Four Different Structural Dietary Polyphenols, Especially Dihydromyricetin, Possess Superior Protective Effect on Ethanol-Induced ICE-6 and AML-12 Cytotoxicity: The Role of CYP2E1 and Keap1-Nrf2 Pathways.

Polyphenols have received attention as dietary supplements for the relief of alcoholic liver disease (ALD) due to various bioactivities. Ethanol-induced rat small intestinal epithelial cell 6 (IEC-6) and alpha mouse liver 12 (AML-12) cell models were pretreated with four dietary polyphenols with different structures to explore their effects on cytotoxicity and potential protective mechanisms. The results showed that polyphenols had potential functions to inhibit ethanol-induced AML-12 and IEC-6 cell damage and oxidative stress, and restore ethanol-induced IEC-6 permeability and tight junction gene expression. Especially, dihydromyricetin (DMY) had the best protective effect on ethanol-induced cytotoxicity, followed by apigenin (API). Western blot results showed that DMY and API had the best ability to inhibit CYP2E1 and Keap1, and promote nuclear translocation of Nrf2, which might be the potential mechanism by which DMY and API attenuate ethanol-induced cytotoxicity. Moreover, the molecular docking results predicted that DMY and API could bind more tightly to the amino acid residues of CYP2E1 and Keap1, which might be one of the inhibitory modes of dietary polyphenols on CYP2E1 and Keap1. This study provided a rationale for the subsequent protective effect of dietary polyphenols on alcohol-induced liver injury in animal models and provided new clues on bioactive components for ALD-protection based on the gut-liver axis.

Phosphodiesterase 4 mRNA Level Suppression is Important for Extract of Black Carrot to Protect Against Hepatic Injury Induced by Ethanol.

Excessive alcohol use often results in alcoholic liver disease (ALD). An early change in the liver due to excessive drinking is hepatic steatosis, which may ultimately progress to hepatitis, liver fibrosis, cirrhosis, and liver cancer. Among these debilitating processes, hepatic steatosis is reversible with the appropriate treatment. Therefore, it is important to find treatments and foods that reverse hepatic steatosis. Black carrot has antioxidant and anti-inflammatory effects. In this study, we examined the effectiveness of black carrot extract (BCE) on hepatic steatosis in in vivo and in vitro ethanol-induced liver injury models. For the in vivo experiments, serum aminotransferase activities enhanced by ethanol- and carbon tetrachloride were significantly suppressed by the BCE diet. Furthermore, morphological changes in the liver hepatic steatosis and fibrosis were observed in the in vivo ethanol-induced liver injury model, however, BCE feeding resulted in the recovery to an almost normal liver morphology. In the in vitro experiments, ethanol treatment induced reactive oxygen species (ROS) levels in hepatocytes at 9 h. Conversely, ROS production was suppressed to control levels and hepatic steatosis was suppressed when hepatocyte culture with ethanol were treated with BCE. Furthermore, we investigated enzyme activities, enzyme protein levels, and messenger RNA levels of alcohol dehydrogenase (ADH), cytochrome p450 2E1 (CYP2E1), and aldehyde dehydrogenase (ALDH) using enzyme assays, western blot, and quantitative reverse transcription-polymerase chain reaction analyses. We found that the activities of ADH, CYP2E1, and ALDH were regulated through the cAMP-PKA pathway at different levels, namely, translational, posttranslational, and transcriptional levels, respectively. The most interesting finding of this study is that BCE increases cAMP levels by suppressing the Pde4b mRNA and PDE4b protein levels in ethanol-treated hepatocytes, suggesting that BCE may prevent ALD.

Fermented Curcuma longa L. Prevents Alcoholic Fatty Liver Disease in Mice by Regulating CYP2E1, SREBP-1c, and PPAR-α.

We examined the efficacy of fermented Curcuma longa L. (FT) on the development of alcoholic fatty liver in mice and investigated the underlying mechanism. The protective potential of FT against ethanol-induced fatty liver was determined using C57BL/6 male mice allocated into four groups (8 mice/group). Control groups received either distilled water or 5 g/kg body weight (b.w.) per day ethanol for 8 days. Treatment groups were administered either 300 mg/kg b.w. per day of milk thistle or FT before receiving ethanol. FT contained a higher amount of caffeic acid and tetrahydrocurcumin than C. longa. FT pretreatment significantly suppressed the elevated hepatic lipid droplets associated with ethanol ingestion. In comparison with ethanol-treated control, FT pretreated mice showed inhibited cytochrome P4502E1 (CYP2E1), sterol regulatory element-binding protein-1 (SREBP-1c), and acetyl-CoA carboxylase production but elevated AMP-activated protein kinase, peroxisome proliferator-activated receptor-alpha (PPAR-α), and carnitine palmitoyltransferase 1 (CPT-1) levels. Taken together, FT is a promising hepatoprotectant for preventing of alcoholic fatty liver through modulating fatty acid synthesis and oxidation.

