The effect of chronic kidney disease on CYP2B expression and activity in male Wistar rats.

Chronic kidney disease (CKD) is characterized by progressive reduction in kidney function over time. CKD affects greater than 10% of the population and its incidence is on the rise due to the growing prevalence of its risk factors. Previous studies demonstrated CKD alters nonrenal clearance of drugs in addition to reducing renal clearance. We assessed the function and expression of hepatic CYP2B enzymes using a rat model of CKD. CKD was induced in Wistar rats by supplementing their chow with adenine and confirmed through the detection of elevated uremic toxins in plasma. Liver enzymes AST and ALT were unchanged by the adenine diet. Bupropion was used as a probe substrate for hepatic CYP2B function using rat liver microsomes. The resulting metabolite, hydroxy-bupropion, and bupropion were quantified by ultra-performance liquid chromatography coupled to time-of-flight mass spectrometry. Level of mRNA and protein were determined by RT-PCR and Western blot, respectively. The results of our study demonstrate that CYP2B1 is downregulated in a rat model of CKD. CYP2B1 mRNA level was significantly decreased (88%, P < 0.001) in CKD relative to control. Similarly, maximal enzymatic velocity (Vmax) for CYP2B was decreased by 46% in CKD relative to control (P < 0.0001). Previous studies involving patients with CKD demonstrated altered bupropion pharmacokinetics compared to control. Hence, our results suggest that these alterations may be mediated by attenuated CYP2B hepatic metabolism. This finding may partially explain the alterations in pharmacokinetics and nonrenal drug clearance frequently observed in patients with CKD.

Effect of Nicotine on CYP2B1 Expression in a Glioma Animal Model and Analysis of CYP2B6 Expression in Pediatric Gliomas.

Cyclophosphamide (CPA) is a pro-drug commonly used in the chemotherapeutic schemes for glioma treatment but has high toxicity and the side effects include brain damage and even death. Since CPA is activated mainly by CY2B6, over-expression of the enzyme in the tumor cells has been proposed to enhance CPA activation. In this study, we explored the induction of the Cyp2b1 (homologous to CYP2B6) by nicotine in an animal rat model with glioma. Gene expression and protein levels were analyzed by RT-PCR and Western blot. Nicotine treatment increased CYP2B1 protein levels in the healthy animals' brain tissue. In the brain tissue of animals with glioma, the CYP2B1 showed a high expression, even before nicotine treatment. Nicotine did not increase significantly the CYP2B1 protein expression in the tumor, but increased its expression in the tumor vicinity, especially around blood vessels in the cortex. We also explored CY2B6 expression in glioma samples derived from pediatric patients. Tumor tissue showed a variable expression of the enzyme, which could depend on the tumor malignancy grade. Induction of the CYP2B6 in pediatric gliomas with lower expression of the enzyme, could be an alternative to improve the antitumoral effect of CPA treatment.

Use of Phenoxyaniline Analogues To Generate Biochemical Insights into the Interactio n of Polybrominated Diphenyl Ether with CYP2B Enzymes.

Human hepatic cytochromes P450 (CYP) are integral to xenobiotic metabolism. CYP2B6 is a major catalyst of biotransformation of environmental toxicants, including polybrominated diphenyl ethers (PBDEs). CYP2B substrates tend to contain halogen atoms, but the biochemical basis for this selectivity and for species specific determinants of metabolism has not been identified. Spectral binding titrations and inhibition studies were performed to investigate interactions of rat CYP2B1, rabbit CYP2B4, and CYP2B6 with a series of phenoxyaniline (POA) congeners that are analogues of PBDEs. For most congeners, there was a <3-fold difference between the spectral binding constants (KS) and IC50 values. In contrast, large discrepancies between these values were observed for POA and 3-chloro-4-phenoxyaniline. CYP2B1 was the enzyme most sensitive to POA congeners, so the Val-363 residue from that enzyme was introduced into CYP2B4 or CYP2B6. This substitution partially altered the protein-ligand interaction profiles to make them more similar to that of CYP2B1. Addition of cytochrome P450 oxidoreductase (POR) to titrations of CYP2B6 with POA or 2'4'5'TCPOA decreased the affinity of both ligands for the enzyme. Addition of cytochrome b5 to a recombinant enzyme system containing POR and CYP2B6 increased the POA IC50 value and decreased the 2'4'5'TCPOA IC50 value. Overall, the inconsistency between KS and IC50 values for POA versus 2'4'5'TCPOA is largely due to the effects of redox partner binding. These results provide insight into the biochemical basis of binding of diphenyl ethers to human CYP2B6 and changes in CYP2B6-mediated metabolism that are dependent on POA congener and redox partner identity.

