeitos dos fármacos , Células Estreladas do Fígado/efeitos dos fármacos , Células Estreladas do Fígado/metabolismo , Resistência à Insulina , Fígado/patologia , Cirrose Hepática/complicações , Cirrose Hepática/patologia , Cirrose Hepática/fisiopatologia , Óxido Nítrico Sintase Tipo III/metabolismo , Hepatopatia Gordurosa não Alcoólica/complicações , Hepatopatia Gordurosa não Alcoólica/patologia , Aumento de Peso/efeitos dos fármacosRESUMO
The liver endothelium plays a key role in the progression and resolution of liver diseases in young and adult individuals. However, its role in older people remains unknown. We have herein evaluated the importance of the sinusoidal endothelium in the pathophysiology of acute liver injury, and investigated the applicability of simvastatin, in aged animals. Eighteen-months-old male Wistar rats underwent 60 minutes of partial warm ischemia followed by 2 hours of reperfusion (WIR). A group of aged rats received simvastatin for 3 days before WIR. Endothelial phenotype, parenchymal injury, oxidative and nitrosative stress, and fenestrae dynamics were analyzed. The effects of WIR and simvastatin were investigated in primary LSEC from aged animals. The results of this study demonstrated that WIR significantly damages the liver endothelium and its effects are markedly worse in old animals. WIR-aged livers exhibited reduced vasodilation and sinusoidal capillarization, associated with liver damage and cellular stress. Simvastatin prevented the detrimental effects of WIR in aged livers. In conclusion, the liver sinusoidal endothelium of old animals is highly vulnerable to acute insult, thus targeted protection is especially relevant in preventing liver damage. Simvastatin represents a useful therapeutic strategy in aging.
Assuntos
Células Endoteliais/efeitos dos fármacos , Fígado/irrigação sanguínea , Fígado/efeitos dos fármacos , Traumatismo por Reperfusão/prevenção & controle , Sinvastatina/farmacologia , Fatores Etários , Animais , Modelos Animais de Doenças , Masculino , Óxido Nítrico/metabolismo , Fenótipo , Ratos , Ratos Wistar , Tirosina/análogos & derivados , Tirosina/metabolismoRESUMO
Non-alcoholic steatohepatitis (NASH) is a common chronic liver disorder in developed countries, with the associated clinical complications driven by portal hypertension (PH). PH may precede fibrosis development, probably due to endothelial dysfunction at early stages of the disease. Our aim was to characterize liver sinusoidal endothelial cell (LSEC) dedifferentiation/capillarization and its contribution to PH in NASH, together with assessing statins capability to revert endothelial function improving early NASH stages. Sprague-Dawley rats were fed with high fat glucose-fructose diet (HFGFD), or control diet (CD) for 8 weeks and then treated with simvastatin (sim) (10 mg·kg-1·day-1), atorvastatin (ato) (10 mg·kg-1·day-1) or vehicle during 2 weeks. Biochemical, histological and hemodynamic determinations were carried out. Sinusoidal endothelial dysfunction was assessed in individualized sorted LSEC and hepatic stellate cells (HSC) from animal groups and in whole liver samples. HFGFD rats showed full NASH features without fibrosis but with significantly increased portal pressure compared with CD rats (10.47 ± 0.37 mmHg vs 8.30 ± 0.22 mmHg; p < 0.001). Moreover, HFGFD rats showed a higher percentage of capillarized (CD32b-/CD11b-) LSEC (8% vs 1%, p = 0.005) showing a contractile phenotype associated to HSC activation. Statin treatments caused a significant portal pressure reduction (sim: 9.29 ± 0.25 mmHg, p < 0.01; ato: 8.85 ± 0.30 mmHg, p < 0.001), NASH histology reversion, along with significant recovery of LSEC differentiation and a regression of HSC activation to a more quiescent phenotype. In an early NASH model without fibrosis with PH, LSEC transition to capillarization and HSC activation are reverted by statin treatment inducing portal pressure decrease and NASH features improvement.
