E4bp4-Cyp3a11 axis in high-fat diet-induced obese mice with weight fluctuation.

OBJECTIVE: Weight regain after weight loss is a challenge in obesity management. The metabolic changes and underlying mechanisms in obese people with weight fluctuation remain to be elucidated. In the present study, we aimed to profile the features and clinical significance of liver transcriptome in obese mice with weight regain after weight loss. METHODS: The male C57BL/6J mice were fed with standard chow diet or high-fat diet (HFD). After 9 weeks, the HFD-induced obese mice were randomly divided into weight gain (WG), weight loss (WL) and weight regain (WR) group. After 10 weeks of dietary intervention, body weight, fasting blood glucose (FBG), intraperitoneal glucose tolerance, triglycerides (TG), total cholesterol (T-CHO) and low-density lipoprotein cholesterol (LDL-C) were measured. Morphological structure and lipid droplet accumulation in the liver were observed by H&E staining and oil red O staining, respectively. The liver transcriptome was detected by RNA sequencing. Protein expressions of liver cytochrome P450 3a11 (Cyp3a11) and E4 promoter-binding protein 4 (E4bp4) were determined by Western blot. RESULTS: After 10 weeks of dietary intervention, the body weight, FBG, glucose area under the curve, T-CHO and LDL-C in WL group were significantly lower than those in WG group (P < 0.05). At 4 weeks of HFD re-feeding, the mice in WR group presented body weight and T-CHO significantly lower than those in WG group, whereas higher than those in WL group (P < 0.05). Hepatic vacuolar degeneration and lipid droplet accumulation in the liver were significantly alleviated in WL group and WR group, compared to those in WG group. The liver transcriptome associated with lipid metabolism was significantly altered during weight fluctuation in obese mice. Compared with those in WG group, Cyp3a11 in the liver was significantly upregulated, and E4bp4 was significantly downregulated in WL and WR groups. CONCLUSION: Obese mice experience weight regain after weight loss by HFD re-feeding, but their glucose and lipid metabolism disorders are milder than those induced by the persistence of obesity. Downregulated E4bp4 and upregulated Cyp3a11 are detected in obese mice after weight loss, suggesting that the E4bp4-Cyp3a11 axis may involved in metabolic mechanisms underlying weight regulation.

Effects of Calamintha incana (Sm.) Helder Ethanolic Extract on the mRNA Expression of Drug-metabolizing cyp450s in the Mouse Livers.

BACKGROUND: Alteration in the expression and activity of drug-metabolizing enzymes (DMEs) can alter the pharmacokinetics and hence the response of the drug. Some chemicals found in herbs and fruits affect the expression of DMEs. Calamintha incana is commonly used in Middle Eastern Arabic countries. There is no report regarding the influence of Calamintha incana on the hepatic expression of DMEs. AIMS: The current investigation aimed to investigate the effect of Calamintha incana consumption on the mRNA expression of major hepatic drug-metabolizing cytochrome (cyp) P450 genes in mice. METHODS: The chemical composition of the ethanoic extract was analyzed using liquid chromatography/ mass spectrometry. Then, 21 BALB/c mice were used for the in vivo experiment. The mice were divided into three groups, each consisting of seven mice. The first group (low-dose group) was treated with 41.6 mg/kg of Calamintha incana extract and the second group was administered the high-dose (125 mg/kg) of the extract for one month. The mice in the third "control" group administrated the vehicle 20% polyethylene glycol 200. Then, the expression of cyp3a11, cyp2c29, cyp2d9, and cyp1a1 was analyzed using the real-time polymerase chain reaction. The relative liver weights of the mice and the hepatic pathohistological alterations were assessed. RESULTS: The ethanolic extract of Calamintha incana contained 27 phytochemical compounds. The most abundant compounds were linolenic acid, myristic acid, and p-cymene. It was found that the low dose of Calamintha incana extract upregulated significantly (P < 0.05) the expression of cyp3a11 by more than ten folds in the liver of treated mice. Furthermore, the histological analysis showed that low- and high-dose administration of the C. incana did not cause pathological alterations. CONCLUSION: It can be concluded from these findings that consumption of low doses of Calamintha incana upregulated the mRNA expression of mouse cyp3a11 without causing histopathological alterations in the livers. Further studies are needed to determine the influence of Calamintha incana on the pharmacokinetics and response of drugs metabolized by cyp3a11.

