Effect of passive smoking exposure on risk of type 2 diabetes: a systematic review and meta-analysis of prospective cohort studies.

Objective: The effect of passive smoking exposure on the risk of type 2 diabetes has not been systematically studied. A meta-analysis was conducted to assess the association between passive smoking exposure and the risk of diabetes. Methods: We searched three major databases up to 31 October 2022 to identify relevant prospective cohort studies on the association between passive smoking and the risk of type 2 diabetes. The pooled relative risk (RR) and 95% confidence interval (CI) for the association between passive smoking exposure and the risk of type 2 diabetes were analyzed using a fixed-effect model. Results: Ten prospective cohort studies were included in this meta-analysis, with a total of 251,620 participants involved. The pooled RR showed a significantly positive association between nonsmokers exposed to passive smoking and type 2 diabetes as compared to non-smokers who were not exposed to passive smoking [RR = 1.27; 95% CI (1.19, 1.36); p < 0.001]. Sensitivity analysis indicated that the pooled RR was not substantially affected by any of the individual studies. Conclusion: Exposure to passive smoking increases the risk of type 2 diabetes. This study may have a positive effect on the prevention of type 2 diabetes. Systematic review registration: https://www.crd.york.ac.uk/PROSPERO/, identifier CRD42023372532.

The Association of Homocysteine and Diabetic Retinopathy in Homocysteine Cycle in Chinese Patients With Type 2 Diabetes.

Objective: This study aimed to explore the relationship between homocysteine (Hcy) and diabetic retinopathy (DR) and the impacts of the Hcy pathway on this relationship against this background. Methods: This study retrieved 1979 patients with type 2 diabetes (T2D) from the First Affiliated Hospital of Liaoning Medical University in Jinzhou, Liaoning Province, China. Multiple logistic regression was used to analyze the effects of Hcy cycle on the relationship between Hcy and DR. Spearman's rank correlation analysis was used to analyze the correlation between risk factors related to DR progression and Hcy. Finally, the results of logistic regression were supplemented by mediation analysis. Results: We found there was a negative correlation between low concentration of Hcy and DR (OR : 0.83, 95%CI: 0.69-1). After stratifying all patients by cysteine (Cys) or Methionine (Met), this relationship remained significant only in low concentration of Cys (OR: 0.75, 95%CI: 0.61-0.94). Through the RCS curve, we found that the effect of Hcy on DR presents a U-shaped curve relationship. Mediating effect in Met and Hcy cycles was also significant [Total effect c (OR: 0.968, 95%CI: 0.938-0.998), Direct effect path c' (OR: 0.969, 95%CI: 0.940-0.999), Path a (OR: 1.047, 95%CI: 1.004-1.091), Path b (OR: 0.964, 95%CI: 0.932-0.998)]. Conclusions: The relationship between Hcy and DR presents a U-shaped curve and the homocysteine cycle pathway has an impact on it. And too low concentration of Hcy indicates a lack of other substances, such as vitamins. It is suggested that the progression of DR is the result of a combination of many risk factors. Further prospective studies are needed to determine the role of Hcy in the pathogenesis of DR.

Hepatic miR-378 modulates serum cholesterol levels by regulating hepatic bile acid synthesis.

Rationale: An improved understanding of thyroid hormone (TH) action on cholesterol metabolism will facilitate the identification of novel therapeutic targets for hypercholesterolemia. TH-regulated microRNAs (miRNAs) have been implicated in TH-controlled biological processes; however, whether and how TH-regulated miRNAs mediate the cholesterol-lowering effect of TH remains unclear. Our aim was to identify TH-regulated microRNAs that have cholesterol-lowering effects and explore the underlying mechanism. Method: Microarray and RNA-seq were performed to identify TH-regulated microRNAs and the genes regulated by mmu-miR-378-3p (miR-378) in the liver of mice, respectively. Recombinant adenoviruses encoding miR-378, Mafg, and shRNA for Mafg, antagomiR-378, liver-specific miR-378 transgenic mice, and miR-378 knockout mice were employed to investigate the roles of hepatic miR-378 and MAFG in cholesterol and bile acid homeostasis. The levels of bile salt species were determined by using UFLC-Triple-time of flight/MS. Results: Here, we show that hepatic miR-378 is positively regulated by TH. Transient overexpression of miR-378 in the liver of mice reduces serum cholesterol levels, accompanied with an increase in the expression of key enzymes in primary bile acid synthetic pathways and corresponding increases in biliary and fecal bile acid levels. Consistently, liver-specific miR-378 transgenic mice with moderate overexpression of hepatic miR-378 display decreased serum cholesterol levels and resistance to diet-induced hypercholesterolemia, while mice lacking miR-378 exhibit defects in bile acid and cholesterol homeostasis. Mechanistically, hepatic miR-378 regulates the expression of key enzymes in both classic and alternative bile acid synthetic pathways through MAFG, a transcriptional repressor, thereby modulating bile acid and cholesterol metabolism. Conclusions: TH-responsive hepatic miR-378 is capable of modulating serum cholesterol levels by regulating both the classic and alternative BA synthetic pathways. Our study not only identifies a previously undescribed role of hepatic miR-378 but also provides new cholesterol-lowering approaches.