Nonlinear molecular dynamics of quercetin in Gynocardia odorata and Diospyros malabarica fruits: Its mechanistic role in hepatoprotection.

Liver performs number of critical physiological functions in human system. Intoxication of liver leads to accumulation of free radicals that eventually cause damage, fibrosis, cirrhosis and cancer. Carbon tetrachloride (CCl4) belongs to hepatotoxin is converted to a highly reactive free radical by cytochrome P450 enzymes that causes liver damage. Plant extracts derived quercetin has substantial role in hepatoprotection. This study highlights the possible mechanism by which quercetin plays significant role in hepatoprotection. HPLC analysis revealed the abundance of quercetin in the fruit extracts of Gynocardia odorata and Diospyros malabarica, were isolated, purified and subjected to liver function analysis on Wistar rats. Post quercetin treatment improved liver function parameters in the hepatotoxic Wistar rats by augmenting bilirubin content, SGOT and SGPT activity. Gene expression profile of quercetin treated rats revealed down regulation of HGF, TIMP1 and MMP2 expressed during CCl4 induction. In silico molecular mechanism prediction suggested that quercetin has a high affinity for cell signaling pathway proteins BCL-2, JAK2 and Cytochrome P450 Cyp2E1, which all play a significant role in CCl4 induced hepatotoxicity. In silico molecular docking and molecular dynamics simulation have shown that quercetin has a plausible affinity for major signaling proteins in liver. MMGBSA studies have revealed high binding of quercetin (ΔG) -41.48±11.02, -43.53±6.55 and -39.89±5.78 kcal/mol, with BCL-2, JAK2 and Cyp2E1, respectively which led to better stability of the quercetin bound protein complexes. Therefore, quercetin can act as potent inhibitor against CCl4 induced hepatic injury by regulating BCL-2, JAK2 and Cyp2E1.

[Extracts of Poria cocos polysaccharides improves alcoholic liver disease in mice via CYP2E1 and NF-κB inflammatory pathways].

The present study investigated the effect of extract of Poria cocos polysaccharides(PCP) on cytochrome P450 2 E1(CYP2 E1) and nuclear factor κB(NF-κB) inflammatory signaling pathways in alcoholic liver disease(ALD) mice and explored its protective effect and mechanism. Sixty male C57 BL/6 N mice of SPF grade were randomly divided into a control group, a model group, a positive drug group(bifendate, 200 mg·kg~(-1)), and high-(200 mg·kg~(-1)) and low-dose(50 mg·kg~(-1)) PCP groups. Gao-binge mo-del was induced and the mice in each group were treated correspondingly. Liver morphological and pathological changes were observed and organ index was calculated. Serum levels of alanine aminotransferase(ALT) and aspartate aminotransferase(AST) were detected. Malondialdehyde(MDA) and superoxide dismutase(SOD) in liver tissues were detected by assay kits. The levels of interleukin-6(IL-6) and tumor necrosis factor-α(TNF-α) were detected by ELISA. The activation of macrophages was observed by immunofluorescence staining and protein expression of CYP2 E1, Toll-like receptor 4(TLR4), NF-κB p65, and phosphorylated NF-κB p65(p-NF-κB p65) were analyzed by Western blot. The ALD model was properly induced. Compared with the model group, the PCP groups significantly improved the pathological injury of liver tissues. Immunofluorescence staining revealed that compared with the model group, the groups with drug intervention showed decreased macrophages in liver tissues. Additionally, the PCP groups showed reduced ALT, AST, MDA, IL-6, and TNF-α(P<0.05), and potentiated activity of SOD(P<0.01). PCP extract has the protective effect against alcoholic liver injury in mice, and the underlying mechanism may be related to the regulation of the expression of CYP2 E1 and inhibition of TLR4/NF-κB inflammatory signaling pathway to reduce oxidative stress and inflammatory injury, thereby inhibiting the development of ALD.

