NAMPT-mediated NAD+ biosynthesis suppresses activation of hepatic stellate cells and protects against CCl4-induced liver fibrosis in mice.

Background: Nicotinamide phosphoribosyltransferase (NAMPT) catalyzes the rate-limiting step in the salvage pathway of mammalian nicotinamide adenine dinucleotide (NAD+) biosynthesis. Through its NAD+-biosynthetic activity, NAMPT is able to regulate the development of hepatic steatosis and inflammation induced by diet or alcohol. However, the roles NAMPT plays in the development of liver fibrosis remain obscure. Purpose: To investigate the roles of NAMPT-mediated NAD+ biosynthesis in hepatic stellate cell (HSC) activation and liver fibrosis. Research Design: Realtime RT-PCR and western blot analyses were performed to analyze the expression of profibrogenic genes. Sirius red staining was conducted to examine the fibrosis in liver. Mouse liver fibrosis was induced by intraperitoneal injection of carbon tetrachloride (CCl4) 2 times a week for 6 weeks. Adenovirus-mediated NAMPT overexpression or nicotinamide mononucleotide (NMN) administration was carried out to study the effects of elevation of NAD+ levels on protecting CCl4-induced liver fibrosis in mice. LX2 cells or primary HSCs were used to study the role of NAMPT overexpression or NMN treatment in reducing profibrogenic gene expression in vitro. ResultsCCl4 administration suppresses NAMPT expression in liver and reduces hepatic NAD+ content. Tgfß1 treatment decreases intracellular NAD+ levels and NAMPT expression in LX2 cells. Adenovirus-mediated NAMPT overexpression augments liver NAD+ levels, inhibits HSC activation and alleviates CCl4-induced liver fibrosis in mice. Administration of NMN also suppresses HSC activation and protects against CCl4-induced liver fibrosis in mice. Conclusions: NAMPT-mediated NAD+ biosynthesis inhibits HSC activation and protects against CCl4-induced liver fibrosis.

Hepatoprotective effect of Spirulina platensis against carbon tetrachloride-induced liver injury in male rats.

OBJECTIVES: Spirulina platensis (SP) is an edible Cyanobacterium with ethnomedicinal significance. This study aims at evaluating the beneficial effect of SP against carbon tetrachloride (CCl4)-induced liver toxicity in male rats. METHODS: Rats received intraperitoneal injections of CCl4 (2 ml/kg body weight [b.w.] per every other day) for 40 days, alone or in combination with oral treatments of SP (400 mg/kg b.w. per day). KEY FINDINGS: SP attenuated haematological disturbances, serum liver markers, hepatic necrosis and inflammation, and dyslipidemia in CCl4-intoxicated rats. SP also reduced CCl4-induced oxidative stress by increasing the activities of antioxidant enzymes, such as glutathione peroxidase, superoxide dismutase and catalase and glutathione content, and inhibiting lipid peroxidation products and nitric oxide levels in the rat liver. Further investigations revealed that SP counteracted CCl4-induced increased hepatic levels of Ki-67 (a parameter of cell proliferation), interleukin-6, and tumour necrosis factor-alpha and cyclooxygenase-2 messenger RNA expression. Noticeably, the supplementation of SP restored the decrease of proapoptotic p53 protein levels in the liver of rats treated with CCl4. CONCLUSIONS: SP prevented liver damage in CCl4-treated rats via augmentation of antioxidant defense mechanisms and inhibition of inflammatory cytokines/mediators and antiproliferative effects.

Zingiber roseum Rosc. rhizome: A rich source of hepatoprotective polyphenols.

Zingiber roseum is native to Bangladesh and widely used in folk medicine. This present study was designed to assess the ameliorative potential of Zingiber roseum rhizome extract in carbon tetrachloride (CCl4) induced hepatotoxicity in mice model. Seven phenolic compounds were identified and quantified by HPLC analysis in the plant extract, including quercetin, myricetin, catechin hydrate, trans-ferulic acid, trans-cinnamic acid, (-) epicatechin, and rosmarinic acid. Hepatotoxicity was induced by administrating a single intraperitoneal injection of CCl4 (10 mL/kg) on 7th day of treatment. The results revealed that plant extract at all doses (100, 200 and 400 mg/kg) significantly reduced (p < 0.05) the elevated serum aspartate aminotransferase (AST), alanine aminotransferase (ALT) and alkaline phosphatase (ALP) concentrations, and these effects were comparable to that of standard drug silymarin. Histopathological examination also revealed the evidence of recovery from CCL4 induced cellular damage when pretreated with Z. roseum rhizome extract. The in-vivo hepatoprotective effects were further investigated by the in-silico study of the aforementioned compounds with liver-protective enzymes such as superoxide dismutase (SOD), peroxiredoxin, and catalase. The strong binding affinities (ranging from -7.3359 to -9.111 KCal/mol) between the phenolic compounds (except trans-cinnamic acid) and oxidative stress enzymes inhibit ROS production during metabolism. The compounds were also found non-toxic in computational prediction, and a series of biological activities like antioxidant, anticarcinogen, cardio-protectant, hepato-protectant have been detected.