Trimethylamine N-Oxide Metabolites in Early Pregnancy and Risk of Gestational Diabetes: A Nested Case-Control Study.

OBJECTIVES: This study aimed to investigate the associations between trimethylamine N-oxide (TMAO) and related metabolites in early pregnancy and the risk of gestational diabetes mellitus (GDM). DESIGN: A prospective cohort of 22,302 pregnant women from 2010 to 2012 in Tianjin, China, was used to perform a nested case-control study. A total of 243 women with GDM and 243 women without GDM matched by maternal age (±1 year) were used as cases and controls, respectively. Conditional logistic regression and restricted cubic spline were used to examine the full-range risk associations between individual TMAOs metabolites at the first antenatal care visit with GDM. Trimethylamine conversion ratio (TMAR) was defined as trimethylamine (TMA)/its precursors, and trimethylamine N-oxide conversion ratio (TMAOR) was defined as TMAO/TMA. An additive interaction between high TMAR and low TMAOR indicates a state of TMA accumulation, and a mathematical interaction between high TMAR and high TMAOR indicates accumulation of TMAO. RESULTS: TMA was linearly associated with GDM, whereas TMA precursors and TMAO were inversely associated with GDM with clear threshold effects, i.e., 16 nmol/mL for TMAO, 200 nmol/mL for betaine, 112 nmol/mL for l-carnitine, and 110 and 270 nmol/mL for cholinechloride (a U-shaped relationship). Copresence of TMAR >0.35 and TMAOR ≤0.15 was associated with a markedly higher OR (11.16; 95% CI, 5.45 to 22.8), compared with TMAR >0.35 only (OR = 1.71; 95% CI, 0.42 to 6.95) or TMAOR ≤0.15 only (OR = 2.06; 95% CI, 1.09 to 3.90), with a significant additive interaction. However, the mathematical interaction was nonsignificant. CONCLUSIONS: TMAO metabolites in the early pregnancy were associated with the risk of GDM, whereas TMA was more likely to play a causal role in GDM.

Caffeine Protects Skin from Oxidative Stress-Induced Senescence through the Activation of Autophagy.