Glabridin attenuates paracetamol-induced liver injury in mice via CYP2E1-mediated inhibition of oxidative stress.

CYP2E1 plays a crucial role in the bio-activation of toxic substances leading to liver damage. In this context, CYP2E1 converts paracetamol (PCM) to N-acetyl-p-benzoquinone imine (NAPQI), which is prone to cause hepatotoxicity. Hence, we aimed to explore the protective effect of glabridin on widely used PCM-induced liver injury model in the present study and, after that, correlated with the role of CYP2E1 toward its efficacy. Glabridin was isolated from Glycyrrhiza glabra and characterized before the investigation in an in-vivo mice model of PCM-induced liver injury. Glabridin after oral treatment at 5-20 mg/kg showed a considerable improvement in serum biochemical parameters (ALT and AST) and oxidative stress markers (MDA, GSH, SOD, and catalase) in comparison to only PCM-treatment. Histopathological examination of the liver depicted that glabridin exhibited substantial protection from PCM-induced liver injury compared to the disease control group. Significant down-regulation of CYP2E1 protein and its mRNA expression levels were observed in the glabridin-treated groups compared to PCM-induced respective elevation of CYP2E1. Moreover, activation of NF-κB was significantly inhibited by glabridin. Therefore, glabridin has the potential to protect PCM-induced liver injury through CYP2E1 inhibition-mediated normalization of oxidative stress. Further research is warranted to establish glabridin as a phytotherapeutics for liver protection for which no effective and safe oral drug is available to date.

Protective effect of Phaeoporus obliquus polysaccharide against acute liver injury induced by carbon tetrachloride and alcohol in mice.

Studied the optimum extraction process of polysaccharide from Phaeoporus obliquus and the effect of Phaeoporus obliquus polysaccharide on carbon tetrachloride (CCl4)- or alcohol-induced acute liver injury in mice. The main factor in influencing the extraction rate of Phaeoporus obliquus polysaccharide were extraction power and time, which was a kind of pyran glucose by infrared spectroscopy. CCl4 and alcohol were employed respectively to establish CCl4 and alcohol-induced acute liver injury mouse models. Compared with model groups mice, Phaeoporus obliquus polysaccharide treatment at the doses of 100mg/kg and 200mg/kg exhibited an obvious reduction liver index, ALP, ALT, AST levels, MDA content and TNF-α level (p<0.01) and SOD activity was increased, which was in a dose-dependent manner. Compared with the model group, the necrosis degree of hepatocytes was obviously reduced and the small fat droplets were formed in some cytoplasm, especially in high dose group, which the liver cells recovered to the level of normal group. Rt-PCR results showed that the expression of CYP2E1 mRNA in liver tissues of Phaeoporus obliquus polysaccharide groups were significantly reduced, and the difference were statistically significant compared with the model group (p<0.05). These results demonstrated that Phaeoporus obliquus polysaccharide has significantly hepatoprotective effect on CCl4 and alcohol-induced acute liver injury in mice.

Alteration of CYP2E1, DBN1, DNMT1, miRNA-335, miRNA-21, c-Fos and Cox-2 gene expression in prefrontal cortex of rats' offspring submitted to prenatal ethanol exposure during their neurodevelopment and the preventive role of nancocurcumin administration: A histological, ultrastructural and molecular study.

Ethanol (EtOH) has been linked to neurotoxic effects on the fetus and prenatal alcohol exposure (PAE) has a negative impact on brain neurodevelopment. Therefore, the present study was aimed to focus on the underlying mechanisms of alcohol-induced oxidative stress and apoptotic cell death in addition to shedding the light on the modulatory effect of nanocurcumin in rats' offspring prefrontal cortices. The current study investigated the effects of prenatal maternal exposure to EtOH intragastric (i.g.) administration of 0.015 mL/g of a 10 % v/v ethanol solution throughout gestation and the concomitant use of nanocurcumin, on 21-day-old offspring Wistar rat prefrontal cortex parameters. CYP2E1, DBN1, DNMT1, miRNA-335, miRNA-21, c-Fos and Cox-2 gene expression as well as the accompanying histological and ultrastructural alterations were assessed. The implemented experimental setting has revealed that ethanol exposure caused significant alterations in the above mentioned parameters. Changes observed in nanocurcumin-treated animals were significantly different to the ethanol-treated group when nanocurcumin was concomitantly administered.

