RESUMEN
The liver is the only visceral organ that exhibits a remarkable capability of regenerating in response to partial hepatectomy (PH) or chemical injury. Improving liver regeneration (LR) ability is the basis for the favourable treatment outcome of patients after PH, which can serve as a potential indicator for postoperative survival. The present study aimed to investigate the protective effects of Yiqi Huoxue recipe (YQHX) on LR after PH in rats and further elucidate its underlying mechanism. A two-thirds PH rat model was used in this study. Wistar rats were randomly divided into four groups: sham-operated, PH, YQHX + PH, and Fuzheng Huayu decoction (FZHY) + PH groups. All rats were sacrificed under anesthesia at 24 and 72 h after surgery. The rates of LR were calculated, and the expression levels of cyclin D1 and c-jun were determined by immunohistochemical staining. The protein levels of p-JNK1/2, JNK1/2, p-c-jun, c-jun, Bax, and Bcl-2 were detected by Western blotting, while the mRNA levels of JNK1, JNK2, c-jun, Bax, and Bcl-2 were examined by real-time polymerase chain reaction (RT-PCR). At the corresponding time points, YQHX and FZHY administration dramatically induced the protein levels of p-JNK1/2 compared to the PH group (p < 0.05), while FZHY + PH group showed prominently increase in p-JNK1/2 protein levels compared to the YQHX + PH group (p < 0.05). A similar trend was observed for the expression levels of p-c-jun. Compared to the PH group, YQHX and FZHY markedly reduced the mRNA and protein expression levels of Bax at 24 h after PH, while those in the FZHY + PH group decreased more obviously (p < 0.05). Besides, in comparison with the PH group, YQHX and FZHY administration predominantly upregulated the mRNA and protein expression levels of Bcl-2 at 24 and 72 h after PH (p < 0.05). In conclusion, YQHX improves LR in rats after PH by inhibiting hepatocyte apoptosis via the JNK signaling pathway.
RESUMEN
OBJECTIVE: To determine the role of IGF-1/PI3K pathway and investigate the molecular mechanism of Fuzhenghuayu (FZHY) therapy in a spontaneous recovery rat model of liver fibrosis. METHODS: The liver fibrosis model was induced in male Wistar rats by administering 8 weeks of twice weekly CCL4 intraperitoneal injections without (untreated model) or with once daily FZHY (treated model). Normal, untreated rats served as the control group. At weeks 4, 6 and 8 (fibrosis) and 10, 12 and 14 (spontaneous recovery) after modeling initiation, effects on protein (a-SMA, IGF-1, PI3K) and mRNA (IGF-1, PI3K) expression levels were evaluated by immunohistochemistry and RT-PCR, respectively. Serum markers of liver function (alanine aminotransferase (ALT) and aspartate aminotransferase (AST)) and liver cell damage (alkaline hydrolysis, HYP) were measured. Histology was performed to assess the degree of inflammation and fibrosis (Ishak scoring system). RESULTS: In the untreated model group, progression of liver fibrosis (weeks 4, 6 and 8) was accompanied by gradual increases in inflammation, necrosis, serum ALT and AST, and hepatic expression of a-SMA protein and IGF-1 and PI3K protein and mRNA; however, during the spontaneous recovery period (weeks 10, 12 and 14) the IGF-1 and PI3K protein and mRNA levels rapidly decreased and the HYP level, Ishak score, and a-SMA hepatic expression also decreased. The FZHY-treated model group showed significantly lower fibrosis-related up-regulation of IGF-1 and PI3K protein and mRNA expression, HYP level, Ishak score, and a-SMA hepatic expression at each time point (vs. untreated model group). CONCLUSION: The IGF-1/PI3K pathway may contribute to progression of liver fibrosis. The mechanism by which FZHY prevents liver fibrosis in a rat model may involve blocking of the IGF/PI3K pathway and inhibiting HSC activation.