Skin cells are vulnerable to oxidative stress-induced senescence, which may lead to abnormal aging or aging-related disorders. Therefore, strategies that can ameliorate oxidative stress-induced senescence are expected to protect skin from damage, holding the promise of treating skin diseases in the clinic. This study aims to investigate whether caffeine, a well-known purine alkaloid, is able to prevent skin from oxidative stress-induced senescence, and to explore the underlying molecular mechanisms. Methods: A free radical inducer 2,2'-Azobis (2-amidinopropane) dihydrochloride (AAPH) was used to induce oxidative stress and cellular senescence in both transformed skin cells and in normal human epidermal keratinocytes (NHEKs). Ultraviolet (UV) irradiation was established as the in vivo oxidative stress model in mouse skin tissues. Cellular senescence was determined by SA ß-galactosidase staining, immunofluorescence and western blotting. Activation of autophagy was confirmed by western blotting, immunofluorescence, and transmission electron microscopy. Reactive oxygen species (ROS) detection by commercial kits, gene knockdown by RNA interference (RNAi) and receptor activation/inactivation by agonist/antagonist treatment were applied in mechanistic experiments. Results: We report that AAPH induced senescence in both transformed skin cells and in NHEKs. Similarly, UV irradiation induced senescence in mouse skin tissues. Remarkably, low dose of caffeine (<10 µM) suppressed cellular senescence and skin damage induced by AAPH or UV. Mechanistically, caffeine facilitated the elimination of ROS by activating autophagy. Using a combination of RNAi and chemical treatment, we demonstrate that caffeine activates autophagy through a series of sequential events, starting from the inhibition of its primary cellular target adenosine A2a receptor (A2AR) to an increase in the protein level of Sirtuin 3 (SIRT3) and to the activation of 5' adenosine monophosphate-activated protein kinase (AMPK). Oral administration of caffeine increased the protein level of SIRT3, induced autophagy, and reduced senescence and tissue damage in UV-irradiated mouse skin. On the other hand, co-administration with autophagy inhibitors attenuated the protective effect of caffeine on UV-induced skin damage in mice. Conclusion: The results reveal that caffeine protects skin from oxidative stress-induced senescence through activating the A2AR/SIRT3/AMPK-mediated autophagy. Our study not only demonstrated the beneficial effect of caffeine using both in vitro and in vivo models, but also systematically investigated the underlying molecular mechanisms. These discoveries implicate the potential of caffeine in the protection of skin disease.

Growth arrest and DNA damage-inducible 45α protects against nonalcoholic steatohepatitis induced by methionine- and choline-deficient diet.

Growth arrest and DNA damage-inducible 45 α (Gadd45α) is a stress-inducible protein that plays an important role in cell survival/death and DNA repair, but its contribution to the development of nonalcoholic steatohepatitis (NASH) has not been investigated. C57BL/6 Gadd45a-null and wild-type (WT) mice were treated with a methionine and choline-deficient diet (MCD) for eight weeks and phenotypic changes examined. Gadd45a-null mice had more severe hepatic inflammation and fibrosis, higher levels of mRNAs encoding pro-inflammatory, pro-fibrotic, and pro-apoptotic proteins, and greater oxidative and endoplasmic reticulum (ER) stress compared with WT mice. Indeed, Gadd45a mRNA was induced in response to ER stress in primary hepatocytes. Lipidomic analysis of NASH livers demonstrated decreased triacylglycerol (TG) in MCD-treated Gadd45a-null mice, which was associated with increased mRNAs encoding phospholipase D (Pld1/2), phosphatidic acid phosphatase type 2A, and choline/ethanolamine phosphotransferase 1 (Cept1), involved in the phosphatidylcholine-phosphatidic acid-diacylglycerol cycle, and decreased mRNAs encoding fatty acid (FA)-binding protein 1 (Fabp1) and FA transport protein 5. Treatment of cultured primary hepatocytes with tumor necrosis factor α, transforming growth factor ß, and hydrogen peroxide led to the corresponding induction of Fabp1, Pld1/2, and Cept1 mRNAs. Collectively, Gadd45α plays protective roles against MCD-induced NASH likely due to attenuating cellular stress and ensuing inflammatory signaling. These results also suggest an interconnection between hepatocyte injury, inflammation and disrupted glycerophospholipid/FA metabolism that yields a possible mechanism for decreased TG accumulation with NASH progression (i.e., "burned-out" NASH).

Role of the lipid-regulated NF-κB/IL-6/STAT3 axis in alpha-naphthyl isothiocyanate-induced liver injury.

Alpha-naphthyl isothiocyanate (ANIT)-induced liver damage is regarded as a useful model to study drug-induced cholestatic hepatitis. Ultra-performance liquid chromatography coupled with electrospray ionization quadrupole mass spectrometry (UPLC-ESI-QTOF MS)-based metabolomics revealed clues to the mechanism of ANIT-induced liver injury, which facilitates the elucidation of drug-induced liver toxicity. 1-Stearoyl-2-hydroxy-sn-glycero-3-phosphocholine (LPC 18:0) and 1-oleoyl-2-hydroxy-sn-glycero-3-phosphocholine (LPC 18:1) were significantly increased in serum from ANIT-treated mice, and this increase resulted from altered expression of genes encoding the lipid metabolism enzymes Chka and Scd1. ANIT also increased NF-κB/IL-6/STAT3 signaling, and in vitro luciferase reporter gene assays revealed that LPC 18:0 and LPC 18:1 can activate NF-κB in a concentration-dependent manner. Activation of PPARα through feeding mice a Wy-14,643-containing diet (0.1%) reduced ANIT-induced liver injury, as indicated by lowered ALT and AST levels, and liver histology. In conclusion, the present study demonstrated a role for the lipid-regulated NF-κB/IL-6/STAT3 axis in ANIT-induced hepatotoxicity, and that PPARα may be a potential therapeutic target for the prevention of drug-induced cholestatic liver injury.

