Borazjani A

References (23)

Title : CES1 Releases Oxylipins from Oxidized Triacylglycerol (oxTAG) and Regulates Macrophage oxTAG\/TAG Accumulation and PGE(2)\/IL-1beta Production - Phillips_2023_ACS.Chem.Biol__
Author(s) : Phillips ME , Adekanye O , Borazjani A , Crow JA , Ross MK
Ref : ACS Chemical Biology , : , 2023
Abstract : Triacylglycerols (TAGs) are storage forms of fat, primarily found in cytoplasmic lipid droplets in cells. TAGs are broken down to their component free fatty acids by lipolytic enzymes when fuel reserves are required. However, polyunsaturated fatty acid (PUFA)-containing TAGs are susceptible to nonenzymatic oxidation reactions, leading to the formation of oxylipins that are esterified to the glycerol backbone (termed oxTAGs). Human carboxylesterase 1 (CES1) is a member of the serine hydrolase superfamily and defined by its ability to catalyze the hydrolysis of carboxyl ester bonds in both toxicants and lipids. CES1 is a bona fide TAG hydrolase, but it is unclear which specific fatty acids are preferentially released during lipolysis. To better understand the biochemical function of CES1 in immune cells, such as macrophages, its substrate selectivity when it encounters oxidized PUFAs in TAG lipid droplets requires study. We sought to identify those esterified oxidized fatty acids liberated from oxTAGs by CES1 because their release can activate signaling pathways that enforce the development of lipid-driven inflammation. Gaining this knowledge will help fill data gaps that exist between CES1 and the lipid-sensing nuclear receptors, PPARgamma and LXRalpha, which are important drivers of lipid metabolism and inflammation in macrophages. Oxidized forms of triarachidonoylglycerol (oxTAG20:4) or trilinoleoylglycerol (oxTAG18:2), which contain physiologically relevant levels of oxidized PUFAs (<5 mol %), were incubated with recombinant CES1 to release oxylipins and nonoxidized arachidonic acid (AA) or linoleic acid (LA). CES1 hydrolyzed each oxTAG, yielding regioisomers of hydroxyeicosatetraenoic acids (5-, 11-, 12-, and 15-HETE) and hydroxyoctadecadienoic acids (9- and 13-HODE). Furthermore, human THP-1 macrophages with deficient CES1 levels exhibited a differential response to extracellular stimuli (oxTAGs, lipopolysaccharide, and 15-HETE) as compared to those with normal CES1 levels, including enhanced oxTAG/TAG lipid accumulation and altered cytokine and prostaglandin E2 profiles. This study suggests that CES1 can metabolize oxTAG lipids to release oxylipins and PUFAs, and it further specifies the substrate selectivity of CES1 in the metabolism of bioactive lipid mediators. We suggest that the accumulation of oxTAGs/TAGs within lipid droplets that arise due to CES1 deficiency enforces an inflammatory phenotype in macrophages.