Chinese herbal formula Fuzheng Huayu alleviates CCl4-induced liver fibrosis in rats: a transcriptomic and proteomic analysis.

Liver fibrosis is a consequence of chronic liver disease that can progress to liver cirrhosis or even hepatocarcinoma. Fuzheng Huayu (FZHY), a Chinese herbal formula, has been shown to exert anti-fibrotic effects. To better understand the molecular mechanisms underlying the anti-fibrotic effects of FZHY, we analyzed transcriptomic and proteomic combination profiles in CCl4-induced liver fibrosis in rats, which were treated with extracted FZHY powder (0.35 g·kg-1·d-1, ig) for 3 weeks. We showed that FZHY administration significantly improved liver function, alleviated hepatic inflammatory and fibrotic changes, and decreased the hydroxyproline content in the livers of CCl4-treated rats. When their liver tissues were examined using microarray and iTRAQ, we found 255 differentially expressed genes (fold change ≥1.5, P<0.05) and 499 differentially expressed proteins (fold change ≥1.2, P<0.05) in the FZHY and model groups. Functional annotation with DAVID (The Database for Annotation, Visualization and Integrated Discovery) showed that 15 enriched gene ontology terms, including drug metabolic process, response to extracellular stimulus, response to vitamins, arachidonic acid metabolic process, response to wounding, and oxidation reduction might be involved in the anti-fibrotic effects of FZHY; whereas KEGG pathway analysis revealed that eight enriched pathways, including arachidonic acid metabolism, retinol metabolism, metabolism of xenobiotics by cytochrome P450, and drug metabolism might also be involved. Moreover, the protein-protein interaction network demonstrated that 10 core genes/proteins overlapped, with Ugt2a3, Cyp2b1 and Cyp3a18 in retinol metabolism pathway overlapped to a higher degree. Compared to the model rats, the livers of FZHY-treated rats had significantly higher mRNA and protein expression levels of Ugt2a3, Cyp2b1 and Cyp3a18. Furthermore, the concentration of retinoic acid was significantly higher in the FZHY-treated rats compared with the model rats. The results suggest that the anti-fibrotic effects of FZHY emerge through multiple targets, multiple functions, and multiple pathways, including FZHY-regulated retinol metabolism, xenobiotic metabolism by cytochrome P450, and drug metabolism through up-regulated Ugt2a3, Cyp2b1, and Cyp3a18. These genes may play important anti-fibrotic roles in FZHY-treated rats.

Evaluation of the human relevance of the constitutive androstane receptor-mediated mode of action for rat hepatocellular tumor formation by the synthetic pyrethroid momfluorothrin.

High dietary levels of the non-genotoxic synthetic pyrethroid momfluorothrin increased the incidence of hepatocellular tumors in male and female Wistar rats. Mechanistic studies have demonstrated that the mode of action (MOA) for momfluorothrin-induced hepatocellular tumors is constitutive androstane receptor (CAR)-mediated. In the present study, to evaluate the potential human carcinogenic risk of momfluorothrin, the effects of momfluorothrin (1-1,000 µM) and a major metabolite Z-CMCA (5-1,000 µM) on hepatocyte replicative DNA synthesis and CYP2B mRNA expression were examined in cultured rat and human hepatocyte preparations. The effect of sodium phenobarbital (NaPB), a prototypic rodent hepatocarcinogen with a CAR-mediated MOA, was also investigated. Human hepatocyte growth factor (hHGF) produced a concentration-dependent increase in replicative DNA synthesis in rat and human hepatocytes. However, while NaPB and momfluorothrin increased replicative DNA synthesis in rat hepatocytes, NaPB, momfluorothrin and Z-CMCA did not increase replicative DNA synthesis in human hepatocytes. NaPB, momfluorothrin and Z-CMCA increased CYP2B1/2 mRNA levels in rat hepatocytes. NaPB and momfluorothrin also increased CYP2B6 mRNA levels in human hepatocytes. Overall, while momfluorothrin and NaPB activated CAR in cultured human hepatocytes, neither chemical increased replicative DNA synthesis. Furthermore, to confirm whether the findings observed in vitro were also observed in vivo, a humanized chimeric mouse study was conducted. Replicative DNA synthesis was not increased in human hepatocytes of chimeric mice treated with momfluorothrin or its close structural analogue metofluthrin. As human hepatocytes are refractory to the mitogenic effects of momfluorothrin, in contrast to rat hepatocytes, the data support the hypothesis that the MOA for momfluorothrin-induced rat liver tumor formation is not relevant for humans.