15,16-dihydrotanshinone I in Danshen ethanol extract aggravated cholestasis by inhibiting Cyp3a11 mediated bile acids hydroxylation.

Our previous study found that high-dose Tanshinones Capsule (TC) aggravated cholestasis in mice. To explore its underlying mechanism, main tanshinones components (15,16-dihydrotanshinone I (DTI), cryptotanshinone (CTS) and tanshinone IIA (TSA)) form TC were studied separately. Bile acids (BAs) that were primarily metabolized by hydroxylation were identified, and then the inhibitory effect of each tanshinones on their hydroxylation were evaluated. The anti-cholestasis effect of each tanshinones were studied in mice, the hepatic concentrations of BAs and tanshinones were measured and analyzed as well. The effect of tanshinones on Cyp3a11 protein expression was investigated. DTI exhibited inhibitory effect on the hydroxylation of lithocholic acid (LCA), taurolithocholic acid (TLCA) and taurochenodeoxycholic acid (TCDCA), their IC50 values were 0.81, 0.36 and 1.29 µM, respectively. The hydroxylation of LCA, TLCA and TCDCA were mediated by Cyp3a11. Low-dose DTI, CTS and TSA ameliorated cholestatic liver injury in mice, while high-dose DTI didn't exhibit anti-cholestatic effect. The hepatic BAs profiles indicated that hydroxylation of BAs was inhibited in high-dose DTI group. DTI and TSA up-regulated the protein expression of Cyp3a11. As the hepatic concentration of DTI increased, the inhibitory effect at enzymatic activity level overwhelmed its up-regulation effect at protein level, thus resulted in worsening of cholestasis.

Glutaredoxin-1 alleviates acetaminophen-induced liver injury by decreasing its toxic metabolites.

Excessive N-acetyl-p-benzoquinone imine (NAPQI) formation is a starting event that triggers oxidative stress and subsequent hepatocyte necrosis in acetaminophen (APAP) overdose caused acute liver failure (ALF). S-glutathionylation is a reversible redox post-translational modification and a prospective mechanism of APAP hepatotoxicity. Glutaredoxin-1 (Glrx1), a glutathione-specific thioltransferase, is a primary enzyme to catalyze deglutathionylation. The objective of this study was to explored whether and how Glrx1 is associated with the development of ALF induced by APAP. The Glrx1 knockout mice (Glrx1-/-) and liver-specific overexpression of Glrx1 (AAV8-Glrx1) mice were produced and underwent APAP-induced ALF. Pirfenidone (PFD), a potential inducer of Glrx1, was administrated preceding APAP to assess its protective effects. Our results revealed that the hepatic total protein S-glutathionylation (PSSG) increased and the Glrx1 level reduced in mice after APAP toxicity. Glrx1-/- mice were more sensitive to APAP overdose, with higher oxidative stress and more toxic metabolites of APAP. This was attributed to Glrx1 deficiency increasing the total hepatic PSSG and the S-glutathionylation of cytochrome p450 3a11 (Cyp3a11), which likely increased the activity of Cyp3a11. Conversely, AAV8-Glrx1 mice were defended against liver damage caused by APAP overdose by inhibiting the S-glutathionylation and activity of Cyp3a11, which reduced the toxic metabolites of APAP and oxidative stress. PFD precede administration upregulated Glrx1 expression and alleviated APAP-induced ALF by decreasing oxidative stress. We have identified the function of Glrx1 mediated PSSG in liver injury caused by APAP overdose. Increasing Glrx1 expression may be investigated for the medical treatment of APAP-caused hepatic injury.