ESTHER : Phillips_2023_ACS.Chem.Biol__
PubMedSearch : Phillips_2023_ACS.Chem.Biol__
PubMedID: 37348046

Title : Cnr1(-\/-) has minimal impact on chlorpyrifos-mediated effects in the mouse endocannabinoid system, but it does alter lipopolysaccharide-induced cytokine levels in splenocytes - Szafran_2023_Chem.Biol.Interact__110425
Author(s) : Szafran BN , Nichols J , Nicaise A , Borazjani A , Carr R , Wilson JR , Ross MK , Kaplan BLF
Ref : Chemico-Biological Interactions , :110425 , 2023
Abstract : Chlorpyrifos (CPF) is an organophosphate pesticide that can inhibit endocannabinoid (eCB) metabolizing enzymes in animal models at levels that do not significantly alter acetylcholinesterase (AChE) in the central nervous system. Previous studies indicated that repeated low-level CPF exposure in developing rats increased the levels of eCBs in the brain. Because eCBs play a role in immune homeostasis through their engagement with cannabinoid receptors, we investigated the role of cannabinoid receptor 1 (CB1, encoded by the Cnr1 gene) on the CPF-mediated effects in the spleen and lung of neonatal and adult female mice. We treated neonatal and adult female Cnr1(-/-) mice with 2.5 mg/kg oral CPF or vehicle for 7 days. Tissues were harvested 4 h after the last CPF dose to evaluate eCB metabolic enzyme activity, levels of eCBs, and tissue immunophenotype. There were a small number of genotype-dependent alterations noted in the endpoints following CPF treatment that were specific to age and tissue type, and differences in eCB metabolism caused by CPF treatment did not correlate to changes in eCB levels. To explore the role of CB1 in CPF-mediated effects on immune endpoints, in vitro experiments were performed with WT murine splenocytes exposed to chlorpyrifos oxon (CPO; oxon metabolite of CPF) and challenged with lipopolysaccharide (LPS). While CPO did not alter LPS-induced pro-inflammatory cytokine levels, inactivation of CB1 by the antagonist SR141716A augmented LPS-induced IFN-gamma levels. Additional experiments with WT and Cnr1(-/-) murine splenocytes confirmed a role for CB1 in altering the production of LPS-induced pro-inflammatory cytokine levels. We conclude that CPF-mediated effects on the eCB system are not strongly dependent on CB1, although abrogation of CB1 does alter LPS-induced cytokine levels in splenocytes.
ESTHER : Szafran_2023_Chem.Biol.Interact__110425
PubMedSearch : Szafran_2023_Chem.Biol.Interact__110425
PubMedID: 36858108

Title : Carboxylesterase 1d Inactivation Augments Lung Inflammation in Mice - Szafran_2022_ACS.Pharmacol.Transl.Sci_5_919
Author(s) : Szafran BN , Borazjani A , Scheaffer HL , Crow JA , McBride AM , Adekanye O , Wonnacott CB , Lehner R , Kaplan BLF , Ross MK
Ref : ACS Pharmacol Transl Sci , 5 :919 , 2022
Abstract : Carboxylesterases are members of the serine hydrolase superfamily and metabolize drugs, pesticides, and lipids. Previous research showed that inhibition of carboxylesterase 1 (CES1) in human macrophages altered the immunomodulatory effects of lipid mediators called prostaglandin glyceryl esters, which are produced by cyclooxygenase-catalyzed oxygenation of the endocannabinoid 2-arachidonoylglycerol (2-AG). Ces1d - the mouse ortholog of human CES1 - is the most abundant Ces isoform in murine lung tissues and alveolar macrophages and a major target of organophosphate poisons. Monoacylglycerol lipase (Magl) is also expressed in murine lung and is the main enzyme responsible for 2-AG catabolism. Several metabolic benefits are observed in Ces1d(-/-) mice fed a high-fat diet; thus, we wondered whether pharmacological and genetic inactivation of Ces1d in vivo might also ameliorate the acute inflammatory response to lipopolysaccharide (LPS). C57BL/6 mice were treated with WWL229 (Ces1d inhibitor) or JZL184 (Magl inhibitor), followed 30 min later by either LPS or saline. Wild-type (WT) and Ces1d(-/-) mice were also administered LPS to determine the effect of Ces1d knockout. Mice were sacrificed at 6 and 24 h, and cytokines were assessed in serum, lung, liver, and adipose tissues. Lipid mediators were quantified in lung tissues, while activity-based protein profiling and enzyme assays determined the extent of lung serine hydrolase inactivation by the inhibitors. WWL229 was shown to augment LPS-induced lung inflammation in a female-specific manner, as measured by enhanced neutrophil infiltration and Il1b mRNA. The marked Ces inhibition in female lung by 4 h after drug treatment might explain this sex difference, although the degree of Ces inhibition in female and male lungs was similar at 6 h. In addition, induction of lung Il6 mRNA and prostaglandin E(2) by LPS was more pronounced in Ces1d(-/-) mice than in WT mice. Thus, WWL229 inhibited lung Ces1d activity and augmented the female lung innate immune response, an effect observed in part in Ces1d(-/-) mice and Ces1d/CES1-deficient murine and human macrophages. In contrast, JZL184 attenuated LPS-induced Il1b and Il6 mRNA levels in female lung, suggesting that Ces1d and Magl have opposing effects. Mapping the immunomodulatory molecules/pathways that are regulated by Ces1d in the context of lung inflammation will require further research.
