bluuebunny/crossref_metadata_2025_split huggingface dataset:
Dual Inhibitors of Cyclooxygenase‐2 and Soluble Epoxide Hydrolase: Studies of Binding Modes at the Active Sites and Time‐dependency of Inhibition, and Development of Water‐soluble Prodrugs
Authors: Hwang Sung Hee , Gaieb Zied , Morisseau Christophe , Yang Jun , Wagner Karen , Gilson Michael K. , Hammock Bruce D. 
Published: June 2021

In recent years, there has been increasing interest in designing multi‐target agents that improve efficacy and safety compared to their single target counterparts.Cyclooxygenase‐2 (COX‐2) selective inhibitors (coxibs) are effective drugs for treating inflammation, pain, and cancer. However, their mechanism‐based side effects limit their use. Meanwhile, soluble epoxide hydrolase (sEH) is involved in the metabolism of fatty acid epoxides that are obtained by cytochrome P450 (CYP) from arachidonic acid. These CYP metabolites, epoxyeicosatrienoic acids (EETs), are anti‐hypertensive, anti‐inflammatory, analgesic and organ‐protective. Therefore, co‐inhibition of both enzymes with a combination treatment is synergistic and is maximized by combining both actions into a single molecule, such as PTUPB. These dual inhibitors showed superior efficacy in several rodent models of inflammatory pain, metabolic syndrome, and cancer.We found that they inhibit COX‐2 tightly and in a time‐dependent manner (25‐fold more potent) and EETs moderately inhibit COX‐2. Computational studies of their binding modes at the active site of enzymes using molecular docking confirm these results. In addition to these studies, water‐soluble prodrugs of these dual inhibitors were developed for their use in animal studies.Support or Funding InformationThis work was supported in part by NIEHS grant R01ES02710, NIEHS Superfund Basic Research Program grant P42 ES04699, National Institute of Neurological Disorders and Stroke (NINDS) U54 NS079202. NIH 5T32DC008072‐05.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Inhibition of Soluble Epoxide Hydrolase Attenuates Food Allergen‐induced Eosinophil Recruitment and Gastrointestinal Inflammation
Authors: Rao Savita , Bastan Idil , Ge Xiao Na , Dileepan Mythili , Greenberg Yana G. , Guedes Alonso G. , Hwang Sung He , Hammock Bruce D. , Washabau Robert J. , Sriramarao P 
Published: June 2021

The prevalence of food allergies is on the rise, affecting nearly 4–6% of the population in the US and impacting their daily life in various ways. Although present in the normal gastrointestinal (GI) mucosa under homeostatic conditions, eosinophils are recruited to the intestinal mucosa in increased numbers in food allergy and contribute to inflammation. Soluble epoxide hydrolase (sEH) is known to play a pro‐inflammatory role during inflammation by metabolizing anti‐inflammatory epoxyeicosatrienoic acids (EETs) to pro‐inflammatory diols. In the current study, the role of sEH in a murine model of food allergy was investigated and the potential therapeutic effect of a highly selective sEH inhibitor (t‐TUCB) was evaluated. Oral exposure of mice on a soy‐free diet to soy protein isolate (SPI) induced expression of intestinal sEH, increased circulating total and antigen‐specific IgE levels and caused significant weight loss. Additionally, SPI induced GI inflammation characterized by increased recruitment of eosinophils and mast cells, elevated eotaxin‐1 levels, mucus hypersecretion and decreased epithelial junction protein expression. Administration of t‐TUCB to SPI‐challenged mice not only inhibited IgE levels and prevented SPI‐induced weight loss but also significantly decreased eosinophilia, mast cell recruitment and mucus secretion relative to untreated mice while SPI‐induced loss of junction protein expression was prevented to variable levels. Inflammatory mediators such as TNF‐α and eotaxin‐1 were found to induce sEH expression in eosinophils. Treatment of eosinophils with t‐TUCB significantly inhibited eosinophil migration, an effect which was mirrored by treatment with 11,12‐EET, by inhibiting intracellular signaling events such as ERK (1/2) activation and eotaxin‐1‐induced calcium flux. These studies suggest that sEH induced by soy proteins promotes allergic responses including eosinophilic GI inflammation and that inhibition of sEH can attenuate these allergen‐induced responses.Support or Funding InformationSupport was provided in part by National Institute of Environmental Health Sciences and National Institute of Diabetes and Digestive and Kidney Diseases to B.D.H. and by the University of Minnesota College of Veterinary Medicine Research Office, Minnesota Agricultural Experiment Station, and General Ag Research Funds to R.J.W.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Pharmacological inhibition of soluble epoxide hydrolase promotes brown adipogenesis
Authors: Overby Haley , Kearns Jamie , Hildreth Kelsey , Chahed Samah , Kodani Sean , Morisseau Christophe , Hammock Bruce , Wang Shu , Bettaieb Ahmed , Zhao Ling 
Published: June 2021