Ameliorative potential of Adhatoda vasica against anti-tubercular drugs induced hepatic impairments in female Wistar rats in relation to oxidative stress and xeno-metabolism.

ETHNOPHARMACOLOGICAL RELEVANCE: Adhatoda vasica Nees is widely used herb of indigenous system to treat various ailments especially upper respiratory tract infections. Not only, anti-tubercular efficacy of crude extract and phytoconstituents of A. vasica has been documented but its hepatoprotective role against various drugs mediated hepatic alterations in different animal models has also been observed. BACKGROUND AND PURPOSE: Isoniazid, rifampicin and pyrazinamide (H-R-Z) are anti-tubercular drugs normally prescribed by health professionals for the treatment of tuberculosis, however along with their medical effectiveness these drugs also exhibit hepatotoxicity among TB patients. Unexpectedly, substantial toxicological data on the metabolism of anti-TB drugs are available but the mystery behind these xenobiotics is too complex and partly implicit. In this study, we further explored the hepatotoxic effects of these xeno-metabolic products and their amelioration by Adhatoda vasica Nees by elucidating its mechanistic action. METHODS: We generated a hepatotoxic rodent model by oral administration of H, R and Z (30.85, 61.7 and 132.65 mg/kg body weight) drugs for 25 days in Wistar rats. Additionally, to achieve hepatoprotection two different doses of Adhatoda vasica Nees ethanolic leaf extract (200 and 300 mg/kg body weight) were used along with H-R-Z dosage, orally and once daily for 25 days and tried to ascertain their mechanistic action. For this, initially phytoconstituents of the extract were evaluated followed by extract standardization using RP-HPLC and FTIR methods. Furthermore, antioxidant activity of the extract was analyzed by DPPH assay. Finally, different treated groups were analyzed for hepatic oxidative stress markers, antioxidant markers, histopathological changes and gene expression study including CYP2E1, CYP7A1, NAT, NR1I2 and UGT1A1 genes involved in phase I and phase II xeno-metabolism. RESULTS: Estimated content of vasicine in RP-HPLC method and free-radical scavenging activity in DPPH assay was found to be 134.519 ± 0.00269µg/10mg of leaf extract and 47.81 µg/mL respectively. In H-R-Z treated group, a significant increase in the levels of thiobarbituric acid, significant reduction in the levels of GSH, and enzymatic markers and marked changes in hepatic histological architecture were observed. In addition, there was significance up-regulation of CYP7A and NAT genes, down-regulation of CYP2E1 gene and insignificant expression levels of NR1I2 and UGT1A1 genes were observed in H-R-Z group. Conversely, high dose of A. vasica extract effectively diminished these alterations by declining oxidative stress and boosting of antioxidant levels. In addition, it acted as bi-functional inducer of both phase I (CYP2E1) and phase II (NAT and UGT1A1) enzyme systems. CONCLUSION: Hence, we concluded that anti-TB drugs exposure has potential to generate reactive metabolites that eventually cause hepatotoxicity by altering oxidant-antioxidant levels and their own metabolism. This study not only emphasized on xeno-metabolism mediated hepatic alterations but also explore the benefit of A. vasica on these toxic insults.

Purified anthocyanins from Zea mays L. cob ameliorates chronic liver injury in mice via modulating of oxidative stress and apoptosis.

BACKGROUND: Purple corn (Zea mays L.) is one of the main economic crops in China and has been used in the treatment of cystitis, urinary infections and obesity. However, purple corncobs, the by-product remaining after processing and having an intense purple-black color, are normally disposed of as waste or used as animal feed. Therefore, to further expand the medicinal value of purple corncob, its content was analyzed and, after purification, the effect and mechanism of purified purple corncob anthocyanins (PPCCA) on CCl4 -induced chronic liver injury in mice were investigated. RESULTS: It was observed that the total anthocyanin content (TAC) from PPCCA (317.51 ± 9.30 mg cyanidin 3-O-glucoside (C-3-G) g-1 dry weight) was significantly higher than that from the purified purple corn seed anthocyanin (266.73 ± 3.67 mg C-3-G g-1 dry weight), of which C-3-G accounted for 90.6% and 90.4% of the TAC, respectively. Furthermore, compared with the CCl4 group, PPCCA treatment significantly reduced liver index, serum total bilirubin, alanine transaminase, aspartate transaminase and liver malondialdehyde levels, but increased liver superoxide dismutase activity. The pathological changes were also improved, such as more regular arrangement of hepatocytes, less swelling, and fewer vacuoles and apoptotic cells. Additionally, mechanistic studies showed that PPCCA downregulated the expression of Caspase-3, Bax and cytochrome P450 2E1 proteins in the liver and upregulated the expression of Bcl-2. CONCLUSION: These results demonstrated that PPCCA could ameliorate CCl4 -induced chronic liver injury by regulating oxidative stress and hepatocyte apoptosis pathways. © 2021 Society of Chemical Industry.