Irinotecan (CPT-11)-induced elevation of bile acids potentiates suppression of IL-10 expression.

Irinotecan (CPT-11) is a first-line anti-colon cancer drug, however; CPT-11-induced toxicity remains a key factor limiting its clinical application. To search for clues to the mechanism of CPT-11-induced toxicity, metabolomics was applied using ultra-performance liquid chromatography coupled with electrospray ionization quadrupole time-of-flight mass spectrometry. Intraperitoneal injection of 50 mg/kg of CPT-11 induced loss of body weight, and intestine toxicity. Changes in gallbladder morphology suggested alterations in bile acid metabolism, as revealed at the molecular level by analysis of the liver, bile, and ileum metabolomes between the vehicle-treated control group and the CPT-11-treated group. Analysis of immune cell populations further showed that CPT-11 treatment significantly decreased the IL-10-producing CD4 T cell frequency in intestinal lamina propria lymphocytes, but not in spleen or mesenteric lymph nodes. In vitro cell culture studies showed that the addition of bile acids deoxycholic acid and taurodeoxycholic acid accelerated the CPT-11-induced suppression of IL-10 secretion by activated CD4(+) naive T cells isolated from mouse splenocytes. These results showed that CPT-11 treatment caused metabolic changes in the composition of bile acids that altered CPT-11-induced suppression of IL-10 expression.

Metabolic mapping of A3 adenosine receptor agonist MRS5980.

(1S,2R,3S,4R,5S)-4-(2-((5-Chlorothiophen-2-yl)ethynyl)-6-(methylamino)-9H-purin-9-yl)-2,3-dihydroxy-N-methylbicyclo[3.1.0]hexane-1-carboxamide (MRS5980) is an A3AR selective agonist containing multiple receptor affinity- and selectivity-enhancing modifications and a therapeutic candidate drug for many inflammatory diseases. Metabolism-related poor pharmacokinetic behavior and toxicities are a major reason for drug R&D failure. Metabolomics with UPLC-MS was employed to profile the metabolism of MRS5980 and MRS5980-induced disruption of endogenous compounds. Recombinant drug-metabolizing enzymes screening experiment were used to determine the enzymes involved in MRS5980 metabolism. Analysis of lipid metabolism-related genes was performed to investigate the reason for MRS5980-induced lipid metabolic disorders. Unsupervised principal components analysis separated the control and MRS5980 treatment groups in feces, urine, and liver samples, but not in bile and serum. The major ions mainly contributing to the separation of feces and urine were oxidized MRS5980, glutathione (GSH) conjugates and cysteine conjugate (degradation product of the GSH conjugates) of MRS5980. The major ions contributing to the group separation of liver samples were phosphatidylcholines. In vitro incubation experiments showed the involvement of CYP3A enzymes in the oxidative metabolism of MRS5980 and direct GSH reactivity of MRS5980. The electrophilic attack by MRS5980 is a minor pathway and did not alter GSH levels in liver or liver histology, and thus may be of minor clinical consequence. Gene expression analysis further showed decreased expression of PC biosynthetic genes choline kinase a and b, which further accelerated conversion of lysophosphatidylcholine to phosphatidylcholines through increasing the expression of lysophosphatidylcholine acyltransferase 3. These data will be useful to guide rational design of drugs targeting A3AR, considering efficacy, metabolic elimination, and electrophilic reactivity.

Structure-inhibition relationship of podophyllotoxin (PT) analogues towards UDP-glucuronosyltransferase (UGT) isoforms.