Brown adipose tissue (BAT) has emerged as a novel target for obesity treatment and prevention. In contrast to white adipocytes, brown adipocytes are responsible for nonshivering thermogenesis, in which ATP synthesis is uncoupled from respiration via uncoupling protein 1 (UCP1) leading to heat production. Soluble epoxide hydrolase (sEH) is a cytosolic enzyme that converts epoxy fatty acids (EpFAs) into less active diols. EpFAs are produced by cytochrome P‐450 epoxygenases from polyunsaturated fatty acids, such as arachidonic acid, eicosapentaenoic acid, or docosahexaenoic acid (DHA). Many EpFAs are autocrine/paracrine lipid signaling molecules that play important roles in physiological processes, including inflammation, vascular dilation, and cell growth and differentiation. Thus, sEH has become a pharmacological target and small and potent sEH inhibitors that stabilize endogenous EpFAs have been shown to be beneficial for many chronic diseases. The objectives of our studies are to study the regulation of sEH expression and the effects of sEH inhibitors on brown adipogenesis. We found that sEH expression was increased during differentiation of both murine and human brown adipocytes. sEH mRNA levels were significantly increased in the interscapular BAT, but not in the epididymal white adipose tissue (WAT), of diet‐induced obese mice. Moreover, sEH pharmacological inhibition using trans‐4‐{4‐[3‐(4‐trifluoromethoxy‐phenyl)‐ureido]‐cyclohexyloxy}‐benzoic acid) (t‐TUCB) and trans‐4‐[4‐(3‐adamantan‐1‐yl‐ureido)‐cyclohexyloxy]‐benzoic acid (t‐AUCB) dose‐dependently promoted brown adipocyte differentiation, possibly through activating PPAR gamma and alpha. In agreement with these findings, our studies further demonstrated higher UCP1 and PRDM16 protein expression in the interscapular BAT under basal conditions in the sEH knockout (KO) mice compared to the wild type (WT) controls. Moreover, the increases in mRNA expression of brown marker genes in response to cold exposure were enhanced in the interscapular BAT and subcutaneous WAT of the sEH KO mice compared to the WT controls. sEH deficiency further enhanced the induction of brown marker mRNA expression by t‐TUCB and 19, 20‐EDP, the most abundant EpFA derived from DHA. Together, our findings suggest that sEH may regulate brown adipogenesis and its inhibition could be novel therapeutic strategies to increase BAT development and thermogenesis.Support or Funding InformationThe work was partially supported by NIH R15AT008733 (to L.Z. and S.W.), NIH K99DK100736 (to A.B.), R00DK100736 (to A.B.), R01ES002710 (to B.D.H), P42ES004699 (to B.D.H), and T32GM099608 (to S.D.K).This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Modulation of Arachidonic Acid Metabolism in the Rat Kidney by Thymoquinone: Implications for Regulation of Blood Pressure
Authors: Elbarbry Fawzy , Ung Aimy , Lack Kelsey 
Published: June 2021