Blockade of periostin-dependent migration and adhesion by curcumol via inhibition of nuclear factor kappa B signaling in hepatic stellate cells.

OBJECTIVES: Curcumol, a guaiane-type sesquiterpenoid hemiketal extracted from the herb Rhizoma Curcumae, exhibits multiple-pharmacological activities. We previously reported that curcumol ameliorated hepatic fibrosis by inhibiting hepatic stellate cell (HSC) activation. In this study, we aimed to investigate the effect of curcumol on HSC migration and adhesion, and reveal its regulation mechanisms. MATERIALS AND METHODS: Cellular viability was determined by Cell Counting Kit-8. Cell migration was detected by boyden chamber and cell scratch experiment. Recombinant human periostin (rh POSTN) and adeno-associated viral (AAV)-GFP-periostin were used to achieve POSTN overexpression in vitro and in vivo, respectively. Nuclear factor kappa B (NF-κB)-p65 overexpression was achieved by using plasmid. ELISA was conducted to detect POSTN level. Immunohistochemistry, qRT-PCR, Western blotting, and immunofluorescence were performed to assess associated factor expression. RESULTS: Curcumol suppressed HSC migration and adhesion, and reduced the secretion and expression of POSTN. By gain of function POSTN in HSCs, using rh POSTN, we found that the inhibition of HSC migration and adhesion by curcumol depended on the decrease of POSTN. Besides, curcumol protection against chronic CCl4-caused hepatic fibrosis could be impaired by POSTN overexpression. Moreover, we showed that curcumol repressed NF-κB signaling and the production of pro-inflammatory factor. Importantly, curcumol down-regulation of POSTN was rescued by knock-in of NF-κB, as well as the inhibition of HSC migration and adhesion. CONCLUSION: These findings reveal the molecular mechanism of curcumol-reduced HSC migration and adhesion, by which points to the possibility of using curcumol based on NF-κB dependent POSTN for the treatment of fibrogenesis.

The involvement of sirtuin 1 and heme oxygenase 1 in the hepatoprotective effects of quercetin against carbon tetrachloride-induced sub-chronic liver toxicity in rats.

The present study was designed to evaluate the therapeutic potential of quercetin in a sub-chronic model of hepatotoxicity. The roles of putative antioxidant enzymes, sirtuin 1 (SIRT1) and heme oxygenase 1 (HO-1), in hepatoprotection were also addressed. Sub-chronic liver injury was induced in rats by intraperitoneal administration of 0.5 ml/kg carbon tetrachloride (CTC), once every 3 days, for 2 weeks. Some CTC rats were concurrently treated with 100 mg/kg quercetin, intragastrically, once every day, for 2 weeks. The effects of these drugs in the liver were evaluated by biochemical, histological, immunohistochemical and molecular biological studies. CTC triggered oxidative damage to the liver as unanimously shown by altered biochemical parameters and liver morphology. Furthermore, CTC highly upregulated HO-1 and SIRT1 expression levels. Concomitant treatment of rats with quercetin downregulated SIRT1 expression and ameliorated the hepatotoxic effects of CTC. However, quercetin did not have any significant effect on HO-1 expression and bilirubin levels. Collectively, these results suggest that the antioxidant and cytoprotective effects of quercetin in CTC treated rats were SIRT1 mediated and less dependent on HO-1. Thus, pharmacologic modulation of SIRT1 could provide a logic therapeutic approach in sub-chronic hepatotoxicity.

Combination of Sitagliptin and Silymarin ameliorates liver fibrosis induced by carbon tetrachloride in rats.

