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BACKGROUND AND AIMS: Accurate risk stratification for hepatocellular carcinoma (HCC) after achieving a sustained viral response (SVR) is necessary for optimal surveillance. We aimed to develop and validate a machine learning (ML) model to predict the risk of HCC after achieving an SVR in individual patients. METHODS: In this multicenter cohort study, 1742 patients with chronic hepatitis C who achieved an SVR were enrolled. Five ML models were developed including DeepSurv, gradient boosting survival analysis, random survival forest (RSF), survival support vector machine, and a conventional Cox proportional hazard model. Model performance was evaluated using Harrel' c-index and was externally validated in an independent cohort (977 patients). RESULTS: During the mean observation period of 5.4 years, 122 patients developed HCC (83 in the derivation cohort and 39 in the external validation cohort). The RSF model showed the best discrimination ability using seven parameters at the achievement of an SVR with a c-index of 0.839 in the external validation cohort and a high discriminative ability when the patients were categorized into three risk groups (P <0.001). Furthermore, this RSF model enabled the generation of an individualized predictive curve for HCC occurrence for each patient with an app available online. CONCLUSIONS: We developed and externally validated an RSF model with good predictive performance for the risk of HCC after an SVR. The application of this novel model is available on the website. This model could provide the data to consider an effective surveillance method. Further studies are needed to make recommendations for surveillance policies tailored to the medical situation in each country. IMPACT AND IMPLICATIONS: A novel prediction model for HCC occurrence in patients after hepatitis C virus eradication was developed using machine learning algorithms. This model, using seven commonly measured parameters, has been shown to have a good predictive ability for HCC development and could provide a personalized surveillance system.
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BACKGROUND AND AIMS: It remains unclear whether obesity increases the risk of hepatocellular carcinoma (HCC) in patients with chronic hepatitis C who achieved a sustained virological response (SVR) with antiviral therapy. METHODS: In this multicenter cohort study, we enrolled patients with chronic hepatitis C who achieved SVR with interferon (IFN)-based therapy (IFN group) or direct-acting antiviral (DAA) therapy (DAA group) between January 1, 1990, and December 31, 2018. The patients underwent regular surveillance for HCC. Cumulative incidence of and the risk factors for HCC development after SVR were assessed using the Kaplan-Meier method and Cox proportional hazard regression analysis, respectively. RESULTS: Among 2,055 patients (840 in the IFN group and 1,215 in the DAA group), 75 developed HCC (41 in the IFN group and 34 in the DAA group) during the mean observation period of 4.1 years. The incidence rates of HCC at 1, 2, and 3 years were 1.2, 1.9, and 3.0%, respectively. Multivariate analysis revealed that in addition to older age, lower albumin level, lower platelet count, higher alpha-fetoprotein level, and absence of dyslipidemia, obesity (body mass index ≥25 kg/m2) and heavy alcohol consumption (≥60 g/day) were independent risk factors for HCC development, with adjusted hazard ratio (HR) of 2.53 (95% confidence interval [CI]: 1.51-4.25) and 2.56 (95% CI: 1.14-5.75), respectively. The adjusted HR was not significant between the 2 groups (DAA vs. IFN; HR 1.19, 95% CI: 0.61-2.33). CONCLUSIONS: Obesity and heavy alcohol consumption increased the risk of HCC development after SVR.
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Use of herbal remedies in the treatment of various diseases has a long tradition in Eastern medicine and the liver diseases are not an exception. In their use, lack of elucidation of mechanism(s) as well as randomized, placebo-controlled clinical trials has been a problem. Recently, we and others reported that inchin-ko-to (TJ-135), one of herbal remedies, suppressed hepatic fibrosis in animal models. In the course of clarifying the mechanism, we directed our focus on hepatic stellate cells (HSCs), playing a pivotal role in hepatic fibrosis, and found that rat HSCs cultured with TJ-135 changed their morphology to star-like configuration with thin, slender and dendritic processes with fewer stress fibers, which might be the features in apoptosis. In fact, TJ-135 induced HSC apoptosis in a time- and concentration-dependent manner as judged by the nuclear morphology, quantitation of cytoplasmic histone-associated DNA oligonucleosome fragments and caspase 3 activity. In HSCs treated with TJ-135, increased expression of p53 and decreased expression of Bcl-2 and phosphorylated Akt and Bad were determined. HSC apoptosis is shown to be involved in the mechanisms of spontaneous resolution of rat hepatic fibrosis and the agent which induces HSC apoptosis has been shown to reduce experimental hepatic fibrosis in rats. Thus, the induction of HSC apoptosis could be the mechanism how TJ-135 works on the resolution of hepatic fibrosis. Our current data may shed light on the novel effect of the herbal remedy.