Effects of Gelsemium elegans Combined with Mussaenda pubescens on Transporter BCRP and Drug-metabolizing Enzyme CYP3A11 / 中国实验方剂学杂志

ObjectiveTo explore the effect of Gelsemium elegans combined with Mussaenda pubescens on efflux transporter breast cancer resistance protein （BCRP） and cytochrome P450 3A11 （CYP3A11） and their attenuation mechanism， and to investigate whether the nuclear receptors were involved in such regulation by intervening it with nuclear receptor activators. MethodC57BL/6 mice were divided into the blank group， G. elegans （GE， 0.25 g·kg-1）group， GE + M. pubescens （MP） （0.25 g·kg-1+10 g·kg-1） group， GE + pregnane X receptor （PXR） activator （rifampicin）（GE + Rif，0.25 g·kg-1+50 mg·kg-1） group， GE + MP + Rif （0.25 g·kg-1+10 g·kg-1+50 mg·kg-1） group， GE + constitutive androstane receptor （CAR） activator （1，4-Bis ［2-（3，5-Dichloropyridyloxy）］ benzene， TCPOBOP）（GE + TCP， 0.25 g·kg-1+0.5 mg·kg-1） group， and GE + MP + TCP （0.25 g·kg-1+10 g·kg-1+0.5 mg·kg-1） group. The medication lasted for 14 successive days. One hour after the last administration， the mice were sacrificed by cervical dislocation and the liver tissue was harvested. The left liver tissue was stained with hematoxylin- eosin （HE） for observing the pathological changes. The right liver tissue was used for BCRP and CYP3A11 mRNA and protein expression detection by real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） and Western blot. ResultThe survival rates of mice in the GE + Rif group， GE group， and GE + MP group were 25% （the lowest）， 40%， and 80%， respectively， and no death was observed in the other groups. Compared with the obvious lesions in the liver cells of the GE group， the pathological changes in liver cells of the GE + MP group were alleviated， while those in the GE + Rif group were worsened. Compared with the GE + Rif group， the GE + MP + Rif group exhibited relieved pathological changes in liver cells. Both the GE + TCP group and the GE + MP + TCP group showed mild liver lesions. The comparison with the GE + MP group revealed that the pathological changes in the GE + MP + TCP group were slightly relieved. Compared with the blank group， the expression of BCRP protein and mRNA in GE group were significantly decreased （P<0.05，P<0.01）.The expression of CYP3A11 protein in GE group were significantly decreased （P<0.01）. Compared with the GE group， the GE + MP group displayed remarkably up-regulated BCRP protein and mRNA expression （P<0.05，P<0.01） and CYP3A11 protein expression （P<0.05）， but slightly up-regulated CYP3A11 mRNA expression. Compared with the GE group， the GE + Rif group exhibited down-regulated BCRP protein expression （P < 0.05）. The protein and mRNA expression levels of BCRP were lower in the GE + MP + Rif group than in the GE + MP group （P<0.05，P<0.01）. The PXR activator rifampicin regulated BCRP before and after the combination of G. elegans with M. pubescens. The CYP3A11 protein and mRNA expression levels in the GE + TCP group were higher than those in the GE group （P<0.05，P<0.01）. Compared with the GE + MP group， the GE + MP + TCP group showed up-regulated CYP3A11 protein and mRNA expression （P<0.05，P<0.01）. CAR activator TCPOBOP also had a regulatory effect on CYP3A11 before and after the compatibility of G. elegans with M. pubescens. ConclusionThe attenuated toxin after the combination of G. elegans with M. pubescens is closely related to the efflux transporter BCRP and the drug-metabolizing enzyme CYP3A11.

Diurnal hepatic CYP3A11 contributes to chronotoxicity of the pyrrolizidine alkaloid retrorsine in mice.