BackgroundWe investigated the effects of thymoquinone (TQ), a Quinone phytochemical in the plant Nigella Sativa, on Arachidonic acid (AA) metabolism in the kidney and its effect on arterial blood pressure, using spontaneously hypertensive rats (SHR) as our animal model system.MethodsRats were treated for 7 weeks with TQ (10, 20, or 40 mg/kg) added to their drinking water; drinking water alone was used as our control. Mean arterial pressure (MAP) was measured at 7‐day intervals throughout the study. At the end of the treatment, rats were euthanized, followed by preparation of kidney microsomes to measure the activity (using LC/UV and fluorescence assay) and expression (using immunoblotting) of enzymes involved in regulation of vasoactive metabolites: CYP4A, the key enzyme in the formation of 20‐hydroxyeicosatetraenoic acid, and the soluble epoxide hydrolase (sEH), which is responsible for the degradation of the vasodilator metabolites such as epoxyeicosatetraenoic acids.ResultsWe found that treatment with TQ leads to significant reductions in both the expression and activity of renal CYP4A isozyme. Only the high dose TQ resulted in a significant reduction in the expression and activity of renal sEH. Consistent with these data, we have found that treatment with TQ resisted the progressive rise in MAP in the developing SHR in a dosedependent manner.ConclusionThis is the first demonstration that TQ modulates the metabolism of AA by both kidney P450 enzymes and sEH in SHR rats. This may represent a novel mechanism by which TQ protects SHR rats against the progressive rise in blood pressure.Support or Funding InformationPacific University Research Incentive GrantMedical Research Foundation of OregonThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Protective effect of APLN against liver X receptor‐mediated hepatic steatosis through HG11/APLNR in human and mouse hepatocytes
Authors: Huang Jin , Kang Saeromi , Park Soo‐Jin , Im Dong‐Soon 
Published: June 2021

BackgroundNon‐alcoholic fatty liver disease is the most commonly occurring chronic liver disease, and hepatic steatosis is associated with obesity and type II diabetes. APLN is an adipokine that acts on a G protein‐coupled receptor named HG11/APLNR, and has been established to play pivotal roles in various physiological conditions. However, the function of APLN in hepatocytes has not been fully investigated.METHODSIn order to assess the functional roles of APLN and APLNR in hepatocytes, we used an in vitro model of liver X receptor (LXR)‐mediated hepatocellular steatosis.RESULTSIn Hep3B human hepatoma cells, T0901317 (a specific LXR activator) induced lipid accumulation and this was inhibited by APLN. T0901317 also induced the expression of SREBP‐1c, a key transcription factor for lipogenesis. APLN not only inhibited SREBP‐1c induction at the mRNA and protein levels but also induced lipolytic PPAR alfa expression. Furthermore, these protective effects of APLN were inhibited by APLN‐F13A (a specific APLNR antagonist). Furthermore, silencing of APLNR by siRNA transfection also inhibited the actions of APLN. Specific inhibitors of cellular signaling components showed inhibition of lipid accumulation by APLN was mediated through Gi/o proteins, AMPK, and SREBP‐1c suppression during the early stage and through AMPK, ERKs, and PPAR alfa induction during the late stage. In addition, the protective effect of APLN was confirmed in mouse primary hepatocytes.CONCLUSIONThe present findings suggest APLN‐APLNR signaling in hepatocytes functions to protect against lipid accumulation in liver through two signaling pathways, that is, via AMPK activation and PPAR alfa induction.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Activation of Pregnane X Receptor Sensitizes Mice to Hemorrhagic Shock Induced Liver Injury
Authors: Xie Yang , Xu Meishu , Deng Meihong , Li Zhigang , Wang Pengcheng , Guo Yan , Ma Xiaochao , Fan Jie , Billiar Timothy , Xie Wen 
Published: June 2021

Hemorrhagic shock (HS) is a life‐threatening condition often associated with traumatic injuries. Hemorrhagic shock causes tissue hypoperfusion, which may lead to injury to multiple organs including the liver. Pregnane X receptor (PXR) is a xenobiotic receptor that regulates the expression of drug‐metabolizing enzymes (DMEs) such as CYP3A. Many of the drugs used in the trauma patients are known to activate PXR and induce CYP3A. However, it is unclear whether and how PXR plays a role in the regulation of DMEs in the setting of HS and if so, whether activation of PXR beneficial or detrimental to the HS‐responsive liver injury. In this study, we showed that genetic or pharmacological activation of PXR sensitized mice to HS‐responsive liver injury, whereas ablation of PXR protected liver from damage following HS. Mechanistically, the sensitizing effect of PXR activation may have been accounted for by the PXR‐responsive induction of Cyp3a and increased oxidative stress in the liver. Pharmacological inhibition of Cyp3a attenuated the sensitizing effect of PXR. Treatment with the antioxidant NACA relieved the oxidative stress and protected mice from HS‐responsive liver injury. Our results suggest that the unavoidable use of PXR‐activating drugs in trauma patients has the potential to exacerbate HS‐responsive liver injury, which can be mitigated by the co‐administration of anti‐oxidative agents.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Evaluation of Carbazeran as an Enzyme‐Selective Substrate of Human Aldehyde Oxidase: Evidence for Regiospecific Oxidation of Carbazeran by CYP1A2, CYP2D6, CYP3A4, and CYP3A5
Authors: Lau Aik Jiang , Xie Jiarong , Saburulla Nur Fazilah D/O , Chen Shiyan , Zhang Harris L. , Wong Siew Ying 
Published: June 2021