Asunto(s)
Medicamentos Herbarios Chinos/farmacología , Cirrosis Hepática Experimental/metabolismo , Hígado/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Animales , Factor I del Crecimiento Similar a la Insulina/metabolismo , Hígado/metabolismo , Hígado/patología , Cirrosis Hepática Experimental/patología , Masculino , Fosfatidilinositol 3-Quinasas/metabolismo , Ratas , Ratas WistarRESUMEN
BACKGROUND: Fuzheng Huayu recipe (FZHY), a compound of Chinese herbal medicine, was reported to improve liver function and fibrosis in patients with hepatitis B virus infection. However, its effect on nutritional fibrosing steatohepatitis is unclear. We aimed to elucidate the role and molecular mechanism of FZHY on this disorder in mice. METHODS: C57BL/6 J mice were fed with methionine-choline deficient (MCD) diet for 8 weeks to induce fibrosing steatohepatitis. FZHY and/or heme oxygenase-1 (HO-1) chemical inducer (hemin) were administered to mice, respectively. The effect of FZHY was assessed by comparing the severity of hepatic injury, levels of hepatic lipid peroxides, activation of hepatic stellate cells (HSCs) and the expression of oxidative stress, inflammatory and fibrogenic related genes. RESULTS: Mice fed with MCD diet for 8 weeks showed severe hepatic injury including hepatic steatosis, necro-inflammation and fibrosis. Administration of FZHY or hemin significantly lowered serum levels of alanine aminotransferase, aspartate aminotransferase, reduced hepatic oxidative stress and ameliorated hepatic inflammation and fibrosis. An additive effect was observed in mice fed MCD supplemented with FZHY or/and hemin. These effects were associated with down-regulation of pro-oxidative stress gene cytochrome P450 2E1, up-regulation of anti-oxidative gene HO-1; suppression of pro-inflammation genes tumor necrosis factor alpha and interleukin-6; and inhibition of pro-fibrotic genes including α-smooth muscle actin, transforming growth factor beta 1, collagen type I (Col-1) and Col-3. CONCLUSIONS: Our study demonstrated the protective role of FZHY in ameliorating nutritional fibrosing steatohepatitis. The effect was mediated through regulating key genes related to oxidative stress, inflammation and fibrogenesis.
Asunto(s)
Antioxidantes/uso terapéutico , Medicamentos Herbarios Chinos/uso terapéutico , Hígado Graso/prevención & control , Hígado/efectos de los fármacos , Actinas/genética , Actinas/metabolismo , Animales , Citocromo P-450 CYP2E1/genética , Citocromo P-450 CYP2E1/metabolismo , Citocinas/genética , Citocinas/metabolismo , Dieta/efectos adversos , Quimioterapia Combinada , Inhibidores Enzimáticos/uso terapéutico , Hígado Graso/etiología , Hígado Graso/metabolismo , Hígado Graso/patología , Fibrosis , Regulación de la Expresión Génica/efectos de los fármacos , Hemo-Oxigenasa 1/antagonistas & inhibidores , Hemo-Oxigenasa 1/genética , Hemo-Oxigenasa 1/metabolismo , Hemina/uso terapéutico , Células Estrelladas Hepáticas/efectos de los fármacos , Células Estrelladas Hepáticas/metabolismo , Células Estrelladas Hepáticas/patología , Hígado/metabolismo , Hígado/patología , Hígado/fisiopatología , Masculino , Proteínas de la Membrana/antagonistas & inhibidores , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Ratones , Ratones Endogámicos C57BL , Necrosis , ARN Mensajero/metabolismo , Distribución AleatoriaRESUMEN
OBJECTIVE: The pathogenesis of non-alcoholic steatohepatitis is still unclear. We have demonstrated previously that peroxisome proliferator activated receptor gamma (PPARγ) ligand protects against inflammation and fibrogenesis in experimental non-alcoholic steatohepatitis. We aim to elucidate the effect and the mechanism of PPARγ itself on nutritional fibrotic steatohepatitis in mice. METHODS: C57BL/6J mice were fed with methionine-choline deficient (MCD) diet for 8 weeks to induce fibrotic steatohepatitis. Mice fed the MCD diet were treated with adenovirus carrying PPARγ (Ad-PPARγ), Ad-PPARγ plus PPARγ agonist rosiglitazone, or PPARγ antagonist 2-chloro-5-nitrobenzaniliden (GW9662), respectively. The effects of up-regulation of PPARγ in the presence or absence of its agonist/or antagonist were assessed by comparing the severity of hepatic injury, activation of hepatic stellate cells and the expression of adiponectin, heme oxygenase-1, and fibrogenic related genes. RESULTS: Mice fed with MCD diet for 8 weeks showed severe hepatic injury including hepatic steatosis, inflammatory infiltration, and fibrosis. Administration of Ad-PPARγ significantly lowered serum alanine aminotransferase level and ameliorated hepatic steatosis, necroinflammation, and fibrosis. These effects were associated with enhanced expression of PPARγ, up-regulated expression of adiponectin and heme oxygenase-1, and down-regulated expression of tumor necrosis factor alpha, interleukin-6, α-smooth muscle actin, transforming growth factor beta 1, matrix metallopeptidase-2, and -9. Administration of GW9662 promoted the severity of liver histology. CONCLUSIONS: The present study provided evidences for the protective role of overexpressing PPARγ in ameliorating hepatic fibrosing steatohepatitis in mice. Modulation of PPARγ expression might serve as a therapeutic approach for fibrotic steatohepatitis.