Liver fibrosis is a common pathological condition that occurs in most conditions associated with chronic liver injury. Silymarin is a herbal product widely used for its hepatoprotective effect. Sitagliptin, a dipeptidyl peptidase-4 inhibitor (DPP4-I), is clinically used as an oral antidiabetic agent. This study was designed to investigate the effects of Sitagliptin, Silymarin, and their combination on established liver fibrosis in carbon tetrachloride (CCl4) rat model. Male albino rats received intraperitoneal injections of CCl4 three times a week for 7 weeks, as well as daily oral treatments of Sitagliptin (100mg/kg) or Silymarin (100mg/kg) or their combination during the 7 weeks of intoxication. Hepatic fibrotic changes were evaluated by measuring hepatic enzymes (ALT, AST, ALP, and GGT) and markers of fibrosis (transforming growth factor ß1 (TGF-ß1), tissue 4-hydroxyproline level, histopathological score), oxidative stress (MDA, GSH, and NOx levels), inflammation (interleukin-6) as well as markers of HSCs activation (α-smooth muscle actin (α-SMA) expression). The injected rats with CCl4 for 7 weeks resulted in a marked elevation of hepatic fibrotic changes and reduction of GSH level, while the combination therapy showed a significant decrease in the former one and a significant increase in the later. In conclusion, this study shows that the combination therapy is more beneficial than monotherapy in ameliorating liver fibrosis in rats. Our findings suggest that Sitagliptin alone or in combination with Silymarin may introduce a new strategy for treating liver fibrosis in humans.

Production of bioactive recombinant rat soluble receptor for advanced glycation end products (rrsRAGE) in Pichia pastoris.

Soluble receptor for advanced glycation end products (sRAGE), a natural inhibitor of RAGE, is considered to be a putative therapeutic molecule for a variety of diseases and a biomarker for certain conditions. To further study the function of sRAGE, recombinant rat sRAGE (rrsRAGE) was expressed and produced in a eukaryotic system. The open reading frame of rat sRAGE was cloned downstream of the methanol-inducible alcohol oxidase promoter of pPICZαA vector, and Pichia pastoris strain X-33 was used as the host strain. The expression of rrsRAGE was achieved by fermentation in a 15-L bioreactor and the resulting fermentation broth was subjected to purification on a cation exchange chromatography column. The purification of rrsRAGE reached 95% after size exclusion chromatography(SEC). The bioactivity of the purified protein was confirmed in a SH-SY5Y cell proliferation assay. The biological function of the purified rrsRAGE protein rat CCl4-induced model was then examined. Treatment with rrsRAGE resulted in significantly lower liver fibrosis and lower serum level of ALT, suggesting that sRAGE prevent liver from injury and fibrosis. In conclusion, we achieved high-efficiency production of bioactive rrsRAGE in P. pastoris.

Obeticholic acid protects against carbon tetrachloride-induced acute liver injury and inflammation.

The farnesoid X receptor (FXR) is a ligand-activated transcription factor that plays important roles in regulating bile acid homeostasis. The aim of the present study was to investigate the effects of obeticholic acid (OCA), a novel synthetic FXR agonist, carbon tetrachloride (CCl4)-induced acute liver injury. Mice were intraperitoneally injected with CCl4 (0.15ml/kg). In CCl4+OCA group, mice were orally with OCA (5mg/kg) 48, 24 and 1h before CCl4. As expected, hepatic FXR was activated by OCA. Interestingly, OCA pretreatment alleviated CCl4-induced elevation of serum ALT and hepatic necrosis. Moreover, OCA pretreatment inhibited CCl4-induced hepatocyte apoptosis. Additional experiment showed that OCA inhibits CCl4-induced hepatic chemokine gene Mcp-1, Mip-2 and Kc. Moreover, OCA inhibits CCl4-induced hepatic pro-inflammatory gene Tnf-α and Il-1ß. By contrast, OCA pretreatment elevated hepatic anti-inflammatory gene Il-4. Further analysis showed that OCA pretreatment inhibited hepatic IκB phosphorylation and blocked nuclear translocation of NF-κB p65 and p50 subunits during CCl4-induced acute liver injury. In addition, OCA pretreatment inhibited hepatic Akt, ERK and p38 phosphorylation in CCl4-induced acute liver injury. These results suggest that OCA protects against CCl4-induced acute liver injury and inflammation. Synthetic FXR agonists may be effective antidotes for hepatic inflammation during acute liver injury.

Antioxidant and hepatoprotective activity of Hamelia patens extracts.