ESTHER : Szafran_2022_ACS.Pharmacol.Transl.Sci_5_919
PubMedSearch : Szafran_2022_ACS.Pharmacol.Transl.Sci_5_919
PubMedID: 36268116

Title : Effects of Chlorpyrifos on Serine Hydrolase Activities, Lipid Mediators, and Immune Responses in Lungs of Neonatal and Adult Mice - Szafran_2021_Chem.Res.Toxicol__
Author(s) : Szafran BN , Borazjani A , Seay CN , Carr RL , Lehner R , Kaplan BLF , Ross MK
Ref : Chemical Research in Toxicology , : , 2021
Abstract : Chlorpyrifos (CPF) is an organophosphate (OP) pesticide that causes acute toxicity by inhibiting acetylcholinesterase (AChE) in the nervous system. However, endocannabinoid (eCB) metabolizing enzymes in brain of neonatal rats are more sensitive than AChE to inhibition by CPF, leading to increased levels of eCBs. Because eCBs are immunomodulatory molecules, we investigated the association between eCB metabolism, lipid mediators, and immune function in adult and neonatal mice exposed to CPF. We focused on lung effects because epidemiologic studies have linked pesticide exposures to respiratory diseases. CPF was hypothesized to disrupt lung eCB metabolism and alter lung immune responses to lipopolysaccharide (LPS), and these effects would be more pronounced in neonatal mice due to an immature immune system. We first assessed the biochemical effects of CPF in adult mice (<=8 weeks old) and neonatal mice after administering CPF (2.5 mg/kg, oral) or vehicle for 7 days. Tissues were harvested 4 h after the last CPF treatment and lung microsomes from both age groups demonstrated CPF-dependent inhibition of carboxylesterases (Ces), a family of xenobiotic and lipid metabolizing enzymes, whereas AChE activity was inhibited in adult lungs only. Activity-based protein profiling (ABPP)-mass spectrometry of lung microsomes identified 31 and 32 individual serine hydrolases in neonatal lung and adult lung, respectively. Of these, Ces1c/Ces1d/Ces1b isoforms were partially inactivated by CPF in neonatal lung, whereas Ces1c/Ces1b and Ces1c/BChE were partially inactivated in adult female and male lungs, respectively, suggesting age- and sex-related differences in their sensitivity to CPF. Monoacylglycerol lipase (MAGL) and fatty acid amide hydrolase (FAAH) activities in lung were unaffected by CPF. When LPS (1.25 mg/kg, i.p.) was administered following the 7-day CPF dosing period, little to no differences in lung immune responses (cytokines and immunophenotyping) were noted between the CPF and vehicle groups. However, a CPF-dependent increase in the amounts of dendritic cells and certain lipid mediators in female lung following LPS challenge was observed. Experiments in neonatal and adult Ces1d(-)(/-) mice yielded similar results as wild type mice (WT) following CPF treatment, except that CPF augmented LPS-induced Tnfa mRNA in adult Ces1d(-)(/-) mouse lungs. This effect was associated with decreased expression of Ces1c mRNA in Ces1d(-)(/-) mice versus WT mice in the setting of LPS exposure. We conclude that CPF exposure inactivates several Ces isoforms in mouse lung and, during an inflammatory response, increases certain lipid mediators in a female-dependent manner. However, it did not cause widespread altered lung immune effects in response to an LPS challenge.