UDP-glucuronosyltransferases (UGTs) are involved in the clearance of many important drugs and endogenous substances, and inhibition of UGTs' activity by herbal components might induce severe herb-drug interactions or metabolic disturbances of endogenous substances. The present study aims to determine the inhibition of UGTs' activity by podophyllotoxin derivatives, trying to indicate the potential herb-drug interaction or metabolic influence towards endogenous substances' metabolism. Recombinant UGT isoforms (except UGT1A4)-catalyzed 4-methylumbelliferone (4-MU) glucuronidation reaction and UGT1A4-catalyzed trifluoperazine (TFP) glucuronidation were employed to firstly screen the podophyllotoxin derivatives' inhibition potential. Structure-dependent inhibition behavior of podophyllotoxin derivatives towards UGT isoforms was detected. Inhibition kinetic type and parameter (Ki) were determined for the inhi- bition of podophyllotoxin towards UGT1A1, and competitive inhibition of podophyllotoxin towards UGT1A1 was observed with the inhibition kinetic parameter (Ki) to be 4.0 µM. Furthermore, podophyllotoxin was demonstrated to exert medium and weak inhibition potential towards human liver microsomes (HLMs)-catalyzed SN-38 glucuronidation and estradiol-3-glucuronidation. In conclusion, podophyllotoxin inhibited UGT1A1 activity, indicating potential herb-drug interactions between podophyllotoxin-containing herbs and drugs mainly undergoing UGT1A1-mediated metabolism.

Activation of intestinal human pregnane X receptor protects against azoxymethane/dextran sulfate sodium-induced colon cancer.

The role of intestinal human pregnane X receptor (PXR) in colon cancer was determined through investigation of the chemopreventive role of rifaximin, a specific agonist of intestinal human PXR, toward azoxymethane (AOM)/dextran sulfate sodium (DSS)-induced colon cancer. Rifaximin treatment significantly decreased the number of colon tumors induced by AOM/DSS treatment in PXR-humanized mice, but not wild-type or Pxr-null mice. Additionally, rifaximin treatment markedly increased the survival rate of PXR-humanized mice, but not wild-type or Pxr-null mice. These data indicated a human PXR-dependent therapeutic chemoprevention of rifaximin toward AOM/DSS-induced colon cancer. Nuclear factor κ-light-chain-enhancer of activated B cells-mediated inflammatory signaling was upregulated in AOM/DSS-treated mice, and inhibited by rifaximin in PXR-humanized mice. Cell proliferation and apoptosis were also modulated by rifaximin treatment in the AOM/DSS model. In vitro cell-based assays further revealed that rifaximin regulated cell apoptosis and cell cycle in a human PXR-dependent manner. These results suggested that specific activation of intestinal human PXR exhibited a chemopreventive role toward AOM/DSS-induced colon cancer by mediating anti-inflammation, antiproliferation, and proapoptotic events.

PPARα-dependent exacerbation of experimental colitis by the hypolipidemic drug fenofibrate.

Fibrates, such as fenofibrate, are peroxisome proliferator-activated receptor-α (PPARα) agonists and have been used for several decades as hypolipidemic agents in the clinic. However, contradictory observations exist on the role of fibrates in host response to acute inflammation, with unclear mechanisms. The role of PPARα in colitis was assessed using fenofibrate and Ppara-null mice. Wild-type or Ppara-null mice were subjected to acute colitis under three distinct protocols, dextran sulfate sodium, trinitrobenzenesulfonic acid, and Salmonella Typhi. Serum and colon lipidomics were analyzed to characterize the metabolic profiles by ultra-performance liquid chromatography-coupled with electrospray ionization quadrupole time-of-flight mass spectrometry. Messenger RNAs of PPARα target genes and genes involved in inflammation were determined by qunatitative PCR analysis. Fenofibrate treatment exacerbated inflammation and tissue injury in acute colitis, and this was dependent on PPARα activation. Lipidomics analysis revealed that bioactive sphingolipids, including sphingomyelins (SM) and ceramides, were significantly increased in the colitis group compared with the control group; this was further potentiated following fenofibrate treatment. In the colon, fenofibrate did not reduce the markedly increased expression of mRNA encoding TNFα found in the acute colitis model, while it decreased hydrolysis and increased synthesis of SM, upregulated RIPK3-dependent necrosis, and elevated mitochondrial fatty acid ß-oxidation, which were possibly related to the exacerbated colitis.

Metabolic profiling of praziquantel enantiomers.