Hamelia patens is widely used in the traditional medicine of Mexico and Central America for the treatment of illnesses associated with inflammatory processes. In this study, antioxidant and hepatoprotective activity were assayed on the methanolic crude (ME), hexane (HE), ethyl acetate (AE), and butanol (BE) extracts of H. patens. The total phenolic content (TPC) as mg of gallic acid equivalents per g of dry extract was determined by Folin-Ciocalteu's method (ME=141.58±11.99, HE=33.96±1.13, AE=375.18±13.09, BE=132.08±3.62), and antioxidant activity by 2,2-diphenyl-1-picryl-hydrazyl (DPPH) free radical-scavenging method (EC(50) ME=77.87±5.67, HE=236.64±26.32, AE=45.87±2.24, BE=50.97±0.85µg/mL). Hepatoprotective activity was evaluated through AST activity on HepG2 cells subjected to damage with CCl(4) (ME=62.5±3.41, HE=72.25±2.87, AE=63.50±4.20, BE=43.74±4.03). BE showed the greater hepatoprotective activity and a good antioxidant capacity, while HE did not show hepatoprotective or antioxidant activity. Cytotoxicity was evaluated on Vero cells cultures; none showed significant toxicity.

Single Dose Oral and Intravenous Pharmacokinetics and Tissue Distribution of a Novel Hesperetin Derivative MTBH in Rats.

BACKGROUND: MTBH, a novel hesperetin derivative, possesses in vivo hepatoprotective effects against carbon tetrachloride (CCl4)-induced acute liver injury in Institute of Cancer Research (ICR) mice. OBJECTIVES: This study investigated the pharmacokinetics and tissue distribution of MTBH and its conjugated metabolites in rats after a single dose of MTBH. METHODS: Male Sprague-Dawley (SD) rats were orally administered (25, 50, 100 mg/kg) or intravenously administered (25 mg/kg) MTBH and blood samples were withdrawn at specific times. Moreover, after a single oral dose of MTBH (200 mg/kg), tissues (heart, liver, spleen, lung, kidney, stomach, intestine, brain and muscle) were collected at scheduled time points. RESULTS: The concentration of MTBH in plasma and tissues was assayed by HPLC before and after hydrolysis with ß-glucuronidase or sulfatase. The glucuronides/sulfates were extensively present in the plasma, moreover, the free form was detectable in the plasma, but in a small amount equivalent to nearly 0.85-1.46 % of the amount of glucuronides/sulfates, the absolute bioavailability of MTBH was approximately 31.27 %. In tissues, the free form appeared in all tissues examined, with trace amount in brain and muscle, and considerable concentration in stomach and lung. Glucuronides/sulfates were the major forms in intestine, kidney and liver, whereas not detectable in heart, brain and muscle. The liver and intestine were found likely to accumulate MTBH at a high concentration among all tissues. CONCLUSIONS: The free form of MTBH was present in the circulation and all assayed organs, whereas its glucuronides/sulfates were the major forms in plasma and intestine, kidney and liver after a single dose.

Resolvin D1 attenuates CCl4-induced acute liver injury involving up-regulation of HO-1 in mice.

Acute hepatic failure involves in excessive oxidative stress and inflammatory responses, leading to a high mortality due to lacking effective therapy. Resolvin D1 (RvD1), an endogenous lipid mediator derived from polyunsaturated fatty acids, has been shown anti-inflammatory and anti-oxidative actions, however, whether RvD1 has protective effects on hepatic failure remains elusive. In this study, the roles and molecular mechanisms of RvD1 were explored in carbon tetrachloride (CCl4)-induced acute liver injury. Our results showed that RvD1 protected mice against CCl4-induced hepatic damage, as evaluated by reduced aminotransferase activities and malondialdehyde content, elevated glutathione and superoxide dismutase activities, and alleviated hepatic pathological damage. Moreover, RvD1 significantly attenuated serum tumor necrosis factor-α and interleukin-6 levels as well as hepatic myeloperoxidase activity, whereas enhanced serum IL-10 level in CCl4-administered mice. Further, RvD1 markedly up-regulated the expression and activity of heme oxygenase-1 (HO-1). However, inhibition of HO-1 activity reversed the protective effects of RvD1 on CCl4-induced liver injury. These results suggest that RvD1 could effectively prevent CCl4-induced liver injury by inhibition of oxidative stress and inflammation, and the underlying mechanism may be related to up-regulation of HO-1.

Melatonin inhibits autophagy and endoplasmic reticulum stress in mice with carbon tetrachloride-induced fibrosis.