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Apoptose/efeitos dos fármacos , Medicamentos de Ervas Chinesas/farmacologia , Hepatócitos/efeitos dos fármacos , Animais , Caspase 3 , Caspases/metabolismo , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Núcleo Celular/patologia , Células Cultivadas , Citoesqueleto/efeitos dos fármacos , Citoesqueleto/patologia , Ensaio de Imunoadsorção Enzimática , Hepatócitos/citologia , Hepatócitos/metabolismo , Immunoblotting , Marcação In Situ das Extremidades Cortadas , Masculino , Proteínas Nucleares/metabolismo , Ratos , Ratos Sprague-DawleyRESUMO
BACKGROUND: Rapamycin is a specific inhibitor of the mammalian target of rapamycin (mTOR). The effect of rapamycin on proliferation and cellular function was studied in hepatocytes stimulated by hepatocyte growth factor (HGF) or transforming growth factor-alpha (TGFalpha). METHODS AND RESULTS: When isolated rat hepatocytes were cultured at low density, the addition of HGF or TGFalpha increased DNA synthesis but did not affect albumin or fibrinogen concentrations in the medium. In contrast, in hepatocytes cultured at high density, the albumin and fibrinogen concentrations, but not DNA synthesis, were increased by HGF or TGFalpha. The HGF- or TGFalpha-induced increase in DNA synthesis and in albumin or fibrinogen concentrations was suppressed by the addition of rapamycin, as well as wortmannin, a phosphatidylinositol-3 kinase inhibitor. CONCLUSION: HGF and TGFalpha stimulate proliferation and function of hepatocytes depending upon the conditions, and rapamycin inhibited these stimulatory effects, possibly by inhibiting the mTOR-dependent signaling pathway.
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Antibióticos Antineoplásicos/farmacologia , Fator de Crescimento de Hepatócito/antagonistas & inibidores , Hepatócitos/efeitos dos fármacos , Sirolimo/farmacologia , Fator de Crescimento Transformador alfa/antagonistas & inibidores , Albuminas/análise , Albuminas/metabolismo , Androstadienos/farmacologia , Animais , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Meios de Cultura/química , Replicação do DNA/efeitos dos fármacos , Fibrinogênio/análise , Fibrinogênio/metabolismo , Fator de Crescimento de Hepatócito/farmacologia , Hepatócitos/metabolismo , Proteínas Quinases/metabolismo , Ratos , Transdução de Sinais/efeitos dos fármacos , Serina-Treonina Quinases TOR , Fator de Crescimento Transformador alfa/farmacologia , WortmaninaRESUMO
Brief periods of tissue ischemia produced tissue resistance to prolonged ischemia and reperfusion, a phenomenon called ischemic preconditioning. The mechanisms of ischemic preconditioning were examined in a rat warm ischemia-reperfusion model as well as the effect of ischemic preconditioning on liver regeneration. Ischemic preconditioning decreased liver injury after warm ischemia-reperfusion, which was reversed by Kupffer cell depletion. Ischemic preconditioning stimulated Kupffer cells to produce reactive oxygen species. Scavengers of reactive oxygen species reversed the effect of ischemic preconditioning, and pretreatment with sublethal dose of hydrogen peroxide mimicked ischemic preconditioning effect. Rat livers were preconditioned by ischemia and subjected to 70% partial hepatectomy. Liver regeneration was then evaluated serially. Ischemic preconditioning promoted liver regeneration, which was reversed by adenosine A2 receptor antagonism and mimicked by adenosine A2 receptor agonism. Promotion of liver regeneration by ischemic preconditioning and adenosine A2 receptor agonism were reversed by Kupffer cell depletion. In conclusion, ischemic preconditioning stimulates Kupffer cells to produce reactive oxygen species, leading to hepatocyte protection against warm ischemia-reperfusion injury; and ischemic preconditioning promoted liver regeneration via adenosine A2 receptor pathway in Kupffer cells.