ESTHER : Szafran_2021_Chem.Res.Toxicol__
PubMedSearch : Szafran_2021_Chem.Res.Toxicol__
PubMedID: 33900070

Title : Inactivation of CES1 Blocks Prostaglandin D(2) Glyceryl Ester Catabolism in Monocytes\/Macrophages and Enhances Its Anti-inflammatory Effects, Whereas the Pro-inflammatory Effects of Prostaglandin E(2) Glyceryl Ester Are Attenuated - Scheaffer_2020_ACS.Omega_5_29177
Author(s) : Scheaffer HL , Borazjani A , Szafran BN , Ross MK
Ref : ACS Omega , 5 :29177 , 2020
Abstract : Human monocytic cells in blood have important roles in host defense and express the enzyme carboxylesterase 1 (CES1). This metabolic serine hydrolase plays a critical role in the metabolism of many molecules, including lipid mediators called prostaglandin glyceryl esters (PG-Gs), which are formed during cyclooxygenase-mediated oxygenation of the endocannabinoid 2-arachidonoylglycerol. Some PG-Gs have been shown to exhibit anti-inflammatory effects; however, they are unstable compounds, and their hydrolytic breakdown generates pro-inflammatory prostaglandins. We hypothesized that by blocking the ability of CES1 to hydrolyze PG-Gs in monocytes/macrophages, the beneficial effects of anti-inflammatory prostaglandin D(2)-glyceryl ester (PGD(2)-G) could be augmented. The goals of this study were to determine whether PGD(2)-G is catabolized by CES1, evaluate the degree to which this metabolism is blocked by small-molecule inhibitors, and assess the immunomodulatory effects of PGD(2)-G in macrophages. A human monocytic cell line (THP-1 cells) was pretreated with increasing concentrations of known small-molecule inhibitors that block CES1 activity [chlorpyrifos oxon (CPO), WWL229, or WWL113], followed by incubation with PGD(2)-G (10 M). Organic solvent extracts of the treated cells were analyzed by liquid chromatography with tandem mass spectrometry to assess levels of the hydrolysis product PGD(2). Further, THP-1 monocytes with normal CES1 expression (control cells) and "knocked-down" CES1 expression (CES1KD cells) were employed to confirm CES1's role in PGD(2)-G catabolism. We found that CES1 has a prominent role in PGD(2)-G hydrolysis in this cell line, accounting for about 50% of its hydrolytic metabolism, and that PGD(2)-G could be stabilized by the inclusion of CES1 inhibitors. The inhibitor potency followed the rank order: CPO > WWL113 > WWL229. THP-1 macrophages co-treated with WWL113 and PGD(2)-G prior to stimulation with lipopolysaccharide exhibited a more pronounced attenuation of pro-inflammatory cytokine levels (interleukin-6 and TNFalpha) than by PGD(2)-G treatment alone. In contrast, prostaglandin E(2)-glyceryl ester (PGE(2)-G) had opposite effects compared to those of PGD(2)-G, which appeared to be dependent on the hydrolysis of PGE(2)-G to PGE(2). These results suggest that the anti-inflammatory effects induced by PGD(2)-G can be further augmented by inactivating CES1 activity with specific small-molecule inhibitors, while pro-inflammatory effects of PGE(2)-G are attenuated. Furthermore, PGD(2)-G (and/or its downstream metabolites) was shown to activate the lipid-sensing receptor PPARgamma, resulting in altered "alternative macrophage activation" response to the Th2 cytokine interleukin-4. These findings suggest that inhibition of CES1 and other enzymes that regulate the levels of pro-resolving mediators such as PGD(2)-G in specific cellular niches might be a novel anti-inflammatory approach.