1. Retrorsine (RTS) is a pyrrolizidine alkaloid (distributed in many medicinal plants) that has significant hepatotoxicity. Here, we aimed to determine the daily variations in RTS hepatotoxicity (chronotoxicity) in mice, and to investigate the role of metabolism in generating RTS chronotoxicity.2. Acute toxicity and pharmacokinetic studies were performed with mice after RTS administration at different times of the day. Hepatotoxicity was assessed by measuring plasma ALT (alanine aminotransferase) and AST (aspartate aminotransferase) levels. mRNA and proteins were determined by qPCR and Western blotting, respectively. Time-dependent in vitro metabolism of RTS was assessed by using mouse liver microsomes.3. We found that RTS toxicity was more severe in the dark phase (zeitgeber time 14 or ZT14 and ZT18) than in the light phase (ZT2 and ZT6). This chronotoxicity was associated with a dosing time difference in the systemic exposures of RTS and a pyrrolic ester metabolite (a cause of hepatotoxicity, measured by the levels of pyrrole-GSH conjugate and pyrrole-protein adducts due to a high chemical reactivity). Moreover, the CYP3A11 (a major enzyme for RTS bioactivation) inhibitor ketoconazole decreased the production of pyrrole-GSH conjugate and abrogated diurnal rhythm in RTS metabolism. In addition, E4bp4 (a circadian regulator of Cyp3a11) ablation abolished the rhythm of CYP3A11 expression and abrogated the dosing time-dependency of RTS toxicity.4. In conclusion, RTS chronotoxicity in mice was attributed to time-varying hepatic metabolism regulated by the circadian clock. Our findings have implications for reducing pyrrolizidine alkaloid-induced toxicity via a chronotherapeutic approach.

PXR is a target of (-)-epicatechin in skeletal muscle.

(-)-Epicatechin (EC) is a flavanol that has shown numerous biological effects such as: decrease risk of cardiovascular dysfunction, metabolism regulation, skeletal muscle (SkM) performance improvement and SkM cells differentiation induction, among others. The described EC acceptor/receptor molecules do not explain the EC's effect on SkM. We hypothesize that the pregnane X receptor (PXR) can fulfill those characteristics, based on structural similitude between EC and steroidal backbone and that PXR activation leads to similar effects as those induced by EC. In order to demonstrate our hypothesis, we: 1) analyzed the possible EC and mouse PXR interaction through in silico strategies, 2) developed an EC's affinity column to isolate PXR, 3) evaluated, in mouse myoblast (C2C12 cells) the inhibition of EC-induced PXR's nucleus translocation by ketoconazole, a specific blocker of PXR and 4) analyzed the effect of EC as an activator of mouse PXR, evaluating the expression modulation of cytochrome 3a11 (Cyp3a11) gen and myogenin protein. (-)-Epicatechin interacts and activates PXR, promoting this protein translocation to the nucleus, increasing the expression of Cyp3a11, and promoting C2C12 cell differentiation through increasing myogenin expression. These results can be the base of further studies to analyze the possible participation of PXR in the skeletal muscle effects shown by EC.

Circadian clock regulates hepatotoxicity of Tripterygium wilfordii through modulation of metabolism.