Praziquantel (PZQ), prescribed as a racemic mixture, is the most readily available drug to treat schistosomiasis. In the present study, ultra-performance liquid chromatography coupled with electrospray ionization quadrupole time-of-flight mass spectrometry (UPLC-ESI-QTOFMS) based metabolomics was employed to decipher the metabolic pathways and enantioselective metabolic differences of PZQ. Many phase I and four new phase II metabolites were found in urine and feces samples of mice 24h after dosing, indicating that the major metabolic reactions encompassed oxidation, dehydrogenation, and glucuronidation. Differences in the formation of all these metabolites were observed between (R)-PZQ and (S)-PZQ. In an in vitro phase I incubation system, the major involvement of CYP3A, CYP2C9, and CYP2C19 in the metabolism of PZQ, and CYP3A, CYP2C9, and CYP2C19 exhibited different catalytic activity toward the PZQ enantiomers. Apparent Km and Vmax differences were observed in the catalytic formation of three mono-oxidized metabolites by CYP2C9 and CYP3A4 further supporting the metabolic differences for PZQ enantiomers. Molecular docking showed that chirality resulted in differences in substrate location and conformation, which likely accounts for the metabolic differences. In conclusion, in silico, in vitro, and in vivo methods revealed the enantioselective metabolic profile of praziquantel.

Inhibition potential of UDP-glucuronosyltransferases (Ugts) 1A isoforms by the analogue of resveratrol, bakuchiol.

Bakuchiol is a promising anti-tumor candidate with resveratrol-like structure. The present study aims to evaluate the inhibition potential of bakuchiol towards UDP-glucuronosyltransferases (UGT) 1A isoforms. An in vitro incubation system using 4-methylumbelliferone (4-MU) glucuronidation was used to evaluate the inhibition capability of bakuchiol towards UGT1A1, 1A3, 1A6, 1A7, 1A8, 1A9 and 1A10. The glucuronidation of trifluoperazine (TFP) was employed as the probe reaction to determine bakuchiol's inhibition towards UGT1A4. At 1 microM and 10 microM of bakuchiol, no or weak inhibition was observed for all the tested UGT1A isoforms. At 100 microM of bakuchiol, the activity of UGT1A1, 1A3, 1A4, 1A6, 1A7, 1A8, 1A9 and 1A10 was inhibited by -46.2%, 74.7%, 17.8%, 98.7%, 70.4%, 99.2%, 75.8%, and 93.3%, respectively. Further inhibition kinetic behaviour was determined for UGT1A6, 1A8, and 1A10. Both Dixon plot and Lineweaver-Burk plot showed the noncompetitive inhibition of bakuchiol towards all these three UGT isoforms. The inhibition kinetic parameters (Ki) were calculated to be 5.3, 1.8, and 92.6 microM for UGT1A6, 1A8, and 1A10, respectively. In combination with the in vivo exposure of bakuchiol, the high possibility of in vivo inhibition of UGT1A6 and 1A8 was predicted. However, relatively low possibility of in vivo inhibition towards UGT1A10 was predicted due to lower in vivo concentration of bakuchiol than its inhibition parameter (Ki). All these information will be helpful for the R&D of bakuchiol as a promising anti-tumor drug.

Metabolomics reveals trichloroacetate as a major contributor to trichloroethylene-induced metabolic alterations in mouse urine and serum.

Trichloroethylene (TCE)-induced liver toxicity and carcinogenesis is believed to be mediated in part by activation of the peroxisome proliferator-activated receptor α (PPARα). However, the contribution of the two TCE metabolites, dichloroacetate (DCA) and trichloroacetate (TCA) to the toxicity of TCE, remains unclear. The aim of the present study was to determine the metabolite profiles in serum and urine upon exposure of mice to TCE, to aid in determining the metabolic response to TCE exposure and the contribution of DCA and TCA to TCE toxicity. C57BL/6 mice were administered TCE, TCA, or DCA, and urine and serum subjected to ultra-performance liquid chromatography coupled with electrospray ionization quadrupole time-of-flight mass spectrometry (UPLC-ESI-QTOFMS)-based global metabolomics analysis. The ions were identified through searching metabolomics databases and by comparison with authentic standards, and quantitated using multiple reactions monitoring. Quantitative polymerase chain reaction of mRNA, biochemical analysis, and liver histology were also performed. TCE exposure resulted in a decrease in urine of metabolites involved in fatty acid metabolism, resulting from altered expression of PPARα target genes. TCE treatment also induced altered phospholipid homeostasis in serum, as revealed by increased serum lysophosphatidylcholine 18:0 and 18:1, and phosphatidylcholine metabolites. TCA administration revealed similar metabolite profiles in urine and serum upon TCE exposure, which correlated with a more robust induction of PPARα target gene expression associated with TCA than DCA treatment. These data show the metabolic response to TCE exposure and demonstrate that TCA is the major contributor to TCE-induced metabolite alterations observed in urine and serum.