Bromuconazole-induced hepatotoxicity is accompanied by upregulation of PXR/CYP3A1 and downregulation of CAR/CYP2B1 gene expression.

Despite widespread use of bromuconazole as a pesticide for food crops and fruits, limited studies have been done to evaluate its toxic effects. Here, we evaluated the hepatotoxic effect of bromuconazole using classical toxicological (biochemical analysis and histopathological examination) and gene-based molecular methods. Male rats were treated either orally or topically with bromuconazole at doses equal to no observed adverse effect level (NOAEL) and 1/10 LD50 for 90 d. Bromuconazole increased activities of liver enzymes (ALT, AST, ALP, and ACP), and levels of bilirubin. It also induced hepatic oxidative stress as evidenced by significant decrease in the activities of superoxide dismutase (SOD), and significant increase in levels of malondialdehyde (MDA) in liver. In addition, bromuconazole caused an increase in liver weights and necrobiotic changes (vacuolation and hepatocellular hypertrophy). It also strongly induced the expression of PXR and its downstream target CYP3A1 gene as well as the activity of CYP3A1. However, it inhibited the expression of CAR and its downstream target CYP2B1 gene without significant changing in CYP2B1 activity. Overall, the oral route showed higher hepatotoxic effect and molecular changes than the dermal route and all changes were dose dependent. This is the first investigation to report that bromuconazole-induced liver oxidative damage is accompanied by upregulation of PXR/CYP3A1 and downregulation of CAR/CYP2B1.

Unmasking efavirenz neurotoxicity: Time matters to the underlying mechanisms.

Efavirenz is an anti-HIV drug that presents relevant short- and long-term central nervous system adverse reactions. Its main metabolite (8-hydroxy-efavirenz) was demonstrated to be a more potent neurotoxin than efavirenz itself. This work was aimed to understand how efavirenz biotransformation to 8-hydroxy-efavirenz is related to its short- and long-term neuro-adverse reactions. To access those mechanisms, the expression and activity of Cyp2b enzymes as well as the thiolomic signature (low molecular weight thiols plus S-thiolated proteins) were longitudinally evaluated in the hepatic and brain tissues of rats exposed to efavirenz during 10 and 36days. Efavirenz and 8-hydroxy-efavirenz plasma concentrations were monitored at the same time points. Cyp2b induction had a delayed onset in liver (p<0.001), translating into increases in Cyp2b activity in liver and 8-hydroxy-efavirenz plasma concentration (p<0.001). Moreover, an increase in S-cysteinyl-glycinylated proteins (p<0.001) and in free low molecular weight thiols was also observed in liver. A distinct scenario was observed in hippocampus, which showed an underexpression of Cyp2b as well as a decrease in S-cysteinylated and S-glutathionylated proteins. Additionally, the observed changes in tissues were associated with a marked increase of S-glutathionylation in plasma. Our data suggest that the time course of efavirenz biotransformation results from different mechanisms for its short- and long-term neurotoxicity. The difference in the redox profile between liver and hippocampus might explain why, despite being mostly metabolized by the liver, this drug is neurotoxic. If translated to clinical practice, this evidence will have important implications in efavirenz short- and long-term neurotoxicity prevention and management.

Peroxisome Proliferator-Activated Receptor α Activation Suppresses Cytochrome P450 Induction Potential in Mice Treated with Gemfibrozil.

Gemfibrozil, a peroxisome proliferator-activated receptor α (PPARα) agonist, is widely used for hypertriglyceridaemia and mixed hyperlipidaemia. Drug-drug interaction of gemfibrozil and other PPARα agonists has been reported. However, the role of PPARα in cytochrome P450 (CYP) induction by fibrates is not well known. In this study, wild-type mice were first fed gemfibrozil-containing diets (0.375%, 0.75% and 1.5%) for 14 days to establish a dose-response relationship for CYP induction. Then, wild-type mice and Pparα-null mice were treated with a 0.75% gemfibrozil-containing diet for 7 days. CYP3a, CYP2b and CYP2c were induced in a dose-dependent manner by gemfibrozil. In Pparα-null mice, their mRNA level, protein level and activity were induced more than those in wild-type mice. So, gemfibrozil induced CYP, and this action was inhibited by activated PPARα. These data suggested that the induction potential of CYPs was suppressed by activated PPARα, showing a potential role of this receptor in drug-drug interactions and metabolic diseases treated with fibrates.