OBJECTIVES: We aimed to determine the diurnal rhythm of Tripterygium wilfordii (TW) hepatotoxicity and to investigate a potential role of metabolism and pharmacokinetics in generating chronotoxicity. METHODS: Hepatotoxicity was determined based on assessment of liver injury after dosing mice with TW at different circadian time points. Circadian clock control of metabolism, pharmacokinetics and hepatotoxicity was investigated using Clock-deficient (Clock-/- ) mice. KEY FINDINGS: Hepatotoxicity of TW displayed a significant circadian rhythm (the highest level of toxicity was observed at ZT2 and the lowest level at ZT14). Pharmacokinetic experiments showed that oral gavage of TW at ZT2 generated higher plasma concentrations (and systemic exposure) of triptolide (a toxic constituent) compared with ZT14 dosing. This was accompanied by reduced formation of triptolide metabolites at ZT2. Loss of Clock gene sensitized mice to TW-induced hepatotoxicity and abolished the time-dependency of toxicity that was well correlated with altered metabolism and pharmacokinetics of triptolide. Loss of Clock gene also decreased Cyp3a11 expression in mouse liver and blunted its diurnal rhythm. CONCLUSIONS: Tripterygium wilfordii chronotoxicity was associated with diurnal variations in triptolide pharmacokinetics and circadian expression of hepatic Cyp3a11 regulated by circadian clock. Our findings may have implications for improving TW treatment outcome with a chronotherapeutic approach.

Cyp3a11 metabolism-based chronotoxicity of brucine in mice.

Brucine is one of the main bioactive and toxic constituents of the herb drug Semen Strychni. Here we aimed to determine dosing time-dependent hepatotoxicity of brucine, and to investigate the role of metabolism in generation of brucine chronotoxicity. Brucine was administered to wild-type or Npas2-/- (a clock disrupted model) mice at different circadian time points for toxicity and pharmacokinetic characterization. The hepatotoxicity was evaluated by plasma alanine aminotransferase and aspartate aminotransferase measurements and histopathological analysis. The role of Cyp3a11 in brucine metabolism was determined by chemical inhibition assays and Cyp3a11-overexpressing HEK293 cells. Hepatic circadian Cyp3a11 mRNA and protein levels were determined by qPCR and Western blotting, respectively. The toxicity of brucine was more severe in the light phase [Zeitgeber time (ZT) 2 and ZT8] than in the dark phase (ZT14 and ZT20). Chemical inhibition and substrate metabolism assays suggested Cyp3a11 as a significant contributor to brucine metabolism. The Cyp3a11 mRNA, protein and activity in the livers of wild-type mice displayed significant circadian fluctuations. Npas2 ablation markedly down-regulated Cyp3a11 mRNA, protein and activity, and abrogated their circadian rhythms. The circadian time differences in brucine pharmacokinetics and liver distribution were lost in Npas2-/- mice, so were the time differences in brucine hepatotoxicity. In conclusion, chronotoxicity of brucine was determined by circadian variations in Cyp3a11 metabolism. The findings have implications in improving brucine (and possibly Semen Strychni) efficacy via dosing time optimization.

Circadian Cyp3a11 metabolism contributes to chronotoxicity of hypaconitine in mice.

Hypaconitine is an active and highly toxic constituent derived from Aconitum species. Here we aimed to determine the chronotoxicity of hypaconitine in mice, and to investigate a potential role of metabolism in hypaconitine chronotoxicity. Cardiac toxicity was assessed by measuring CK (creatine kinase) and LDH (lactate dehydrogenase) levels after hypaconitine administration to wild-type and Bmal1-/- (a clock disrupted model) mice at different times of day. The mRNA and protein levels of Cyp3a11 in mouse livers were determined by qPCR and western blotting, respectively. In vitro metabolism was assessed using liver microsomes. Pharmacokinetic study of hypaconitine was performed with wild-type mice. We observed injection time-dependent toxicity (i.e., a more severe toxicity during the light phase than the dark phase) for hypaconitine in mice. The chronotoxicity was attributed to a difference in systemic exposure of hypaconitine caused by time of day-dependent metabolism. Furthermore, circadian metabolism of hypaconitine was accounted for by the diurnal expression of Cyp3a11, a major enzyme for hypaconitine detoxification in the liver. Moreover, Bmal1 ablation in mice abolished the daily rhythm of Cyp3a11 expression and abrogated the time-dependency of hypaconitine toxicity. In conclusion, circadian Cyp3a11 metabolism contributed to chronotoxicity of hypaconitine in mice. This metabolism-based chronotoxicity would facilitate the formulation of best timing for drug administration.