Metabolic map and bioactivation of the anti-tumour drug noscapine.

BACKGROUND AND PURPOSE: Noscapine is a promising anti-tumour agent. The purpose of the present study was to describe the metabolic map and investigate the bioactivation of noscapine. EXPERIMENTAL APPROACH: Ultra-performance liquid chromatography coupled with electrospray ionization quadrupole time-of-flight mass spectrometry-based metabolomics was used to analyse the in vitro incubation mixtures, urine and faeces samples from mice treated with noscapine. Recombinant drug-metabolizing enzymes were employed to identify those involved in noscapine metabolism. Hepatic GSH levels and serum biochemistry were also carried out to determine reactive metabolites of noscapine. KEY RESULTS: Several novel phase I metabolites of noscapine were detected after oral gavage of mice, including an N-demethylated metabolite, two hydroxylated metabolites, one metabolite undergoing both demethylation and cleavage of the methylenedioxy group and a bis-demethylated metabolite. Additionally, several novel glucuronides were detected, and their structures were elucidated through MS/MS fragmentology. Recombinant enzymes screening showed the involvement of several cytochromes P450, flavin-containing mono-oxygenase 1 and the UDP-glucuronosyltransferases UGT1A1, UGT1A3, UGT1A9 and UGT2B7, in noscapine metabolism. In vitro glutathione trapping revealed the existence of an ortho-quinone reactive intermediate formed through further oxidation of a catechol metabolite. However, this bioactivation process of noscapine does not occur in vivo. Similar to this result, altered glutathione levels in liver and serum biochemistry revealed no evidence of hepatic damage, thus indicating that, at least in mice, noscapine does not induce hepatotoxicity through bioactivation. CONCLUSIONS AND IMPLICATIONS: A comprehensive metabolic map and bioactivation evaluation provides important information for the development of noscapine as an anti-tumour drug.

Gossypol exhibits a strong influence towards UDP-glucuronosyltransferase (UGT) 1A1, 1A9 and 2B7-mediated metabolism of xenobiotics and endogenous substances.

Gossypol, the polyphenolic constituent isolated from cottonseeds, has been used as a male antifertility drug for a long time, and has been demonstrated to exhibit excellent anti-tumor activity towards multiple cancer types. The toxic effects of gossypol limit its clinical utilization, and enzyme inhibition is an important facet of this. In the present study, in vitro human liver microsomal incubation system supplemented with UDPGA was used to investigate the inhibition of gossypol towards UGT1A1, 1A9 and 2B7-mediated metabolism of xenobiotics and endogenous substances. Estradiol, the probe substrate of UGT1A1, was selected as representative endogenous substance. Propofol (a probe substrate of UGT1A9) and 3'-azido-3'-deoxythimidine (AZT, a probe substrate of UGT2B7) were employed as representative xenobiotics. The results showed that gossypol noncompetitively inhibits UGT-mediated estradiol-3-glucuronidation and propofol O-glucuronidation, and the inhibition kinetic parameters (K(i)) were calculated to be 34.2 and 16.4 µM, respectively. Gossypol was demonstrated to exhibit competitive inhibition towards UGT-mediated AZT glucuronidation, and the inhibition kinetic parameter (K(i)) was determined to be 14.0 µM. All these results indicated that gossypol might induce metabolic disorders of endogenous substances and alteration of metabolic behaviour of co-administered xenobiotics through inhibition of UGTs' activity.

Reversible inhibition of four important human liver cytochrome P450 enzymes by diethylstilbestrol.