The clinical value and potential molecular mechanism of the downregulation of MAOA in hepatocellular carcinoma tissues.

BACKGROUND: Hepatocellular carcinoma (HCC) remains one of the most common cancers worldwide and tends to be detected at an advanced stage. More effective biomarkers for HCC screening and prognosis assessment are needed and the mechanisms of HCC require further exploration. The role of MAOA in HCC has not been intensively investigated. METHODS: In-house tissue microarrays, genechips, and RNAsequencing datasets were integrated to explore the expression status and the clinical value of MAOA in HCC. Immunohistochemical staining was utilized to determine MAOA protein expression. Intersection genes of MAOA related co-expressed genes and differentially expressed genes were obtained to perform functional enrichment analyses. In vivo experiment was conducted to study the impact of traditional Chinese medicine nitidine chloride (NC) on MAOA in HCC. RESULTS: MAOA was downregulated and possessed an excellent discriminatory capability in HCC patients. Decreased MAOA correlated with poor prognosis in HCC patients. Downregulated MAOA protein was relevant to an advanced TNM stage in HCC patients. Co-expressed genes that positively related to MAOA were clustered in chemical carcinogenesis, where CYP2E1 was identified as the hub gene. In vivo experiment showed that nitidine chloride significantly upregulated MAOA in a nude mouse HCC model. CONCLUSIONS: A decreased MAOA level is not only correlated with aggressive behaviors in males but also serves as a promising biomarker for the diagnosis and prognosis of HCC patients. Moreover, MAOA may play a role in AFB1 toxic transformation through its synergistic action with co-expressed genes, especially CYP3A4. MAOA also serves as a potential therapy target of NC in HCC patients.

Salidroside protects mice against CCl4-induced acute liver injury via down-regulating CYP2E1 expression and inhibiting NLRP3 inflammasome activation.

Salidroside (Sal), a natural phenolic compound isolated from Rhodiola sachalinensis, has been utilized as anti-inflammatory and antioxidant for centuries, however, its effects against liver injury and the underlying mechanisms are unclear. This study was designed to evaluate the protective effects and underlying mechanisms of Sal on carbon tetrachloride (CCl4)-induced acute liver injury (ALI) in mice. C57BL/6 mice were pretreated with Sal before CCl4 injection, the serum and liver tissue were collected to evaluate liver damage and molecular indices. The results showed that Sal pretreatment dose-dependently attenuated CCl4-induced acute liver injury, as indicated by lowering the activities of serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT), and inhibiting hepatic pathological damage and apoptosis. In addition, Sal alleviated CCl4-primed oxidative stress and inflammatory response by restoring hepatic glutathione (GSH), superoxide dismutase (SOD), catalase (CAT), malondialdehyde (MDA), and inhibiting cytokines. Finally, Sal also down-regulated the expression of cytochrome P4502E1 (CYP2E1), and Nod-like receptor protein 3 (NLRP3) inflammasome activation in the liver of mice by CCl4. Our study demonstrates that Sal exerts its hepatoprotective effects on ALI through its antioxidant and anti-inflammatory effects, which might be mediated by down-regulating CYP2E1 expression and inhibiting NLRP3 inflammasome activation.

Flavonoids isolated from loquat (Eriobotrya japonica) leaves inhibit oxidative stress and inflammation induced by cigarette smoke in COPD mice: the role of TRPV1 signaling pathways.