The transcription factor E4bp4 regulates the expression and activity of Cyp3a11 in mice.

Human CYP3A4 (Cyp3a11 in mice) is one of the most important enzymes for drug metabolism and detoxification. Here, we aimed to investigate a potential role for E4bp4 in regulation of Cyp3a11 expression and activity. The regulatory effects of E4bp4 on Cyp3a11 enzyme were assessed using E4bp4-/- mice and Hepa-1c1c7 cells. The mRNA and protein levels were quantified using qPCR and Western blotting, respectively. In vitro microsomal Cyp3a11 activity was probed using its specific substrates midazolam and testosterone. Pharmacokinetic studies were performed with wild-type and E4bp4-/- mice after midazolam administration. Global deletion of E4bp4 led to significant upregulation of Cyp3a11 mRNA and protein in major metabolic organs (i.e., the liver, kidney and small intestine). E4bp4 ablation also caused an increased microsomal Cyp3a11 activity consistent with the enzyme's expression change. Overexpression of E4bp4 in Hepa-1c1c7 cells resulted in reduced levels of Cyp3a11 mRNA and protein, whereas E4bp4 knockdown caused upregulation of Cyp3a11 expression. In addition, the systemic exposure of midazolam was lowered in E4bp4-/- mice compared with wild-type mice. This was accompanied by enhanced formation of its metabolite 1'-hydroxymidazolam. Furthermore, luciferase reporter and mobility shift assays revealed that E4bp4 repressed Cyp3a11 transcription via direct binding to C-site (-1539/-1529 bp) in the promoter region. In conclusion, E4bp4 negatively regulates Cyp3a11 expression, thereby impacting drug metabolism and pharmacokinetics.

Liver-specific knockout of histone methyltransferase G9a impairs liver maturation and dysregulates inflammatory, cytoprotective, and drug-processing genes.

Methyltransferase G9a is essential for a key gene silencing mark, histone H3 dimethylation at lysine-9 (H3K9me2). Hepatic G9a expression is down-regulated by xenobiotics and diabetes. However, little is known about the role of G9a in liver. Thus, we generated mice with liver-specific knockout (Liv-KO) of G9a. Adult G9a Liv-KO mice had marked loss of H3K9me2 proteins in liver, without overt liver injury or infiltration of inflammatory cells. However, G9a-null livers had ectopic induction of certain genes normally expressed in neural and immune systems. Additionally, G9a-null livers had moderate down-regulation of cytoprotective genes, markedly altered expression of certain important drug-processing genes, elevated endogenous reactive oxygen species, induction of ER stress marker Chop, but decreased glutathione and nuclear Nrf2. microRNA-383, a negative regulator of the PI3K/Akt pathway, was strongly induced in G9a Liv-KO mice. After LPS treatment, G9a Liv-KO mice had aggravated lipid peroxidation and proinflammatory response. Taken together, the present study demonstrates that G9a regulates liver maturation by silencing neural and proinflammatory genes but maintaining/activating cytoprotective and drug-processing genes, in which the G9a/miR-383/PI3K/Akt/Nrf2 (Chop) pathways may play important roles. G9a deficiency due to genetic polymorphism and/or environmental exposure may alter xenobiotic metabolism and aggravate inflammation and liver dysfunction.

Zonation of the drug-metabolizing enzyme cytochrome P450 3A in infant mice begins in pre-weaning period.