This study aimed to investigate whether inhibition of autophagy and endoplasmic reticulum (ER stress) associates with the antifibrogenic effect of melatonin in mice treated with carbon tetrachloride (CCl4 ). Mice received CCl4 5 µL/g body wt i.p. twice a week for 4 wk or 6 wk. Melatonin was given at 5 or 10 mg/kg/day i.p, beginning 2 wk after the start of CCl4 administration. Treatment with CCl4 resulted in fibrosis evidenced by the staining of α-smooth muscle actin (α-SMA)-positive cells. CCl4 induced an autophagic response measured as the presence of autophagic vesicles, protein 1 light chain 3 (LC3) staining, conversion of LC3-I to autophagosome-associated LC3-II, changes in expression of beclin-1, UV radiation resistance-associated gene (UVRAG), ubiquitin-like autophagy-related (Atg5), Atg12, Atg16L1, sequestosome 1 (p62/SQSTM1), and lysosome-associated membrane protein (LAMP)-2, and increased phosphorylation of the mammalian target of rapamycin (mTOR). There was an increase in the expression of the ER stress chaperones CCAAT/enhancer-binding protein homologous protein (CHOP), immunoglobulin-heavy-chain-binding protein (BiP/GRP78), and 94-kDa glucose-regulated protein (GRP94), and in the mRNA levels of pancreatic ER kinase (PERK), activating transcription factor 6 (ATF6), ATF4, inositol-requiring enzyme 1 (IRE1), and spliced X-box-binding protein-1 (XBP1). Phospho-IRE1, ATF6, and phospho-PERK protein concentration also increased significantly. Immunohistochemical staining of α-SMA indicated an abrogation of hepatic stellate cells activation by melatonin. Furthermore, treatment with the indole resulted in significant inhibition of the autophagic flux and the unfolded protein response. Findings from this study give new insight into molecular pathways accounting for the protective effect of melatonin in fibrogenesis.

MicroRNA-30 Protects Against Carbon Tetrachloride-induced Liver Fibrosis by Attenuating Transforming Growth Factor Beta Signaling in Hepatic Stellate Cells.

Transforming growth factor beta (TGF-ß) is crucial for transdifferentiation of hepatic stellate cells (HSCs) and the blunting of TGF-ß signaling in HSCs can effectively prevent liver fibrosis. Krüppel-like factor 11 (KLF11) is an early response transcription factor that potentiates TGF-ß/Smad signaling by suppressing the transcription of inhibitory Smad7. Using a mouse model of carbon tetrachloride (CCl4)-induced liver fibrosis, we observed significant upregulation of KLF11 in the activated HSCs during liver fibrogenesis. Meanwhile, the downregulation of miR-30 was observed in the HSCs isolated from fibrotic liver. Adenovirus-mediated ectopic expression of miR-30 was under the control of smooth muscle α-actin promoter, showing that the increase in miR-30 in HSC greatly reduced CCl4-induced liver fibrosis. Subsequent investigations showed that miR-30 suppressed KLF11 expression in HSC and led to a significant upregulation of Smad7 in vivo. Mechanistic studies further confirmed that KLF11 was the direct target of miR-30, and revealed that miR-30 blunted the profibrogenic TGF-ß signaling in HSC by suppressing KLF11 expression and thus enhanced the negative feedback loop of TGF-ß signaling imposed by Smad7. Finally, we demonstrated that miR-30 facilitated the reversal of activated HSC to a quiescent state as indicated by the inhibition of proliferation and migration, the loss of activation markers, and the gain of quiescent HSC markers. In conclusion, our results define miR-30 as a crucial suppressor of TGF-ß signaling in HSCs activation and provide useful insights into the mechanisms underlying liver fibrosis.

Efecto hepatoprotector del Noni-C® en la intoxicación experimental inducida por tetracloruro de carbono / Noni-C® hepatoprotective effect in experimental intoxication induced by carbon tetrachloride