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Precondicionamento Isquêmico/métodos , Fígado/fisiopatologia , Traumatismo por Reperfusão/prevenção & controle , Animais , Modelos Animais de Doenças , Hepatectomia , Peróxido de Hidrogênio/farmacologia , Células de Kupffer/metabolismo , Regeneração Hepática , Ratos , Espécies Reativas de Oxigênio/metabolismo , Receptores Purinérgicos P1/metabolismoRESUMO
Branched-chain amino acids (BCAAs) modulate various cellular functions, in addition to providing substrates for the production of proteins. In this study, we examined the effect of BCAAs on the secretion of hepatocyte growth factor (HGF) by hepatic stellate cells. A hepatic stellate cell clone was cultured in medium supplemented with various concentrations of valine, leucine, or isoleucine. Of these BCAAs, leucine markedly induced an increase in the levels of HGF in the medium in a dose-dependent manner. The addition of valine or isoleucine had no significant effect on HGF levels in the medium. The difference in levels of HGF in the medium between leucine-treated and non-treated cells was enhanced by the incubation period. These results demonstrate that, among BCAAs, leucine stimulates the secretion of HGF by cultured hepatic stellate cells.
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Fator de Crescimento de Hepatócito/metabolismo , Leucina/química , Animais , Meios de Cultura/farmacologia , Relação Dose-Resposta a Droga , Ensaio de Imunoadsorção Enzimática , Isoleucina/metabolismo , Leucina/farmacologia , Leucina/fisiologia , Fígado/citologia , Modelos Estatísticos , Ratos , Fatores de Tempo , Valina/metabolismoRESUMO
BACKGROUND AND AIMS: Hepatic ischemic preconditioning decreases sinusoidal endothelial cell injury and Kupffer cell activation after cold ischemia/reperfusion, leading to improved survival of liver transplant recipients in rats. Ischemic preconditioning also protects livers against warm ischemia/reperfusion injury, in which hepatocyte injury is remarkable. We aimed to determine whether ischemic preconditioning directly protects hepatocytes and to elucidate its mechanisms. METHODS: Rats were injected with gadolinium chloride to deplete Kupffer cells or with N -acetyl- l -cysteine, superoxide dismutase, or catalase to scavenge reactive oxygen species. Livers were then preconditioned by 10 minutes of ischemia and 10 minutes of reperfusion. Subsequently, livers were subjected to 40 minutes of warm ischemia and 60 minutes of reperfusion in vivo or in a liver perfusion system. In other rats, livers were preconditioned by H(2)O(2) perfusion instead of ischemia. In the other experiments, livers were perfused with nitro blue tetrazolium to detect reactive oxygen species formation. RESULTS: Ischemic preconditioning decreased injury in hepatocytes, but not in sinusoidal endothelial cells. Kupffer cell depletion itself did not change hepatocyte injury after ischemia/reperfusion, indicating no contribution of Kupffer cells to ischemia/reperfusion injury. However, Kupffer cell depletion reversed hepatoprotection by ischemic preconditioning. Reactive oxygen species formation occurred in Kupffer cells after ischemic preconditioning. Scavenging of reactive oxygen species reversed the effect of ischemic preconditioning, and H(2)O(2) preconditioning mimicked ischemic preconditioning. CONCLUSIONS: Ischemic preconditioning directly protected hepatocytes after warm ischemia/reperfusion, which is not via suppression of changes in sinusoidal cells as in cold ischemia/reperfusion injury. This hepatocyte protection was mediated by reactive oxygen species produced by Kupffer cells.
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Hepatócitos/citologia , Precondicionamento Isquêmico , Células de Kupffer/citologia , Espécies Reativas de Oxigênio/metabolismo , Acetilcisteína/farmacologia , Animais , Catalase/farmacologia , Sequestradores de Radicais Livres/farmacologia , Gadolínio/farmacologia , Hepatócitos/efeitos dos fármacos , Hepatócitos/fisiologia , Peróxido de Hidrogênio/farmacologia , Células de Kupffer/efeitos dos fármacos , Células de Kupffer/fisiologia , Masculino , Ratos , Ratos Sprague-Dawley , Superóxido Dismutase/farmacologiaRESUMO
Hepatocyte growth factor (HGF) has pleiotropic effects. Up-regulation of HGF activity in vivo may be beneficial. Branched-chain amino acids (BCAAs) are known to modulate various cellular functions. When starved rats received intraperitoneal injections of valine, leucine or isoleucine, only leucine treatment increased both hepatic and circulating levels of HGF in a dose-dependent manner, up to 1.5 and 2.3 times higher, respectively, than in controls. When young growing rats with free access to food were injected with leucine once a day for a week, HGF levels and liver weights were significantly higher than those of control rats. Furthermore, 1 week of leucine treatment of adult rats resulted in elevated serum albumin levels with an increase in HGF levels. Taken together with our previous report showing that leucine stimulates HGF production by hepatic stellate cells in culture, leucine, among BCAAs, may induce an increase in HGF production by the liver in vivo.