Aldehyde oxidase‐1 (AOX‐1) is a cytosolic xenobiotic‐metabolizing enzyme that has gained increasing interest because of its emerging role in drug metabolism and toxicity. It is a catalyst of carbazeran 4‐oxidation, resulting in the formation of 4‐oxo‐carbazeran. However, cytochrome P450 is also a catalyst of oxidative reactions. Therefore, we tested the hypothesis that cytochrome P450 and AOX‐1 catalyze carbazeran 4‐oxidation in the endoplasmic reticulum and cytosol, respectively. Incubations containing human liver microsomes with or without NADPH yielded 4‐oxo‐carbazeran, but to a lesser extent than that in incubations containing human liver cytosol. Cytochrome P450 inhibitors (1‐aminobenzotriazole and ketoconazole) did not decrease the extent of carbazeran 4‐oxidation in liver microsomes, whereas an AOX‐1 inhibitor (hydralazine) decreased it in liver cytosol. Human recombinant CYP1A2, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP3A4, and CYP3A5 did not catalyze carbazeran 4‐oxidation, in contrast to the catalysis by recombinant AOX‐1. The extent of carbazeran 4‐oxidation in incubations containing liver microsomes was attributed to the presence of AOX‐1 in the microsomal preparations, as indicated by the decrease in carbazeran 4‐oxidation by an AOX‐1 inhibitor (hydralazine) and the detection of AOX‐1 protein in liver microsomes (at a level that was up to one‐third of that in liver cytosol). Cytosolic sulfotransferases‐catalyzed dehydroepiandrosterone sulfation increased with increasing amount of microsomes, indicating that the two lots of hepatic microsomes are contaminated with not only AOX‐1, but also other cytosolic enzymes. An isomer of 4‐oxo‐carbazeran, putatively identified as 5‐hydroxy‐carbazeran or 8‐hydroxy‐carbazeran, was detected in incubations containing carbazeran and CYP1A2, CYP2D6, CYP3A4, or CYP3A5. The 5‐hydroxy‐carbazeran or 8‐hydroxy‐carbazeran peak was detected in human liver microsomal incubation and the peak area increased with increasing incubation time. The 5‐hydroxy‐carbazeran or 8‐hydroxy‐carbazeran peak was decreased by a pan‐CYP inhibitor (1‐aminobenzotriazole), but not by an AOX‐1 inhibitor (hydralazine). The addition of 1‐aminobenzotriazole increased the 4‐oxo‐carbazeran formation, indicating a shunting of carbazeran metabolism to AOX‐1 in the presence of the CYP inhibitor. In conclusion, carbazeran is a dual substrate of AOX‐1 and cytochrome P450. However, AOX‐1, but not cytochrome P450, catalyzes carbazeran 4‐oxidation.Support or Funding InformationSingapore Ministry of Education Academic Research Fund Tier 1 [Grant R‐148‐000‐218‐112 to AJL]This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Physical Organization of Heme Oxygenase 1, NADPH‐Cytochrome P450 Reductase, and the Cytochromes P450 in the Endoplasmic Reticulum
Authors: Connick John Patrick , Backes Wayne L. 
Published: June 2021