Assuntos
Inibidores de Hidroximetilglutaril-CoA Redutases/uso terapêutico , Hipertensão Portal/tratamento farmacológico , Fígado/patologia , Hepatopatia Gordurosa não Alcoólica/tratamento farmacológico , Animais , Dieta Hiperlipídica , Carboidratos da Dieta/administração & dosagem , Hipertensão Portal/complicações , Masculino , Hepatopatia Gordurosa não Alcoólica/complicações , Hepatopatia Gordurosa não Alcoólica/patologia , Fenótipo , Ratos , Ratos Sprague-DawleyRESUMO
In cirrhosis, liver microvascular dysfunction is a key factor increasing hepatic vascular resistance to portal blood flow, which leads to portal hypertension. De-regulated inflammatory and pro-apoptotic processes due to chronic injury play important roles in the dysfunction of liver sinusoidal cells. The present study aimed at characterizing the effects of the pan-caspase inhibitor emricasan on systemic and hepatic hemodynamics, hepatic cells phenotype, and underlying mechanisms in preclinical models of advanced chronic liver disease. We investigated the effects of 7-day emricasan on hepatic and systemic hemodynamics, liver function, hepatic microcirculatory function, inflammation, fibrosis, hepatic cells phenotype, and paracrine interactions in rats with advanced cirrhosis due to chronic CCl4 administration. The hepato-protective effects of emricasan were additionally investigated in cells isolated from human cirrhotic livers. Cirrhotic rats receiving emricasan showed significantly lower portal pressure than vehicle-treated animals with no changes in portal blood flow, indicating improved vascular resistance. Hemodynamic improvement was associated with significantly better liver function, reduced hepatic inflammation, improved phenotype of hepatocytes, liver sinusoidal endothelial cells, hepatic stellate cells and macrophages, and reduced fibrosis. In vitro experiments demonstrated that emricasan exerted its benefits directly improving hepatocytes' expression of specific markers and synthetic capacity, and ameliorated nonparenchymal cells through a paracrine mechanism mediated by small extracellular vesicles released by hepatocytes. Conclusion: This study demonstrates that emricasan improves liver sinusoidal microvascular dysfunction in cirrhosis, which leads to marked amelioration in fibrosis, portal hypertension and liver function, and therefore encourages its clinical evaluation in the treatment of advanced chronic liver disease.
RESUMO
The socioeconomic and medical improvements of the last decades have led to a relevant increase in the median age of worldwide population. Although numerous studies described the impact of aging in different organs and the systemic vasculature, relatively little is known about liver function and hepatic microcirculatory status in the elderly. In this study, we aimed at characterizing the phenotype of the aged liver in a rat model of healthy aging, particularly focusing on the microcirculatory function and the molecular status of each hepatic cell type in the sinusoid. Moreover, major findings of the study were validated in young and aged human livers. Our results demonstrate that healthy aging is associated with hepatic and sinusoidal dysfunction, with elevated hepatic vascular resistance and increased portal pressure. Underlying mechanisms of such hemodynamic disturbances included typical molecular changes in the cells of the hepatic sinusoid and deterioration in hepatocyte function. In a specific manner, liver sinusoidal endothelial cells presented a dysfunctional phenotype with diminished vasodilators synthesis, hepatic macrophages exhibited a proinflammatory state, while hepatic stellate cells spontaneously displayed an activated profile. In an important way, major changes in sinusoidal markers were confirmed in livers from aged humans. In conclusion, our study demonstrates for the first time that aging is accompanied by significant liver sinusoidal deregulation suggesting enhanced sinusoidal vulnerability to chronic or acute injuries.
Assuntos
Envelhecimento/fisiologia , Fígado/anatomia & histologia , Fígado/irrigação sanguínea , Microcirculação , Animais , Translocação Bacteriana , Células Endoteliais/patologia , Regulação da Expressão Gênica no Desenvolvimento , Hemodinâmica , Células Estreladas do Fígado/metabolismo , Células Estreladas do Fígado/ultraestrutura , Hepatócitos/metabolismo , Hepatócitos/patologia , Imunidade Inata , Inflamação/patologia , Fígado/ultraestrutura , Masculino , Modelos Animais , Fenótipo , Ratos WistarRESUMO
Hepatic stellate cells (HSC) play a key role in the development of chronic liver disease (CLD). Liraglutide, well-established in type 2 diabetes, showed anti-inflammatory and anti-oxidant properties. We evaluated the effects of liraglutide on HSC phenotype and hepatic microvascular function using diverse pre-clinical models of CLD. Human and rat HSC were in vitro treated with liraglutide, or vehicle, and their phenotype, viability and proliferation were evaluated. In addition, liraglutide or vehicle was administered to rats with CLD. Liver microvascular function, fibrosis, HSC phenotype and sinusoidal endothelial phenotype were determined. Additionally, the effects of liraglutide on HSC phenotype were analysed in human precision-cut liver slices. Liraglutide markedly improved HSC phenotype and diminished cell proliferation. Cirrhotic rats receiving liraglutide exhibited significantly improved liver microvascular function, as evidenced by lower portal pressure, improved intrahepatic vascular resistance, and marked ameliorations in fibrosis, HSC phenotype and endothelial function. The anti-fibrotic effects of liraglutide were confirmed in human liver tissue and, although requiring further investigation, its underlying molecular mechanisms suggested a GLP1-R-independent and NF-κB-Sox9-dependent one. This study demonstrates for the first time that liraglutide improves the liver sinusoidal milieu in pre-clinical models of cirrhosis, encouraging its clinical evaluation in the treatment of chronic liver disease.