ESTHER : Scheaffer_2020_ACS.Omega_5_29177
PubMedSearch : Scheaffer_2020_ACS.Omega_5_29177
PubMedID: 33225149
Gene_locus related to this paper: human-CES1

Title : Endocannabinoid hydrolases in avian HD11 macrophages identified by chemoproteomics: inactivation by small-molecule inhibitors and pathogen-induced downregulation of their activity - Lee_2018_Mol.Cell.Biochem_444_125
Author(s) : Lee JH , Hou X , Kummari E , Borazjani A , Edelmann MJ , Ross MK
Ref : Molecular & Cellular Biochemistry , 444 :125 , 2018
Abstract : The endocannabinoids (eCBs) are endogenous arachidonoyl-containing lipid mediators with important roles in host defense. Macrophages are first-line defenders of the innate immune system and biosynthesize large amounts of eCBs when activated. The cellular levels of eCBs are controlled by the activities of their biosynthetic enzymes and catabolic enzymes, which include members of the serine hydrolase (SH) superfamily. The physiologic activity of SHs can be assessed in a class-specific way using chemoproteomic activity-based protein profiling (ABPP) methods. Here, we have examined avian (chicken) HD11 macrophages, a widely used cell line in host-pathogen research, using gel-based ABPP and ABPP-multidimensional protein identification technology (MudPIT) to profile the changes in SH activities under baseline, chemical-inhibitor-treated, and pathogen-challenged conditions. We identified alpha/beta-hydrolase domain 6 (ABHD6) and fatty acid amide hydrolase (FAAH) as the principal SHs responsible for 2-arachidonoylglycerol (2AG) hydrolysis, thereby regulating the concentration of this lipid in HD11 cells. We further discovered that infection of HD11 macrophages by Salmonella Typhimurium caused the activities of these 2AG hydrolases to be downregulated in the host cells. ABHD6 and FAAH were potently inhibited by a variety of small-molecule inhibitors in intact live cells, and thus these compounds might be useful host-directed adjuvants to combat antimicrobial resistance in agriculture. 2AG was further shown to augment the phagocytic function of HD11 macrophages, which suggests that pathogen-induced downregulation of enzymes controlling 2AG hydrolytic activity might be a physiological mechanism to increase 2AG levels, thus enhancing phagocytosis. Together these results define ABHD6 and FAAH as 2AG hydrolases in avian macrophages that can be inactivated pharmacologically and decreased in activity during Salmonella Typhimurium infection.
ESTHER : Lee_2018_Mol.Cell.Biochem_444_125
PubMedSearch : Lee_2018_Mol.Cell.Biochem_444_125
PubMedID: 29196970

Title : Characterization of Endocannabinoid-Metabolizing Enzymes in Human Peripheral Blood Mononuclear Cells under Inflammatory Conditions - Szafran_2018_Molecules_23_
Author(s) : Szafran BN , Lee JH , Borazjani A , Morrison P , Zimmerman G , Andrzejewski KL , Ross MK , Kaplan BLF
Ref : Molecules , 23 : , 2018
Abstract : Endocannabinoid-metabolizing enzymes are downregulated in response to lipopolysaccharide (LPS)-induced inflammation in mice, which may serve as a negative feedback mechanism to increase endocannabinoid levels and reduce inflammation. Increased plasma levels of the pro-inflammatory cytokine interleukin-6 (IL-6) and decreased fatty acid amide hydrolase (FAAH) activity in peripheral lymphocytes from individuals diagnosed with Huntington's disease (HD) suggests that a similar negative feedback system between inflammation and the endocannabinoid system operates in humans. We investigated whether CpG- (unmethylated bacterial DNA) and LPS-induced IL-6 levels in peripheral blood mononuclear cells (PBMCs) from non-HD and HD individuals modulated the activities of endocannabinoid hydrolases monoacylglycerol lipase (MAGL) and carboxylesterase (CES). Baseline plasma IL-6 levels and 2-arachidonoylglycerol (2-AG) hydrolytic activity in PBMC lysates were not different in HD and non-HD individuals. Inhibition of MAGL and CES1 activity in PBMCs using the inhibitors JZL184 and WWL113, respectively, demonstrated that MAGL was the dominant 2-AG hydrolytic enzyme in PBMCs, regardless of disease state. Correlative analyses of 2-AG hydrolytic activity versus enzyme abundance confirmed this conclusion. Flow cytometric analysis of PBMCs showed that MAGL and CES1 were primarily expressed in monocytes and to a lesser extent in lymphocytes. In conclusion, these data suggest that IL-6 did not influence 2-AG hydrolytic activity in human PBMCs; however, monocytic MAGL was shown to be the predominant 2-AG hydrolytic enzyme.