Chronic obstructive pulmonary disease (COPD) is a chronic, progressive lung disease with few successful treatments, and is strongly associated with cigarette smoking (CS). Since the novel coronavirus has spread worldwide seriously, there is growing concern that patients who have chronic respiratory conditions like COPD can easily be infected and are more prone to having severe illness and even mortality because of lung dysfunction. Loquat leaves have long been used as an important material for both pharmaceutical and functional applications in the treatment of lung disease in Asia, especially in China and Japan. Total flavonoids (TF), the main active components derived from loquat leaves, showed remarkable anti-inflammatory and antioxidant activities. However, their protective activity against CS-induced COPD airway inflammation and oxidative stress and its underlying mechanism still remain not well-understood. The present study uses a CS-induced mouse model to estimate the morphological changes in lung tissue. The results demonstrated that TF suppressed the histological changes in the lungs of CS-challenged mice, as evidenced by reduced generation of pro-inflammatory cytokines including interleukin 6 (IL-6), IL-1ß, tumor necrosis factor α (TNF-α), nitric oxide (NO), and inducible nitric oxide synthase (iNOS) and diminished the protein expression of transient receptor potential vanilloid 1 (TRPV1). Moreover, TF also inhibited phosphorylation of IKK, IκB and NFκB and increased p-Akt. Interestingly, TF could inhibit CS-induced oxidative stress in the lungs of COPD mice. TF treatment significantly inhibited the level of malondialdehyde (MDA) and increased the activity of superoxide dismutase (SOD). In addition, TF markedly downregulated TRPV1 and cytochrome P450 2E1 (CYP2E1) and upregulated the expression of SOD-2, while the p-JNK level was observed to be inhibited in COPD mice. Taken together, our findings showed that the protective effect and putative mechanism of the action of TF resulted in the inhibition of inflammation and oxidative stress through the regulation of TRPV1 and the related signal pathway in lung tissues. It suggested that TF derived from loquat leaves could be considered to be an alternative or a new functional material and used for the treatment of CS-induced COPD.

All trans retinoic acid modulates TNF-α and CYP2E1 pathways and enhances regression of ethanol-induced fibrosis markers in hepatocytes and HSCs in abstaining rodent model.

Context: Our previous studies showed that all trans retinoic acid (ATRA) ameliorates alcohol-induced toxicity. Hence, we evaluated the efficacy of ATRA and abstention in the regression of alcohol-induced hepatotoxicity. Materials and methods: After ethanol administration to rats for 90 days, the regression of alcohol-induced toxicity was studied by supplementing ATRA at a dose of 100 µg/kg body weight for 30 days. It was also compared with animals in abstention. Results and discussion: Ethanol administration enhanced oxidative stress, activated HSCs and increased collagen deposition. All these alterations were reversed to a certain extent by ATRA supplementation. Conclusions: ATRA had better efficacy than just abstention in reducing ethanol-induced toxicity. The mechanism might be downregulation of CYP2E1, leading to reduced oxidative stress in the hepatocytes and thus impeding NFκB activation, cytokine production, activation of HSC and resulting in the reduction of inflammation and remodelling of fibrosis by modulating MMP and TIMP.

Sagittaria sagittifolia polysaccharide protects against isoniazid- and rifampicin-induced hepatic injury via activation of nuclear factor E2-related factor 2 signaling in mice.

ETHNOPHARMACOLOGICAL RELEVANCE: The Sagittaria sagittifolia L. polysaccharide (SSP) is a purified form of a homogeneous polysaccharide isolated from the root tubers of S. sagittifolia, which has been used as a protectant against hepatotoxicity induced by coadministration of isoniazid and rifampicin. However, the protective effect of SSP against isoniazid- and rifampicin-induced liver injury has never been studied. AIM OF THE STUDY: In this study, the hepatoprotective effect of SSP and its underlying mechanism were investigated in mice with isoniazid- and rifampicin-induced liver injury. MATERIALS AND METHODS: Liver injury was induced in mice by intragastric administration of isoniazid and rifampicin, and the mice were divided into the following six groups: standard control (administration of saline by gavage), model (intragastric administration of isoniazid and rifampicin at 100 mg/kg/day each), positive control (100 mg/kg/day silymarin by gavage 4 h after isoniazid and rifampicin administration), and SSP-treated (200, 400, or 800 mg/kg/day SSP by gavage after isoniazid and rifampicin administration). Subsequently, blood and liver samples were collected from all the animals and were assessed. RESULTS: SSP significantly alleviated the liver injury, as evidenced by decreased activities of alanine aminotransferase, aspartate aminotransferase, and lactate dehydrogenase in the serum and a decreased level of malondialdehyde in the liver, as well as by an increased level of glutathione and increased activities of superoxide dismutase and catalase in the liver. SSP also effectively reduced the pathological tissue damage. The gene and protein expression of cytochrome P450 (CYP) 2E1 and CYP3A4 was inhibited by SSP. The gene and protein expression of nuclear factor erythroid 2-related factor 2 (NRF2), glutamate-cysteine ligase, and heme oxygenase-1 were induced by SSP, whereas that of Kelch-like ECH-associated protein 1 was inhibited. CONCLUSIONS: SSP exerts a protective effect against isoniazid- and rifampicin-induced liver injury in mice. The underlying mechanisms may involve activation of NRF2 and its target antioxidant enzymes and inhibition of the expression of CYPs.