Impacts of Blast-Induced Traumatic Brain Injury on Expressions of Hepatic Cytochrome P450 1A2, 2B1, 2D1, and 3A2 in Rats.

The hepatic cytochrome P450 (CYP450) enzyme superfamily is one of the most important drug-metabolizing enzyme systems, which is responsible for the metabolism of a large number of clinically relevant medications used in traumatic brain injury (TBI) therapy. Modification of CYP450 expression may have important influences on drug metabolism and lead to untoward effects on those with narrow therapeutic windows. However, the impact of blast-induced TBI (bTBI) on the expression of CYP450 has received little attention. The subfamilies of CYP1A, 2B, 2D, and 3A account for about 85 % of all human drug metabolism of clinical significance. Therefore, in this study, we investigated the expressions of hepatic CYP1A2, CYP2B1, CYP2D1, and CYP3A2 in rats suffering bTBI. Meanwhile, we also measured some important cytokines in serum after injury, and calculated the correlation between these cytokines and the expressions of CYP1A2, CYP2B1, CYP2D1, and CYP3A2. The results showed that bTBI could significantly reduce mRNA expressions of CYP1A2, CYP2D1, and CYP3A2 at the early stage and induce the expressions from 48 h to 1 week after injury. The protein expressions of these CYP450s had all been downregulated from 24 to 48 h post- injury, and then began to elevate at 48 h after bTBI. The cytokines, IL-1ß, IL-2, IL-6, and TNF-α, increased significantly in the early phase, and began to reduce at the delayed phase of bTBI. The serum levels of IL-1ß, IL-6, and TNF-α but not IL-2 were significantly negative correlated with the mRNA expressions of CYP2B1 and CYP2D1 and the proteins expressions of CYP1A2, CYP2B1, CYP2D1, and CYP3A2. In conclusion, our work has, for the first time, indicated that bTBI has significant impact on the expressions of CYP1A2, CYP2B1, CYP2D1, and CYP3A2, which may be related to the cytokines induced by the injury.

Aryl hydrocarbon receptor agonists trigger avoidance of novel food in rats.

The aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor that mediates the toxicity of dioxins, but also plays important physiological roles, which are only beginning to unfold. Previous studies have surprisingly unveiled that low doses of the potent AHR agonist TCDD induce a strong and persistent avoidance of novel food items in rats. Here, we further examined the involvement of the AHR in the avoidance response in Sprague-Dawley rats with three established AHR agonists: 6-formylindolo(3,2-b)carbazole (FICZ), ß-naphthoflavone (BNF) and benzo[a]pyrene (BaP); with a novel selective AHR modulator (C2); and with an activator of another nuclear receptor, CAR: 2,4,6-tryphenyldioxane-1,3 (TPD). As sensitive indices of AHR or CAR activity, we used Cyp1a1 and Cyp2b1 gene expression, as they are, respectively, the drug-metabolizing enzymes specifically regulated by them. We further attempted to address the roles played by enhanced neophobia and conditioned taste aversion (CTA) in the avoidance behaviour. All AHR agonists triggered practically total avoidance of novel chocolate, but the durations varied. Likewise, acutely subtoxic doses of C2, differing by 25-fold, all elicited a similar outcome. In contrast, TPD did not influence chocolate consumption at all. If rats were initially accustomed to chocolate for 6h after single FICZ or BNF exposure, avoidance was still clearly present two weeks later when chocolate was offered again. Hence, the avoidance response appears to specifically involve the AHR instead of being triggered by induction of intestinal or hepatic nuclear receptor signalling in general. It is also shared by both endogenous and exogenous AHR activators. Moreover, this behavioural change in rats seems to contain elements of both CTA and enhanced neophobia, but further clarification of this is still required.

Higher gene expression of CYP1A2, 2B1 and 2D2 in the brain of female compared with male rats.