Asunto(s)
Hígado Graso/prevención & control , Vectores Genéticos/administración & dosificación , PPAR gamma/biosíntesis , PPAR gamma/uso terapéutico , Adenoviridae/genética , Anilidas/administración & dosificación , Animales , Colina , Dieta , Hígado Graso/etiología , Hígado Graso/genética , Hígado Graso/metabolismo , Inflamación/genética , Inflamación/fisiopatología , Cirrosis Hepática/etiología , Cirrosis Hepática/genética , Cirrosis Hepática/metabolismo , Cirrosis Hepática/prevención & control , Cirrosis Hepática Experimental , Masculino , Metionina/deficiencia , Ratones , Ratones Endogámicos C57BL , PPAR gamma/administración & dosificación , PPAR gamma/genética , Distribución Aleatoria , Rosiglitazona , Tiazolidinedionas/administración & dosificación , Transfección , beta-Galactosidasa/administración & dosificación , beta-Galactosidasa/genéticaRESUMEN
OBJECTIVE: To investigate the potential role of heme oxygenase-1 on preventing non-alcoholic steatohepatitis (NASH) in mice. METHODS: Experimental models of NASH were established by feeding male C57BL/6J mice with choline-methionine deficient diet (MCD) for four weeks. Control animals were fed with choline-methionine supplemented diet. The treatment groups were fed with MCD diet combined with HO-1 inducer hemin or inhibitor zinc protoporphyrin IX (ZnPP-IX). Serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were tested by enzymic method with automatic biochemistry analyzer. The degree of hepatic steatosis, inflammation and fibrosis were examined under HE staining. The hepatic mRNA and protein expressions of HO-1, TNFalpha and IL-6 were analyzed by RT-PCR and Western blot respectively. MCD fed mice showed increased serum ALT and AST levels and moderate to severe hepatic steatosis with inflammatory infiltration, hepatic spot or focal necrosis, light portal and sinus hepaticus fibrosis in the liver sections, which associated with enhanced expression of HO-1, TNFalpha and IL-6 mRNA and protein (1.13+/-0.11, 1.74+/-0.05; 0.20+/-0.01, 1.92+/-0.10; 0.58+/-0.02, 2.06+/-0.05 vs 0.43+/-0.02, 0.75+/-0.05; 0.08+/-0.00, 0.59+/-0.02; 0.22+/-0.01, 0.91+/-0.02). Administration of hemin significantly decreased serum ALT and AST levels and attenuated hepatic steatosis and necroinflammation which associated with up-regulation of antioxidative gene HO-1 and down-regulation of pro-inflammatory cytokines TNFalpha and IL-6 (P < 0.01). A contrary effect on serum aminotransferase levels and liver histopathology was observed in mice injected with ZnPP-IX (P < 0.01). CONCLUSIONS: The effect was associated with suppressed HO-1 expression and increased TNFaLPHA and IL-6 expression. The data provided a biochemical, morphological and molecular biological evidence for the protective role of HO-1 in ameliorating hepatic steatosis, necroinflammation in experimental nutritional steatohepatitis.
Asunto(s)
Hígado Graso/metabolismo , Hígado Graso/patología , Hemo-Oxigenasa 1/metabolismo , Proteínas de la Membrana/metabolismo , Animales , Interleucina-6/metabolismo , Hígado/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Enfermedad del Hígado Graso no Alcohólico , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
OBJECTIVE: Hepatic oxidative stress plays a key role in the development of non-alcoholic steatohepatitis (NASH). However, the protective effects of antioxidants on NASH are largely unknown. The aim of this study was to elucidate the effect and mechanism of antioxidants on NASH in mice. MATERIAL AND METHODS: C57BL6/J mice were fed a methionine-choline-deficient (MCD) diet for 10 days or 3 weeks to induce steatohepatitis. Antioxidants (vitamin E, ABT, or vitamin E plus ABT) were supplemented in mice fed a MCD diet, respectively. The effect of antioxidants on oxidative stress and apoptosis was assessed, and activation of adiponectin and expressions of inflammatory factors, apoptosis-related genes, and fibrosis-related genes were assayed. RESULTS: MCD feeding in mice showed increasing serum alanine aminotransferase (ALAT) and aspartate aminotransferase (ASAT) levels, and progressive hepatic injury including hepatic steatosis and inflammatory infiltration. Administration of antioxidants vitamin E and/or ABT significantly lowered serum ALAT and ASAT levels (p<0.001) and ameliorated hepatic steatosis and necroinflammation. These effects were associated with repressed hepatic lipid peroxides through reducing hepatic MDA content and enhancing hepatic superoxide dismutase (SOD) activity; down-regulated inflammatory factor COX-2, lowered activity of NF-kappaB, up-regulated anti-apoptotic gene Bcl-2, and down-regulated pro-apoptotic gene Bax suppressed expression of the fibrotic genes TGF-beta1 and MMP2. Moreover, expression of the anti-inflammatory factor adiponectin was also induced by vitamin E or ABT. A combination of vitamin E and ABT showed an additive effect on preventing liver injury. CONCLUSIONS: The present study provides morphological and molecular biological evidence for the protective role of the antioxidant vitamins E and ABT in ameliorating oxidative stress, hepatic apoptosis, and necroinflammation in experimental nutritional steatohepatitis.