Black Radish (Raphanus sativus L. var. niger) Extract Mediates Its Hepatoprotective Effect on Carbon Tetrachloride-Induced Hepatic Injury by Attenuating Oxidative Stress.

Nonalcoholic fatty liver disease is a serious liver disorder associated with oxidative stress. Black radish (Raphanus sativus L. var. niger) extract (BRE) can lower the risk of this disease. The hepatoprotective effect of BRE containing 3-(E)-(methylthio)methylene-2-pyrrolidinethione was evaluated in human hepatocyte carcinoma (HepG2) cells and in rat livers with carbon tetrachloride (CCl4)-induced hepatic injury. BRE was administered at 125, 250, 500, and 1000 µg/mL to the oleic acid-induced HepG2 cells. Male Sprague-Dawley rats were randomly divided into seven groups: the control group, BRE group, CCl4 group, and BRE + CCl4 group. BRE was administered orally at 125, 250, 500, and 1000 mg/kg/day once daily for 7 consecutive days, followed by a single oral treatment of 1.5 mL/kg CCl4. Inhibition of lipid accumulation, serum markers of liver injury, histological evaluations, levels of oxidative stress related enzymatic and nonenzymatic antioxidants in HepG2 cells and liver tissue were investigated. The protein expression of main liver P450 isoenzymes such as cytochrome p450(CYP)2E1, the expression of nuclear factor erythroid 2-related factor-2(Nrf-2) and heme oxygenase-1(HO-1) were also studied. BRE has an inhibitory effect on lipid accumulation and caused acute hepatotoxicity manifested by increased levels of lipid peroxidation, serum alanine aminotransferase, and aspartate aminotransferase with corresponding histopathological changes and high levels of oxidative stress. BRE treatment significantly increased the level of CYP2E1, Nrf-2, and HO-1 in a dose-dependent manner. Besides, 3-(E)-(methylthio)methylene-2-pyrrolidinethione significantly increased radical-scavenging effects and the expression of Nrf-2 in oleic acid-treated HepG2 cells. These results suggest that BRE treatment reduces lipid accumulation in oleic acid-induced steatosis of HepG2 cells, and has a hepatoprotective effect against CCl4-induced liver injury in rats, possibly through Nrf-2/HO-1-mediated antioxidant effects.

Kaempferol's Protective Effect on Ethanol-Induced Mouse Primary Hepatocytes Injury Involved in the Synchronous Inhibition of SP1, Hsp70 and CYP2E1.

The mechanism of ethanol-induced hepatotoxicity was complicated, accompanied by the over-expressions of the cytochrome P450 2E1 (CYP2E1), heat shock protein 70 (Hsp70) and the nuclear factor specificity protein 1 (SP1). Kaempferol (Kaem) could protect the ethanol-induced hepatotoxicity likely by inhibiting the CYP2E1 expression and activity. This study investigated the protective mechanism(s) of kaempferol on ethanol-induced toxicity by dynamic alteration of SP1, Hsp70 and CYP2E1 among the nucleus and different organelles in hepatocytes. After ethanol treatment alone and co-incubation hepatocytes with kaempferol, protein levels of CYP2E1, Hsp70, and SP1 were determined in vitro (western blotting and immunofluorescence). Hepatocytes' viability was assessed by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) methods. Glutathione (GSH) levels were evaluated for ethanol-induced oxidative stress. In the ethanol-treated hepatocytes, kaempferol decreased protein levels of CYP2E1 in both microsome and mitochondria, cytosolic Hsp70 and SP1 in nuclear and cytosol, and the oxidative stress and increased the cell viability compared to those of ethanol group. Collectively, our findings propose that the protective mechanism of kaempferol is involved in the synchronous, early and persistent inhibitions of mitochondrial and microsomal CYP2E1, cytosolic Hsp70 and nuclear and cytosolic SP1 in mouse primary hepatocytes' injury induced by ethanol.

Supplementation of all trans retinoic acid ameliorates ethanol-induced endoplasmic reticulum stress.