Cytochrome P450 (CYP) in the brain plays an essential role in the local metabolism of various compounds, including clinically used drugs, toxins, and endogenous substances. In the present study, we compared the expression profiles of mRNAs for several CYP subtypes in the brain between male and female rats. The expression of CYP1A2, CYP2B1, and CYP2D2 in females was significantly higher than that in males. On the other hand, the expression level of the other CYP subtypes examined in the male brain was similar to that in the female brain. These results strongly suggest that marked gender differences exist in the expression profiles of some CYP subtypes in rat brain.

Effects of Nonalcoholic Fatty Liver Disease on Hepatic CYP2B1 and in Vivo Bupropion Disposition in Rats Fed a High-Fat or Methionine/Choline-Deficient Diet.

Nonalcoholic fatty liver disease (NAFLD) refers to hepatic pathologies, including simple fatty liver (SFL), nonalcoholic steatohepatitis (NASH), fibrosis, and cirrhosis, that may progress to hepatocellular carcinoma. These liver disease states may affect the activity and expression levels of drug-metabolizing enzymes, potentially resulting in an alteration in the pharmacokinetics, therapeutic efficacy, and safety of drugs. This study investigated the hepatic cytochrome P450 (CYP) 2B1-modulating effect of a specific NAFLD state in dietary rat models. Sprague-Dawley rats were given a methionine/choline-deficient (MCD) or high-fat (HF) diet to induce NASH and SFL, respectively. The induction of these disease states was confirmed by plasma chemistry and liver histological analysis. Both the protein and mRNA levels of hepatic CYP2B1 were considerably reduced in MCD diet-fed rats; however, they were similar between the HF diet-fed and control rats. Consistently, the enzyme-kinetic and pharmacokinetic parameters for CYP2B1-mediated bupropion metabolism were considerably reduced in MCD diet-fed rats; however, they were also similar between the HF diet-fed and control rats. These results may promote a better understanding of the influence of NAFLD on CYP2B1-mediated metabolism, which could have important implications for the safety and pharmacokinetics of drug substrates for the CYP2B subfamily in patients with NAFLD.

[Effect of xenobiotics on microRNA expression in rat liver].

Using bioinformatics analysis we selected microRNAs which could bind 3'-UTR-region of cytochrome P450 (CYP) genes. Three microRNA miR-21, -221, -222, their potential targets might be mRNA for CYP1A1, and two microRNA miR-143, miR-152 for CYP2B1 accordingly were selected for experimental verification. Expression level of these microRNAs in rat liver upon benzo(a)pyrene (BP), phenobarbital (PB), and DDT induction was determined using RT-qPCR method. In rats treated by both BP, and DDT the hepatic content of miR-21, -221, -222 significantly demonstrated a 2-3-fold decrease. The decrease in miR expression was accompanied by a considerable (5.5-8.7-fold) increase in the CYP1A1-mediated EROD activity. The expression of miR-143 remained unchanged after the PB treatment, while the expression of miR-152 increased by 2 times, however, the (10.5-fold) increase in PROD activity of CYP2B was much higher. In the DDT-treated liver PROD activity increased by 20 times, the expression of miR-152 didn't change, and the expression of miR-143 increased by 2 times. The bioinformatics analysis of interactions between microRNAs and targets showed that the studied miRs can potentially bind 3'-end of AhR, ESR1, GR, CCND1, PTEN mRNA. Thus, the expression profile of miR-21, -221, -222, -143, -152 might change under the xenobiotics exposure. In silico analysis confirmed, that microRNAs target not only cytochrome P450 mRNA but also other genes, including those involved in hormonal carcinogenesis, they also can be regulated with studied miRs.

Subacute nicotine co-exposure has no effect on 2,2',3,5',6- pentachlorobiphenyl disposition but alters hepatic cytochrome P450 expression in the male rat.