The drug-metabolizing enzyme CYP3A is a heterogeneous enzyme found in the liver that displays local characteristics referred to as "zonation." Zonation contributes to improved energy efficiency in metabolism. The objective of this study was to determine a scientific basis for the safety of fetuses and nursing infants in cases in which the use of pharmaceuticals by pregnant and nursing mothers is unavoidable. In addition, we analyzed CYP3A zonation in the liver using mice from the fetus stage to the nursing stage. The livers of mice ranging from day 13.5 of the fetal stage to day 7 of the nursing stage were resected and immunostained using rabbit anti-rat CYP3A2 Ab, which can detect CYP3A11, CYP3A13, CYP3A16, CYP3A25, CYP3A41 and CYP3A44. The results indicated that zonation did not begin in the fetus stage up to day 3 of the nursing stage, and began on day 7 of infancy. This study revealed that changes in the metabolic activity of CYP3A in the liver between the fetal and nursing stages are partly related to zonation. Further studies are needed to establish standards for the proper use of pharmaceuticals by pregnant and nursing mothers.

Relationship between low midazolam metabolism by cytochrome P450 3A in mice and the high incidence of birth defects.

The use of midazolam in early stages of pregnancy has resulted in a high incidence of birth defects; however, the underlying reason is unknown. We investigated expression changes of the CYP3A molecular species and focused on its midazolam metabolizing activity from the foetal period to adulthood. CYP3A16 was the only CYP3A species found to be expressed in the liver during the foetal period. However, CYP3A11 is upregulated in adult mice, but has been found to be downregulated during the foetal period and to gradually increase after birth. When CYP3A16 expression was induced in a microsomal fraction of cells used to study midazolam metabolism by CYP3A16, its activity was suppressed. These results showed that the capacity to metabolize midazolam in the liver during the foetal period is very low, which could hence result in a high incidence of birth defects associated with the use of midazolam during early stages of pregnancy.

Identification of Taurine-Responsive Genes in Murine Liver Using the Cdo1-Null Mouse Model.

The cysteine dioxygenase (Cdo1)-null mouse is unable to synthesize hypotaurine and taurine by the cysteine/cysteine sulfinate pathway and has very low taurine levels in all tissues. The lack of taurine is associated with a lack of taurine conjugation of bile acids, a dramatic increase in the total and unconjugated hepatic bile acid pools, and an increase in betaine and other molecules that serve as organic osmolytes. We used the Cdo1-mouse model to determine the effects of taurine deficiency on expression of proteins involved in sulfur amino acid and bile acid metabolism. We identified cysteine sulfinic acid decarboxylase (Csad), betaine:homocysteine methytransferase (Bhmt), cholesterol 7α-hydroxylase (Cyp7a1), and cytochrome P450 3A11 (Cyp3a11) as genes whose hepatic expression is strongly regulated in response to taurine depletion in the Cdo1-null mouse. Dietary taurine supplementation of Cdo1-null mice restored hepatic levels of these four proteins and their respective mRNAs to wild-type levels, whereas dietary taurine supplementation had no effect on abundance of these proteins or mRNAs in wild-type mice.

Colonization by non-pathogenic bacteria alters mRNA expression of cytochromes P450 in originally germ-free mice.

Gut microbiota provides a wide range of beneficial function for the host and has an immense effect on the host's health state. It has also been shown that gut microbiome is often involved in the biotransformation of xenobiotics; however, the molecular mechanisms of the interaction between the gut bacteria and the metabolism of drugs by the host are still unclear. To investigate the effect of microbial colonization on messenger RNA (mRNA) expression of liver cytochromes P450 (CYPs), the main drug-metabolizing enzymes, we used germ-free (GF) mice, lacking the intestinal flora and mice monocolonized by non-pathogenic bacteria Lactobacillus plantarum NIZO2877 or probiotic bacteria Escherichia coli Nissle 1917 compared to specific pathogen-free (SPF) mice. Our results show that the mRNA expression of Cyp1a2 and Cyp2e1 was significantly increased, while the expression of Cyp3a11 mRNA was decreased under GF conditions compared to the SPF mice. The both bacteria L. plantarum NIZO2877 and E. coli Nissle 1917 given to the GF mice decreased the level of Cyp1a2 mRNA and normalized it to the control level. On the other hand, the colonization by these bacteria had no effect on the expression of Cyp3a11 mRNA in the liver of the GF mice (which remained decreased). Surprisingly, monocolonization with chosen bacterial strains has shown a different effect on the expression of Cyp2e1 mRNA in GF mice. Increased level of Cyp2e1 expression observed in the GF mice was found also in mice colonized by L. plantarum NIZO2877; however, the colonization with probiotic E. coli Nissle 1917 caused a decrease in Cyp2e1 expression and partially restored the SPF mice conditions.