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Fator de Crescimento de Hepatócito/biossíntese , Leucina/farmacologia , Animais , Fator de Crescimento de Hepatócito/sangue , Injeções Intraperitoneais , Isoleucina/administração & dosagem , Isoleucina/farmacologia , Cinética , Leucina/administração & dosagem , Fígado/efeitos dos fármacos , Fígado/metabolismo , Masculino , Tamanho do Órgão/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Albumina Sérica/metabolismo , Inanição , Valina/administração & dosagem , Valina/farmacologiaRESUMO
Hepatic stellate cells (HSCs) play a central role in the development of hepatic fibrosis. Recent evidence has revealed that HSCs also play a role in its resolution, where HSC apoptosis was determined. Moreover, induction of HSC apoptosis caused a reduction of experimental hepatic fibrosis in rats. Thus knowing the mechanism of HSC apoptosis might be important to clarify the pathophysiology and establish the therapeutic strategy for hepatic fibrosis. In HSCs, Rho and Rho kinase are known to regulate contraction, migration, and proliferation with modulation of cell morphology. Controversy exists as to the participation of Rho and Rho kinase on cell survival, and little is known regarding this matter in HSCs. In this study, we directed our focus on the role of the Rho pathway in the regulation of HSC survival. C3, an inhibitor of Rho, increased histone-associated DNA fragmentation and caspase 3 activity with enhanced condensation of nuclear chromatin in rat cultured HSCs. Moreover, Y-27632, an inhibitor of Rho kinase, had the same effects, suggesting that inhibition of the Rho/Rho kinase pathway causes HSC apoptosis. On the other hand, lysophosphatidic acid, which stimulates the Rho/Rho kinase pathway, decreased histone-associated DNA fragmentation in HSCs. Inhibition of the Rho/Rho kinase pathway did not affect p53, Bcl-2, or Bax levels in HSCs. Thus we concluded that the Rho/Rho kinase pathway may play a role in the regulation of HSC survival.
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Apoptose/fisiologia , Hepatócitos/fisiologia , Proteínas Serina-Treonina Quinases/fisiologia , Proteínas rho de Ligação ao GTP/fisiologia , ADP Ribose Transferases/farmacologia , Amidas/farmacologia , Animais , Apoptose/efeitos dos fármacos , Sangue , Toxinas Botulínicas/farmacologia , Células Cultivadas , Meios de Cultura Livres de Soro , Inibidores Enzimáticos/farmacologia , Hepatócitos/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular , Lisofosfolipídeos/farmacologia , Masculino , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Piridinas/farmacologia , Ratos , Ratos Sprague-Dawley , Proteína Supressora de Tumor p53/metabolismo , Proteína X Associada a bcl-2 , Proteínas rho de Ligação ao GTP/antagonistas & inibidores , Quinases Associadas a rhoRESUMO
Although structural changes are most important to determine vascular resistance in portal hypertension, vasoactive mediators also contribute to its regulation. Hepatic stellate cells (HSCs) are assumed to play a role in modulating intrahepatic vascular resistance based on their residence in the space of Disse and capacity to contract. Because sphingosine 1-phosphate (S1P) has been shown to stimulate HSC contractility, we wondered if S1P could regulate portal pressure. S1P at 0.5-5 microM increased portal pressure in isolated rat perfused liver. This effect was abrogated in the presence of a binding antagonist for S1P2, JTE-013. Perfusion of isolated rat liver with 5 microM S1P increased Rho activity in the liver, and co-perfusion with JTE-013 cancelled S1P-induced Rho activation. Because S1P is present in human plasma at approximately 0.2 microM, S1P might readily regulate portal vascular tone in physiological and pathological status. The antagonist for S1P2 merits consideration for treatment of portal hypertension.