Polychlorinated biphenyls (PCBs) are metabolized by cytochrome P450 2B enzymes (CYP2B) and nicotine is reported to alter CYP2B activity in the brain and liver. To test the hypothesis that nicotine influences PCB disposition, 2,2',3,5',6-pentachlorobiphenyl (PCB 95) and its metabolites were quantified in tissues of adult male Wistar rats exposed to PCB 95 (6mg/kg/d, p.o.) in the absence or presence of nicotine (1.0mg/kg/d of the tartrate salt, s.c.) for 7 consecutive days. PCB 95 was enantioselectively metabolized to hydroxylated (OH-) PCB metabolites, resulting in a pronounced enrichment of E1-PCB 95 in all tissues investigated. OH-PCBs were detected in blood and liver tissue, but were below the detection limit in adipose, brain and muscle tissues. Co-exposure to nicotine did not change PCB 95 disposition. CYP2B1 mRNA and CYP2B protein were not detected in brain tissues but were detected in liver. Co-exposure to nicotine and PCB 95 increased hepatic CYP2B1 mRNA but did not change CYP2B protein levels relative to vehicle control animals. However, hepatic CYP2B protein in animals co-exposed to PCB 95 and nicotine were reduced compared to animals that received only nicotine. Quantification of CYP2B3, CYP3A2 and CYP1A2 mRNA identified significant effects of nicotine and PCB 95 co-exposure on hepatic CYP3A2 and hippocampal CYP1A2 transcripts. Our findings suggest that nicotine co-exposure does not significantly influence PCB 95 disposition in the rat. However, these studies suggest a novel influence of PCB 95 and nicotine co-exposure on hepatic cytochrome P450 (P450) expression that may warrant further attention due to the increasing use of e-cigarettes and related products.

DEHP reduces thyroid hormones via interacting with hormone synthesis-related proteins, deiodinases, transthyretin, receptors, and hepatic enzymes in rats.

Di-(2-ethylhexyl) phthalate (DEHP) is used extensively in many personal care and consumer products, resulting in widespread nonoccupational human exposure through multiple routes and media. Limited studies suggest that exposure to DEHP may be associated with altered thyroid function, but detailed mechanisms are unclear. In order to elucidate potential mechanisms by which DEHP disturbs thyroid hormone homeostasis, Sprague-Dawley (SD) rats were dosed with DEHP by gavage at 0, 250, 500, and 750 mg/kg/day for 30 days and sacrificed within 24 h after the last dose. Gene expressions of thyroid hormone receptors, deiodinases, transthyretin, and hepatic enzymes were measured by RT-PCR; protein levels of transthyretin were also analyzed by Western blot. Results showed that DEHP caused histological changes in the thyroid and follicular epithelial cell hypertrophy and hyperplasia were observed. DEHP significantly reduced thyroid hormones (T3, T4) and thyrotropin releasing hormone (TRH) levels, whereas thyroid stimulating hormone (TSH) was not affected. After exposure to DEHP, biosynthesis of thyroid hormones was suppressed, and sodium iodide symporter (NIS) and thyroid peroxidase (TPO) levels were significantly reduced. Additionally, levels of deiodinases and transthyretin were also affected. TSH receptor (TSHr) level was downregulated, while TRH receptor (TRHr) level was upregulated. Metabolism of thyroid hormones was accelerated due to elevated gene expression of hepatic enzymes (UDPGTs and CYP2B1) by DEHP. Taken together, observed findings indicate that DEHP could reduce thyroid hormones through influencing biosynthesis, biotransformation, biotransport, receptor levels, and metabolism of thyroid hormones.

Phase I/II clinical trial of encapsulated, cytochrome P450 expressing cells as local activators of cyclophosphamide to treat spontaneous canine tumours.

Based upon promising preclinical studies, a clinical trial was performed in which encapsulated cells overexpressing cytochrome P450 enzyme isoform 2B1 were implanted around malignant mammary tumours arising spontaneously in dogs. The dogs were then given cyclophosphamide, one of the standard chemotherapeutic agents used for the treatment of mammary tumours. The dogs were assessed for a number of clinical parameters as well as for reduction in tumour size. The treatment was well tolerated with no evidence of adverse reactions or side effects being associated with the administration of the encapsulated cells. Reductions in tumour size of more than 50% were observed for 6 out of the 11 tumours analysed while 5 tumours showing minor responses, i.e. stable disease. In contrast, the tumours that received cyclophosphamide alone showed only stable disease. Taken together, this data suggests that encapsulated cytochrome P450 expressing cells combined with chemotherapy may be useful in the local treatment of a number of dog mammary tumours and support the performance of further clinical studies to evaluate this new treatment.

PCB153 and p,p'-DDE disorder thyroid hormones via thyroglobulin, deiodinase 2, transthyretin, hepatic enzymes and receptors.