CONTEXT: Molecular pathogenesis of chronic alcoholism is linked to increased endoplasmic reticulum stress. Ethanol is a competitive inhibitor of vitamin A metabolism and vitamin A supplementation aggravates existing liver problems. Hence, we probed into the impact of supplementation of all trans retinoic acid (ATRA), the active metabolite of vitamin A on ethanol-induced endoplasmic reticulcum stress. METHODS: Male Sprague-Dawley rats were divided into four groups - I: Control; II: Ethanol; III: ATRA; IV: ATRA + Ethanol. After 90 days the animals were sacrificed to study markers of lipid peroxidation in hepatic microsomal fraction and expression of ER stress proteins and apoptosis in liver. RESULTS AND CONCLUSION: Ethanol caused hepatic hyperlipidemia, enhanced microsomal lipid peroxidation, upregulated expression of unfolded protein response associated proteins and that of apoptosis. Ethanol also led to downregulation of retinoid receptors. ATRA supplementation reversed all these alterations indicating the decrease in ethanol-induced endoplasmic reticulum stress.

A novel sesquiterpene glycoside from Loquat leaf alleviates oleic acid-induced steatosis and oxidative stress in HepG2 cells.

Loquat (Eriobotrya japonica) leaf has displayed beneficial effect on metabolic syndrome. In our previously study, total sesquiterpene glycosides (TSG) isolated from Loquat leaf exhibited therapeutic effect on Non-alcoholic fatty liver disease (NAFLD) in vivo, but the accurate active compound remains unknown. Sesquiterpene glycoside 1 (SG1) is a novel compound, which is exclusively isolated from Loquat leaf, but its biological activity has been rarely reported. The present study was designed to evaluate the pharmacological effect of SG1, the main component of TSG, in oleic acid (OA)-induced HepG2 cell model of NAFLD with its related mechanisms of action. In this study, both SG1 and TSG were found to significantly reduce the lipid deposition in the cell model. They could also decrease total cholesterol (TC), triglyceride (TG) and intracellular free fatty acid (FFA) contents. Compared with OA-treated cells, the superoxide dismutase (SOD) level increased, and the malondialdehyde (MDA) and 4-hydroxynonenal levels respectively decreased after the administration of SG1 or TSG. The high dose of SG1 (140 µg/mL) displayed a similar therapeutic effect as TSG at 200 µg/mL. Both SG1 and TSG were found to suppress the expression of cytochrome P450 2E1 (CYP2E1) and the phosphorylation of c-jun terminal kinase (JNK) and its downstream target c-Jun in OA-treated cell. These results demonstrate again that TSG are probably the main responsible chemical profiles of Loquat leaf for the treatment of NAFLD, for which it can effectively improve OA-induced steatosis and reduce oxidative stress, probably by downregulating of CYP2E1 expression and JNK/c-Jun phosphorylation, while SG1 may be the principle compound.

Identification of novel therapeutic target genes and pathway in pancreatic cancer by integrative analysis.

BACKGROUND: Gene alterations are crucial to the molecular pathogenesis of pancreatic cancer. The present study was designed to identify the potential candidate genes in the pancreatic carcinogenesis. METHODS: Gene Expression Omnibus database (GEO) datasets of pancreatic cancer tissue were retrieval and the differentially expressed genes (DEGs) from individual microarray data were merged. Gene Ontology (GO) enrichment analysis, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, protein-protein interaction (PPI) networks, and gene coexpression analysis were performed. RESULTS: Three GEO datasets, including 74 pancreatic cancer samples and 55 controls samples were selected. A total of 2325 DEGs were identified, including 1383 upregulated and 942 downregulated genes. The GO terms for molecular functions, biological processes, and cellular component were protein binding, small molecule metabolic process, and integral to membrane, respectively. The most significant pathway in KEGG analysis was metabolic pathways. PPI network analysis indicated that the significant hub genes including cytochrome P450, family 2, subfamily E, polypeptide 1 (CYP2E1), mitogen-activated protein kinase 3 (MAPK3), and phospholipase C, gamma 1 (PLCG1). Gene coexpression network analysis identified 4 major modules, and the potassium channel tetramerization domain containing 10 (KCTD10), kin of IRRE like (KIRREL), dipeptidyl-peptidase 10 (DPP10), and unc-80 homolog (UNC80) were the hub gene of each modules, respectively. CONCLUSION: Our integrative analysis provides a comprehensive view of gene expression patterns associated with the pancreatic carcinogenesis.