INTRODUCCIÓN: las enfermedades hepáticas son un serio problema de salud y la carencia de un tratamiento efectivo en la medicina moderna hace que aumenten los esfuerzos por hallar medicamentos naturales apropiados. OBJETIVO: determinar el efecto hepatoprotector del producto natural Noni-C® en la intoxicación experimental por tetracloruro de carbono (CCl4). MÉTODOS: estudio experimental en ratas Wistar macho; se emplearon cuatro grupos de trabajo, uno control negativo, uno control positivo tratado con CCl4 y dos experimentales tratados con Noni-C® a las dosis 200 y 400 mg/kg de peso corporal durante 21 días, más CCl4 postratamiento con Noni-C® por 3 días a la dosis 0,5 mL/kg intraperitoneal. Se determinaron los niveles de transaminasa glutámico pirúvica y glutámico oxalacética. Se realizó análisis histopatológico para determinar lesión hepática y renal de diferente grado. RESULTADOS: se obtuvo reducción significativa de la transaminasa glutámico pirúvica al comparar las dosis de 200 y 400 mg/kg. Se observó disminución de las lesiones histopatológicas hepáticas y renales, de esteatosis hepática severa a leve y moderada, y de necrosis tubular aguda a tumefacción celular moderada, respectivamente, a la dosis 400 mg/kg de Noni-C®. CONCLUSIÓN: el tratamiento preventivo con Noni-C® a la dosis 400 mg/kg reduce la gravedad del daño hepático resultante de la intoxicación por CCl4. Por las características químico-físicas del producto y la variedad de compuestos identificados en el fruto, entre ellos vitaminas y minerales que contribuyen con su capacidad antioxidante, se recomienda su estudio en la prevención de enfermedades hepáticas.

INTRODUCTION: liver diseases are a serious health problem and the lack of effective treatment in modern medicine drives up efforts to find suitable natural medicines. OBJECTIVE: determine the hepatoprotective effect of Noni-c® natural product in experimental poisoning carbon tetrachloride (CCL4). METHODS: an experimental study was conducted in wistar male rats; four working groups were formed: one negative control, a positive control treated with CCL4 and two experimental Noni-c® treated at doses 200 and 400 mg/kg body weight for 21 days, plus CCL4 after treatment with Noni-C® for 3 days in 0.5 mL/kg dose intraperitoneally. Levels of glutamic oxaloacetic and glutamic pyruvic transaminase were determined. Histopathological analysis was performed to determine liver and kidney damage at different levels. RESULTS: significant reduction in glutamic pyruvic transaminase was observed when comparing 200 and 400 mg/kg doses. Decrease liver and kidney histopathological lesions, severe to mild and moderate hepatic steatosis were observed; acute tubular necrosis or moderate cell swelling, respectively, at 400 mg/kg Noni-c® dose was also observed. CONCLUSION: preventive treatment at 400 mg/kg Noni-c® dose reduced the severity of liver damage resulting from CCl4 poisoning. Due to the chemical-physical product features and variety of compounds identified in this fruit, including vitamins and minerals which contribute as antioxidant, its study is recommended in preventing liver diseases.

Myricitrin exhibits antioxidant, anti-inflammatory and antifibrotic activity in carbon tetrachloride-intoxicated mice.

Myricetin-3-O-α-rhamnoside (myricitrin) is a naturally occurring phenolic compound which possesses antioxidant and anti-inflammatory activity. The aim of this study was to determine the hepatoprotective effects of myricitrin. Myricitrin at doses of 10, 30 and 100 mg/kg and silymarin at dose of 100mg/kg were administered to BALB/cN mice by oral gavage, once daily for two consecutive days following carbon tetrachloride (CCl4)-intoxication. Myricitrin significantly ameliorated CCl4-induced increase in serum aspartate transaminase (AST) and alanine transaminase (ALT) levels and histopathological changes in the liver. Hepatic oxidative stress was reduced by myricitrin, as evidenced by the decrease in lipid peroxidation, with concomitant increase in glutathione (GSH) level and cytochrome P450 2E1 (CYP2E1) expression. In addition, cyclooxygenase-2 (COX-2) and tumor necrosis factor-alpha (TNF-α) overexpression in the liver was reduced, suggesting the suppression of inflammation. The expression of transforming growth factor-beta1 (TGF-ß1) and alpha-smooth muscle actin (α-SMA) was markedly ameliorated, indicating the inhibition of profibrotic response. Myricitrin also improved the regeneration of hepatic tissue after CCl4-intoxication, as evidenced by increased proliferating cell nuclear antigen (PCNA) expression. The results of the current study suggest that myricitrin exhibits a significant hepatoprotective activity. Myricitrin provided better hepatoprotection when compared to silymarin, which is consistent with its higher in vitro antioxidant potential.

The protective effects of Acanthus ilicifolius alkaloid A and its derivatives on pro- and anti-inflammatory cytokines in rats with hepatic fibrosis.