Polychlorinated biphenyls (PCBs) and DDT are widespread environmental persistent organic pollutants that have various adverse effects on reproduction, development and endocrine function. In order to elucidate effects of PCBs and DDT on thyroid hormone homeostasis, Sprague-Dawley rats were dosed with PCB153 and p,p'-DDE intraperitoneally (ip) for five consecutive days and sacrificed within 24 h after the last dose. Results indicated that after combined exposure to PCB153 and p,p'-DDE, total thyroxine , free thyroxine, total triiodothyronine, and thyroid-stimulating hormone in serum were decreased, whereas free triiodothyronine and thyrotropin-releasing hormone were not affected. Thyroglobulin and transthyretin levels in serum were significantly reduced. mRNA expression of deiodinases 2 (D2) was also suppressed, while D1 and D3 levels were not significantly influenced after combined exposure. PCB153 and p,p'-DDE induced hepatic enzymes, UDPGTs, CYP1A1, CYP2B1, and CYP3A1 mRNA expressions being significantly elevated. Moreover, TRα1, TRß1, and TRHr expressions in the hypothalamus displayed increasing trends after combined exposure to PCB153 and p,p'-DDE. Taken together, observed results indicate that PCB153 and p,p'-DDE could disorder thyroid hormone homeostasis via thyroglobulin, deiodinase 2, transthyretin, hepatic enzymes, and hormone receptors.

Extravascular use of drug-eluting beads: a promising approach in compartment-based tumor therapy.

Intraperitoneal carcinomatosis (PC) may occur with several tumor entities. The prognosis of patients suffering from PC is usually poor. Present treatment depends on the cancer entity and includes systemic chemotherapy, radiation therapy, hormonal therapy and surgical resection. Only few patients may also benefit from hyperthermic intraperitoneal chemotherapy with a complete tumor remission. These therapies are often accompanied by severe systemic side-effects. One approach to reduce side effects is to target chemotherapeutic agents to the tumor with carrier devices. Promising experimental results have been achieved using drug-eluting beads (DEBs). A series of in vitro and in vitro experiments has been conducted to determine the suitability of their extravascular use. These encapsulation devices were able to harbor CYP2B1 producing cells and to shield them from the hosts immune system when injected intratumorally. In this way ifosfamide--which is transformed into its active metabolites by CYP2B1--could be successfully targeted into pancreatic tumor growths. Furthermore DEBs can be used to target chemotherapeutics into the abdominal cavity for treatment of PC. If CYP2B1 producing cells are proven to be save for usage in man and if local toxic effects of chemotherapeutics can be controlled, DEBs will become promising tools in compartment-based anticancer treatment.

[Effect of shenfu injection on CYP450s of rat liver].

The paper is to report the study of the effect of Shenfu injection on the enzyme activity of liver CYP450 and its mRNA level of rat liver. Microsome of rat liver was prepared after intravenous administration of Shenfu injection for 7 days. The enzyme activity was quantified by Cocktail method. Meanwhile, the mRNA expression of CYP1A2, CYP2B1/2, CYP2C11 and CYP3A1 in the liver was detected by RT-PCR. Shenfu injection obviously induced the enzyme activities of CYP2B and CYP2C. Meantime Shenfu injection decreased the enzyme activities of CYP1A2 and CYP3A. The mRNA levels of CYP2B and CYP2C were also induced in rats treated with Shenfu injection. But it obviously inhibited the mRNA level of CYP1A2 and CYP3A. Since the enzyme activity and mRNA level were obviously changed after administration, the potential effect of drug-drug interaction should be concerned.

CREBH determines the severity of sulpyrine-induced fatal shock.

Although the pyrazolone derivative sulpyrine is widely used as an antipyretic analgesic drug, side effects, including fatal shock, have been reported. However, the molecular mechanism underlying such a severe side effect is largely unclear. Here, we report that the transcription factor CREBH that is highly expressed in the liver plays an important role in fatal shock induced by sulpyrine in mice. CREBH-deficient mice were resistant to experimental fatal sulpyrine shock. We found that sulpyrine-induced expression of cytochrome P450 2B (CYP2B) family genes, which are involved in sulpyrine metabolism, in the liver was severely impaired in CREBH-deficient mice. Moreover, introduction of CYP2B in CREBH-deficient liver restored susceptibility to sulpyrine. Furthermore, ectopic expression of CREBH up-regulated CYP2B10 promoter activity, and in vivo knockdown of CREBH in wild-type mice conferred a significant resistance to fatal sulpyrine shock. These data demonstrate that CREBH is a positive regulator of CYP2B in response to sulpyrine administration, which possibly results in fatal shock.