Diethylstilbestrol (DES), a synthetic estrogen clinically used to treat threatened abortion between the 1940s and the 1970s, has been restricted to treat certain cases of prostatic and breast cancer due to its adverse drug responses such as teratogenicity and carcinogenicity. Some reports have demonstrated that the addition of DES to docetaxel could modify tubulin composition and improve response of prostate cancer to chemotherapy. Given that DES might be co-administered with other drugs such as docetaxel, the present study focused on CYP-based drug-drug interaction (DDI). In vitro inhibitory effects of DES on CYP isoforms were investigated, and the results showed that DES could competitively inhibit CYP3A4, CYP2C8, CYP2C9 and CYP2E1. The inhibition constants (Ki) were calculated to be 4.4 microM, 3.0 microM, 5.0 microM and 8.0 microM for CYP3A4, CYP2C9, CYP2E1 and CYP2C8, respectively. Based on peak serum DES level after drip influsion of 500 mg of fosfestrol (DES diphosphate) in patients, [I]/Ki was calculated to be 4.3, 6.2, 3.7 and 2.3 for CYP3A4, CYP2C9, CYP2E1 and CYP2C8, which suggested that DES was likely to induce in vivo DDI through inhibition of these four major CYP isoforms. These results collectively demonstrate that adverse drug responses might exist when DES is co-administered with other drugs.

Cycloartane triterpenoids from Cimicifuga yunnanensis induce apoptosis of breast cancer cells (MCF7) via p53-dependent mitochondrial signaling pathway.

The present study was carried out to investigate the antitumor activity of five cycloartane triterpenoids isolated from Cimicifuga yunnanensis on the breast cancer cell line MCF7 and its corresponding drug resistant subline R-MCF7, including cimigenol-3-O-ß-D-xylopyranoside (compound 1), 25-O-acetylcimigenol-3-O-ß-D-xylopyranoside (compound 2), 25-chlorodeoxycimigenol-3-O-ß-D-xylopyranoside (compound 3), 25-O-acetylcimigenol-3-O-α-L-arabinopyranoside (compound 4) and 23-O-acetylcimigenol-3-O-ß-D-xylopyranoside (compound 5). The results showed that compounds 2-5 have relatively high antitumor activity on both MCF7 and R-MCF7 cells. The involvement of apoptosis as a major cause of cycloartane triterpenoids-induced cell death was further confirmed. The results of RT-PCR showed that compounds 2-5 increased the expression of p53 and bax, which led to the loss of mitochondrial potential and then resulted in the activation of caspase-7. These findings collectively demonstrated that compounds 2-5 induced apoptosis of MCF7 via p53-dependent mitochondrial pathway.

Reversible inhibition of three important human liver cytochrome p450 enzymes by tiliroside.

Tiliroside, an active flavonoid extensively found in many medicinal plants including Helichrysum italicum, Geranium mexicanum and Helianthemum glomeratum, has been demonstrated to exert multiple biological effects including antiinflammatory, antimicrobial, antioxidant and antitumor activities. Cytochrome P450 (CYP) enzymes play an important role in the Phase I oxidation metabolism of a wide range of xenobiotics and inhibition of CYP isoforms might influence the elimination of drugs and induce serious adverse drug response. The inhibition of seven CYP isoforms (CYP3A4, CYP1A2, CYP2A6, CYP2D6, CYP2C9, CYP2C8 and CYP2E1) by tiliroside was investigated using in vitro human liver microsomal incubation assays. The results showed that tiliroside strongly inhibited the activity of CYP3A4 (IC(50) = 9.0 ± 1.7 µm), CYP2C8 (IC(50) = 12.1 ± 0.9 µm) and CYP2C9 (IC(50) = 10.2 ± 0.9 µm) with other CYP isoforms negligibly influenced. Further kinetic analysis showed that inhibition of these three CYP isoforms by tiliroside is best fit to a competitive way. The K(i) value was calculated to be 5.5 µm, 3.3 µm, 9.4 µm for CYP3A4, CYP2C9 and CYP2C8, respectively. The relatively low K(i) values suggested that tiliroside might induce drug-drug interactions with many clinically used drugs which are mainly metabolized by these three CYP isoforms. Therefore, attention should be given to the probable drug-drug interaction between tiliroside-containing herbs and substrates of CYP3A4, CYP2C9 and CYP2C8.