Assuntos
Células Estreladas do Fígado/efeitos dos fármacos , Incretinas/farmacologia , Liraglutida/farmacologia , Cirrose Hepática/tratamento farmacológico , Animais , Proliferação de Células/efeitos dos fármacos , Modelos Animais de Doenças , Células Estreladas do Fígado/metabolismo , Humanos , Fígado/irrigação sanguínea , Cirrose Hepática/fisiopatologia , Masculino , Microvasos/efeitos dos fármacos , Ratos WistarRESUMO
Warm ischemia and reperfusion (WIR) causes hepatic damage and may lead to liver failure, however the mechanisms involved are largely unknown. Here we have characterized the microcirculatory status and endothelial phenotype of livers undergoing WIR, and evaluated the use of simvastatin in WIR injury prevention. Male Wistar rats received simvastatin, or vehicle, 30 min before undergoing 60 min of partial warm ischemia (70%) followed by 2 h or 24 h of reperfusion. Hepatic and systemic hemodynamics, liver injury (AST, ALT, LDH), endothelial function (vasodilatation in response to acetylcholine), KLF2 and nitric oxide pathways, oxidative stress, inflammation (neutrophil and macrophage infiltration) and cell death were evaluated. Profound microcirculatory dysfunction occurred rapidly following WIR. This was evidenced by down-regulation of the KLF2 vasoprotective pathway, impaired vasodilatory capability and endothelial activation, altogether leading to increased hepatic vascular resistance and liver inflammation, with significant leukocyte infiltration, oxidative stress and cell death. Simvastatin preserved the hepatic endothelial phenotype, and blunted the detrimental effects of WIR on liver hemodynamics and organ integrity. In conclusion, WIR-induced injury to liver sinusoidal endothelial cells is mitigated by pre-treatment with Simvastatin probably through a KLF2-dependent mechanism.
Assuntos
Fígado/irrigação sanguínea , Traumatismo por Reperfusão/prevenção & controle , Sinvastatina/farmacologia , Isquemia Quente/efeitos adversos , Animais , Células Cultivadas , Avaliação Pré-Clínica de Medicamentos , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Fatores de Transcrição Kruppel-Like/metabolismo , Células de Kupffer/efeitos dos fármacos , Células de Kupffer/metabolismo , Fígado/efeitos dos fármacos , Fígado/imunologia , Fígado/patologia , Masculino , Microcirculação , Infiltração de Neutrófilos , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo , Ratos Wistar , Sinvastatina/uso terapêuticoRESUMO
INTRODUCTION: Autophagy is a cellular process essential for survival and homeostasis that confers cellular protection toward a wide range of deleterious stimuli. It has a highly complex regulation with several autophagic proteins also belonging to other main signaling pathways as cell proliferation or apoptosis. In addition, autophagy has an important role in cell metabolism. Interest in the study of this process is rapidly rising and, in the past few years, autophagy has been implicated in a variety of hepatic diseases. AREAS COVERED: The review covers the research and investigational use of pharmacological strategies that modify autophagy in the treatment of liver diseases. Autophagy modulation in steatosis, steatohepatitis, viral hepatitis, fibrogenesis, cirrhosis, hepatocellular carcinoma and ischemia/reperfusion injury will be described, critically analyzed and discussed. Papers included in the present manuscript were selected from the PubMed search: liver + (macro)autophagy + each of the pathologies described above. EXPERT OPINION: The complexity of autophagy creates significant controversy on the potential of its pharmacological modulation. A major requirement for drugs regulating autophagy in the treatment of liver diseases is that these should be liver-specific; moreover, they should primarily target one specific hepatic cell type.