NADPH‐Cytochrome P450 reductase (POR) is an endoplasmic reticulum(ER)‐bound enzyme responsible for providing electrons to reactions catalyzed by the cytochromes P450 (P450s) as well as heme oxygenase‐1 (HO‐1). Previous work has revealed that some P450s form physical complexes with each other, leading to changes in their interactions with POR and subsequent modification of P450 activities. Because HO‐1 is expressed in the same location and must compete with the P450s for POR binding, we hypothesized that increased HO‐1 expression could inhibit the POR‐binding of individual P450s and that increased P450 expression could inhibit the POR‐binding of HO‐1. Additionally, HO‐1 has the potential to form homomeric complexes or heteromeric complexes with individual P450s as we have seen previously with the P450s. The goal of this study was to better understand the organization of these proteins, how they might interact in the membrane environment, and to determine if these interactions affected protein function. Bioluminescence resonance energy transfer (BRET) was used to examine these potential protein‐protein interactions. BRET involves expressing a pair of proteins, one with a green fluorescent protein tag and one with a Renilla luciferase tag, in HEK293T cells. Because energy transfer between the two protein tags requires them to be in close proximity, we can determine whether the tagged proteins exist as stable complexes in the cells. Co‐transfecting a third, untagged protein allows us to determine whether this protein destabilizes the complex formed by the tagged proteins. To date, we have determined that HO‐1 forms homomeric complexes that can be disrupted by excess POR, and that HO‐1 appears to form complexes with CYP1A1, CYP1A2, and CYP2D6, but not with CYP2A6 or CYP2C9. At least the HO‐1•CYP1A1 complex appears to be stable in the presence of POR. While further work is required to paint a complete picture of this system, the early results suggest the presence of a previously undescribed network of protein‐protein interactions involving HO‐1, POR, and the P450s.Support or Funding InformationNIH NIEHS ES004344NIH NIEHS ES013648This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Identification of the enzyme isoforms responsible for the N‐oxide detoxification pathway in tobacco specific nitrosamines (TSNAs)
Authors: Perez‐Paramo Yadira Xitlalli , Watson Christy J. W. , Chen Gang , Lazarus Philip 
Published: June 2021

Tobacco‐specific nitrosamines (TSNAs), are among the most potent carcinogens found in tobacco products. Among them are N′‐nitrosonornicotine (NNN), N′‐nitrosoanabasine (NAB), N′‐nitrosoanabatine (NAT), 4‐(methylnitrosamino)‐1‐(3‐pyridyl)‐1‐butanone (NNK) and 4‐ (methylnitrosamino)‐1‐(3‐pyridyl)‐1‐butanol (NNAL), a major metabolite of NNK. TSNAs are detoxified by a number of metabolic pathways, including glucuronidation by the UDP‐glucuronosyltransferase (UGT) family of enzymes, as well as N‐oxidation by an as yet unidentified enzyme or family of enzymes. TSNAs share a pyridine ring in their chemical structure that can be further metabolized to a detoxification N‐oxide product. TSNA‐N‐oxides have been identified in the urine of rodents exposed to TSNAs and NNAL‐N‐oxide was found in the urine of smokers, however, the enzymes responsible for this conversion have not yet been determined. To identify enzymes that catalyze TSNA‐N‐oxide formation, we used specific inhibitors for the major hepatic cytochrome P450 (CYP) enzymes (CYPs 1A2, 2A6, 2B6, 2C8, 2C9, 2C19, 2D6, 2E1, and 3A4) in oxidation reactions with pooled human liver microsomes, including phenacetin, coumarin, bupropion, amodiaquine, diclofenac, omeprazole, dextrometrphan, chlorzoxazone and midazolam. Inhibition of NNK‐N‐oxide and NNAL‐N‐oxide formation was observed when using inhibitors for CYPs 1A2, 2A6, 2E1, 2C19 and 3A4. To confirm results from these inhibition studies, NNK‐N‐oxide and NNAL‐N‐oxide formation was examined using microsomal fractions from CYP over‐expressing HEK293 cells. Microsomes from cells over‐expressing CYPs 1A2, 2C19, 2E1 and 3A4 isoforms exhibited TSNA‐N‐oxide formation. The isoforms 2C19 and 2E1 presented an approximate two‐fold higher activity than the other active isoforms. These data suggest that several cytochrome P450 enzymes may play an important role in the N‐oxidation of TSNAs, and that inter‐individual variability in the activity of these enzymes could potentially be important in TSNA‐induced cancer risk.Support or Funding Information

National Institutes of Health, National Institutes of Environmental Health Sciences [Grant R01‐ES025460]
Health Sciences and Services Authority of Spokane, WA [Grant WSU002292]
This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.