Effect of Gehua Jiejue Dizhi decoction on the liver fatty deposition and expression of PXR in themousealcoholic fatty liver / 中国比较医学杂志

Objective To explore the effect of a herbalcompound Gehua Jiejue Dizhi Decoction (GJDD) on the liver fat deposition and the expression of PXR, and the mRNA and protein expression of its target genes CYP3A11 and CYP3A25in the liver tissues of mouse models of alcoholic fatty liver.Methods Twenty-nine healthy male C57BL/6J mice were randomly divided into control group (n=5), model group (n=8), high dose GJDD group (n=8)and low dose GJDD group (n=8).The mouse model of alcoholic fatty liver was prepared according to the National Institute on Alcohol Abuse and Alcoholism (NIAAA) method.Then, the mice were treated with the high dose and low dose GJDD for 9 days.Serum glutamic-pyruvic transaminase (AST) and aspartate aminotransferase (AST) were detected by enzyme-linked immunosorbent assay (ELISA).Liver fat deposition was detected by oil red O staining.Real-time RT-PCR and immunohistochemistry were performed to examine the expressions of PXR, CYP3A11 and CYP3A25.Results Compared with the model group, the liver fat deposition in the intervention groups was significantly reduced in a dose-dependent manner, with a significant increase of the expression of PXR and CYP3A25 (P < 0.01).The serum ALT level was significantly reduced in the model group (P < 0.01), while the transcriptional levels of CYP3A11 mRNA in the groups were similar (P ≥ 0.05).Conclusions Gehua Jiejue Dizhi Decoction has obvious therapeutic effect on the AFLD in mice, which may be related to the activation of PXR and its target genes CYP3A25.

Effect of Cyp27A1 gene dosage on atherosclerosis development in ApoE-knockout mice.

In humans, sterol 27-hydroxylase (CYP27A1) deficiency leads to cholesterol deposition in tendons and vasculature. Thus, in addition to its role in bile acid synthesis, where it converts cholesterol to 27-hydroxycholesterol (27-OHC), CYP27A1 may also be atheroprotective. Cyp27A1-deficient (Cyp27A1(-/-)) mice were crossed with apolipoprotein E (apoE)-deficient mice. Cyp27A1(+/+)/apoE(-/-) [ApoE-knockout (KO)], Cyp27A1(+/-)/apoE(-/-) heterozygous (het), and Cyp27A1(-/-)/apoE(-/-) [double-knockout (DKO)] mice were challenged with a Western diet (WD) for 3 and 6 mo. ApoE-KO mice fed a chow diet or a WD were used as the control. The severity of atherosclerosis in DKO mice was reduced 10-fold. Compared with the control, the DKO mice had no 27-OHC, total plasma cholesterol and low-density lipoprotein and very low density lipoprotein (LDL/VLDL) concentrations were reduced 2-fold, and HDL was elevated 2-fold. Expression of hepatic CYP7A1, CYP3A, and CYP8B1 were 5- to 10-fold higher. 3-Hydroxy-3-methyl-glutaryl-CoA reductase (HMGR) activity increased 4-fold. Fecal cholesterol was increased. In contrast, het mice fed a WD developed accelerated atherosclerosis and severe skin lesions, possibly because of reduced reverse cholesterol transport due to diminished 27-OHC production. CYP27A1 activity is involved in the control of cholesterol homeostasis and development of atherosclerosis with a distinct gene dose-dependent effect.