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Hepatócitos/fisiologia , Fígado/fisiologia , Lisofosfolipídeos/farmacologia , Pressão na Veia Porta/efeitos dos fármacos , Receptores de Superfície Celular/metabolismo , Esfingosina/farmacologia , Proteínas rho de Ligação ao GTP/metabolismo , Animais , Células Cultivadas , Técnicas de Cultura , Relação Dose-Resposta a Droga , Hepatócitos/efeitos dos fármacos , Fígado/efeitos dos fármacos , Masculino , Taxa de Depuração Metabólica/efeitos dos fármacos , Pressão na Veia Porta/fisiologia , Pirazóis/farmacologia , Piridinas/farmacologia , Ratos , Ratos Sprague-Dawley , Receptores Acoplados a Proteínas G , Receptores de Lisoesfingolipídeo , Esfingosina/análogos & derivados , Receptores de Esfingosina-1-FosfatoRESUMO
BACKGROUND & AIMS: Sphingosine 1-phosphate (S1P), a ligand for G protein-coupled endothelial differentiation gene-1 (Edg-1), Edg-3, Edg-5, Edg-6, and Edg-8, elicits a variety of responses by cells. Prominent among these is cell proliferation. S1P is abundantly stored in platelets and released upon their activation, suggesting that S1P plays a pathophysiologic role in vivo. Because the major part of injected S1P was distributed into the liver in mice, we wondered whether the liver would be one of its targets. The effects of S1P on hepatocytes, the major constituent cells in the liver, were examined. METHODS & RESULTS: Northern blot analysis revealed the expression of Edg-1 and Edg-5 messenger RNA (mRNA) in cultured rat hepatocytes, in which S1P decreased DNA synthesis induced by hepatocyte growth factor (HGF) or epidermal growth factor (EGF) without affecting total protein synthesis. This inhibitory effect was attenuated by inactivation of small GTPase Rho with C3 exotoxin but not by inactivation of G(i) with pertussis toxin. Moreover, in the presence of JTE-013, a newly developed and specific binding antagonist for Edg-5, the inhibitory effect was also cancelled. Finally, the administration of S1P after 70% partial hepatectomy in rats reduced the peak of DNA synthesis in hepatocytes with increased Rho activity. Furthermore, Edg-5 but not Edg-1 mRNA expression was enhanced in hepatocytes 24-72 hours after partial hepatectomy, which coincides with decreasing hepatocyte proliferation. CONCLUSIONS: S1P has an antiproliferative property in rat hepatocytes by activating Rho via Edg-5. Our results raise the possibility that S1P is a negative regulator in liver regeneration.
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Hepatócitos/citologia , Lisofosfolipídeos , Receptores de Superfície Celular/fisiologia , Receptores Acoplados a Proteínas G , Esfingosina/análogos & derivados , Esfingosina/fisiologia , Proteínas rho de Ligação ao GTP/fisiologia , ADP Ribose Transferases/farmacologia , Animais , Toxinas Botulínicas/farmacologia , Células CHO , Divisão Celular/efeitos dos fármacos , Divisão Celular/fisiologia , Células Cultivadas , Cricetinae , DNA/antagonistas & inibidores , DNA/biossíntese , Hepatócitos/metabolismo , Fígado/metabolismo , Regeneração Hepática/fisiologia , Masculino , Toxina Pertussis/farmacologia , Pirazóis/farmacologia , Piridinas/farmacologia , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Receptores de Superfície Celular/genética , Receptores de Lisofosfolipídeos , Esfingosina/farmacologiaRESUMO
Using a rat myofibroblast-like hepatic stellate cell line, we studied the actomyosin-based cytoskeletal actions mediated by Rho-kinase and/or myosin light chain kinase (MLCK). Calmodulin/MLCK inhibitors W-7 and ML-7 attenuated cell migration dose-relatedly at concentrations from 10(-6) to 10(-4)M and collagen gel-contraction by the cells at 10(-4)M, respectively. Rho-kinase inhibitors Y-27632 and HA1077 attenuated the gel-contraction at concentrations from 10(-6) to 10(-4) M, respectively. These Rho-kinase inhibitors attenuated cell migration at 10(-7)M but enhanced the migration at 10(-4)M, respectively. They altered cell morphology showing prominent peripheral actin bundles and sparse central stress fibers, in comparison with the calmodulin/MLCK inhibitors. Both ML-7 and Y-27632 attenuated phosphorylation of myosin regulatory light chain and cell attachment to extracellular substrate. ML-7 attenuated the activation of GTP-binding protein Rac, while Y-27632 did not. These findings suggest that the actomyosin-based cytoskeletal actions can be functionally diverse depending on the Rho-kinase-mediated pathway and the MLCK-mediated pathway.