This study was designed to investigate the protective effects of Acanthus ilicifolius alkaloid A [4-hydroxy-2-benzoxazolone (HBOA)] and its acetylated derivatives including 4-acetoxy-2-benzoxazolone (TC-2) and 3-acetyl-4-acetoxy-2-benzoxazolone (TC-3) on carbon tetrachloride (CCl4 )-induced liver fibrosis in rats. Sprague-Dawley rats were given CCl4 twice per week for 8 weeks to induce liver fibrosis. Then, they were treated with HBOA, TC-2, and TC-3 daily for 4 weeks, respectively. The serum indicators including total protein (TP), albumin (Alb), globulin, hyaluronic acid (HA), and laminin (LN) were measured by commercial kits. The messenger ribonucleic acid expression of adiponectin, peroxisome proliferator-activated receptor-γ (PPAR-γ), tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), transforming growth factor-ß1 (TGF-ß1 ) and Toll-like receptor 4 (TLR4 ) was determined by reverse-transcriptase -PCR. The proteins of adiponectin, TGF-ß1 , α-smooth muscle actin (α-SMA), and TLR4 were also detected by the immunohistochemical assay. The results showed that HBOA, TC-2, and TC-3 significantly attenuated the fibrotic degree induced by CCl4 as evidenced by higher levels of TP, Alb, adiponectin, and PPAR-γ, which in turn decreased the proliferation of hepatic stellate cells. Moreover, those drugs markedly decreased the levels of HA, LN, TNF-α, IL-6, TGF-ß1 , α-SMA, and TLR4 . Our study indicates that HBOA, TC-2, and TC-3 have beneficial effects against liver fibrosis, and the mechanisms may be related to the inhibition of inflammatory response.

Leishmanial lipid affords protection against oxidative stress induced hepatic injury by regulating inflammatory mediators and confining apoptosis progress.

Persistence of liver injury alters the internal milieu, promotes deregulation of inflammatory factors, and leads to dysplastic lesions like fibrosis, cirrhosis to hepatocellular carcinoma. Our previous study revealed that leishmanial lipid (pLLD) exerts potential anti-inflammatory activity in sepsis associated hepatic injury. We now show that pLLD gives protection against chemical induced hepatotoxicity in murine system. The beneficial effect of treatment with pLLD on such hepatic injury in mice was analyzed using different assays including ELISA, FACS, western blot and immunohistochemical analysis. pLLD significantly suppressed serum enzymes and rectified the histopathological alteration to induce the antioxidant level in CCl4 intoxicated liver. Levels of several growth factors including TGF-ß, HGF, and EGF were significantly improved in serum and hepatic tissue with consequent reduction of caspase activities and expressions of Bad, Bax, p53, and NF-κBp65. Moreover, pLLD modulated inflammatory responses by decreasing the production of several cytokines and chemokines, thus preventing the infiltration of immune cells to the damaged area. It accelerated the repair process in liver damage with modulation of signalling cascade via alteration of apoptotic factors. Our experimental approaches suggest that pLLD effectively prevents liver injury mainly through down regulation of oxidative stress and inflammatory response towards anti-apoptotic changes.

Amelioration of carbon tetrachloride-induced pulmonary toxicity with Oxalis corniculata.

This research work was planned to investigate the antioxidant potential of methanolic crude extract of Oxalis corniculata (OCME) against lung injuries initiated by carbon tetrachloride (CCl4) in rats at histological and biochemical level. A total of 42 female Sprague Dawley rats were randomly distributed in to seven groups and each group comprised of six rats. Experiment was completed in 22 days (10 doses at alternate days). Group I was not treated (control rats), while group II was administered with vehicles (olive oil and dimethyl sulfoxide), groups III, IV, and V were treated with 1 ml kg(-1) body weight (b.w.) of CCl4 (20% in olive oil). Group III received only CCl4, whereas groups IV and V were administered with 100 and 200 mg kg(-1) b.w. of OCME, respectively. Group VI was administered with OCME (200 mg kg(-1) b.w.) alone. Group VII was treated with sylimarin (50 mg kg(-1) b.w.). CCl4 enhanced the lipid peroxidation while reduced the glutathione in lung samples. Activities of antioxidant enzymes, catalase, peroxidase, superoxide dismutase, and glutathione-S-transferase decreased in lung homogenates with CCl4. Treatment of CCl4 induced deleterious changes in the microanatomy of lungs by rupturing the alveolar septa, thickening of alveolar walls, and damaging the cells with subsequent collapse of blood vessels due to the accumulation of degenerated blood cells. OCME, dose dependently, prevented the alterations in these parameters. These results suggest that OCME protected the lungs due to its intrinsic properties by scavenging of free radicals generated by CCl4.

Hepatoprotective activity of ginsenosides from Panax ginseng adventitious roots against carbon tetrachloride treated hepatic injury in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Ginseng (Panax ginseng C. A. Meyer) has a beneficial role in the treatment of various diseases including liver disorders like acute/chronic hepatotoxicity, hepatitis, hepatic fibrosis/cirrhosis and hepatocellular carcinoma. MATERIALS AND METHODS: Tissue culture raised mountain ginseng adventitious root (TCMGARs) extract with ginsenosides in abundance was used as an experimental material. 'Sprague-Dawley' male rats were used as experimental systems and were fed with TCMGARs extracts at doses of 30, 100, 300mg/kg body weight for two weeks to test the effect on carbon tetrachloride (CCl4) induced acute liver damage. Field cultivated Korean ginseng root extract fed rats (100mg/kg) were used as positive control. Plasma enzyme levels, alanine aminotransferase (ALT), aspartate aminotransferase (AST) and alkaline phosphatase (ALP) were assessed. Glutathione (GSH) and malondialdehyde (MDA) concentrations were also evaluated. RESULTS: TCMGARs extracts remarkably prevented the elevation of ALT, AST, ALP and liver peroxides in CCl4-treated rats. Hepatic glutathione levels were significantly increased by the treatment with the extracts in experimental groups. CONCLUSION: The TCMGARs rich in varied ginsenosides can afford protection against CCl4-induced hepatocellular injury.

Mistletoe alkaloid fractions alleviates carbon tetrachloride-induced liver fibrosis through inhibition of hepatic stellate cell activation via TGF-ß/Smad interference.

ETHNOPHARMACOLOGICAL RELEVANCE: Mistletoe (Viscum coloratum (Kom.) Nakai) has long been categorized as a traditional herbal medicine in Asia. In addition to its application in cancer therapy, mistletoe has also been used in the treatment of chronic hepatic disorders in China. In the present study, we investigated the antifibrotic effect and mechanisms of action of mistletoe extracts in a rat model of carbon tetrachloride (CCl4)-induced hepatotoxicity. MATERIALS AND METHODS: An experimental model of hepatic fibrosis was established by intraperitoneal injection of rats with CCl4 for 8 weeks. Rats were subsequently treated with a mistletoe alkaloid fraction preparation via oral administration (120mg/kg daily for 8 weeks) or with distilled water as a control. Histopathological changes were observed by hematoxylin and eosin staining and Masson׳s trichrome staining. The expression of markers relevant to hepatic stellate cell (HSC) activation in the liver was assessed by real-time reverse transcription-polymerase chain reaction, immunohistochemistry and western blotting. The anti-fibrosis activity and mechanisms of action of mistletoe alkaloid fractions were further investigated in the HSC-T6 HSC line, following treatment with mistletoe alkaloid fractions (12mg/ml) for 48h. RESULTS: Hepatic fibrosis decreased markedly in CCl4-treated animals following treatment with mistletoe alkaloid fractions, compared to controls. The mRNA levels of transforming growth factor-ß1 (TGF-ß1), procollagen I and tissue inhibitors of metalloproteinases (TIMPs) were significantly downregulated, by about 40%, 40% and 45%, respectively, in liver tissues from rats treated with mistletoe alkaloid fractions. Furthermore, significant downregulation of TGF-ß1, TGF-ß1 receptor, phosphorylated Smad 2 and alpha smooth muscle actin (α-SMA) proteins, by about 45%, 30% and 40%, respectively, was also observed in liver tissues from mistletoe alkaloid fractions-treated rats. In contrast, Smad 7 levels were significantly increased by about 30% in mistletoe alkaloid fractions-treated rats. Treatment of HSC-T6 cells with mistletoe alkaloid fractions significantly induced Smad 7 expression and inhibited the expression of α-SMA, TGFß1, TGF-ß1 receptor, Smad 2 and TIMP-1, in vitro. CONCLUSION: We demonstrate that mistletoe alkaloid fractions decrease extracellular matrix accumulation by inhibiting HSC activation. Mechanistically, this may occur via inhibition of TGF-ß1/Smad 2 and Smad 7 signal transduction, thereby blocking the synthesis of procollagen I and TIMP-1. These findings suggest that mistletoe alkaloid fractions may be a potential therapeutic agent for the treatment of hepatic fibrosis.