ESTHER : Szafran_2018_Molecules_23_
PubMedSearch : Szafran_2018_Molecules_23_
PubMedID: 30513753

Title : Silencing carboxylesterase 1 in human THP-1 macrophages perturbs genes regulated by PPARgamma\/RXR and RAR\/RXR: down-regulation of CYP27A1-LXRalpha signaling - Mangum_2018_Biochem.J_475_621
Author(s) : Mangum LC , Hou X , Borazjani A , Lee JH , Ross MK , Crow JA
Ref : Biochemical Journal , 475 :621 , 2018
Abstract : Macrophage foam cells store excess cholesterol as cholesteryl esters, which need to be hydrolyzed for cholesterol efflux. We recently reported that silencing expression of carboxylesterase 1 (CES1) in human THP-1 macrophages [CES1KD (THP-1 cells with CES1 expression knocked down) macrophages] reduced cholesterol uptake and decreased expression of CD36 and scavenger receptor-A in cells loaded with acetylated low-density lipoprotein (acLDL). Here, we report that CES1KD macrophages exhibit reduced transcription of cytochrome P45027A1 (CYP27A1) in nonloaded and acLDL-loaded cells. Moreover, levels of CYP27A1 protein and its enzymatic product, 27-hydroxycholesterol, were markedly reduced in CES1KD macrophages. Transcription of LXRalpha (liver X receptor alpha) and ABCA1 (ATP-binding cassette transporter A1) was also decreased in acLDL-loaded CES1KD macrophages, suggesting reduced signaling through PPARgamma-CYP27A1-LXRalpha. Consistent with this, treatment of CES1KD macrophages with agonists for PPARgamma, RAR, and/or RAR/RXR partially restored transcription of CYP27A1 and LXRalpha, and repaired cholesterol influx. Conversely, treatment of control macrophages with antagonists for PPARgamma and/or RXR decreased transcription of CYP27A1 and LXRalpha Pharmacologic inhibition of CES1 in both wild-type THP-1 cells and primary human macrophages also decreased CYP27A1 transcription. CES1 silencing did not affect transcript levels of PPARgamma and RXR in acLDL-loaded macrophages, whereas it did reduce the catabolism of the endocannabinoid 2-arachidonoylglycerol. Finally, the gene expression profile of CES1KD macrophages was similar to that of PPARgamma knockdown cells following acLDL exposures, further suggesting a mechanistic link between CES1 and PPARgamma. These results are consistent with a model in which abrogation of CES1 function attenuates the CYP27A1-LXRalpha-ABCA1 signaling axis by depleting endogenous ligands for the nuclear receptors PPARgamma, RAR, and/or RXR that regulate cholesterol homeostasis.
ESTHER : Mangum_2018_Biochem.J_475_621
PubMedSearch : Mangum_2018_Biochem.J_475_621
PubMedID: 29321244
Gene_locus related to this paper: human-CES1

Title : Interaction of the serine hydrolase KIAA1363 with organophosphorus agents: Evaluation of potency and kinetics - Ross_2016_Arch.Biochem.Biophys_590_72
Author(s) : Ross MK , Pluta K , Bittles V , Borazjani A , Allen Crow J
Ref : Archives of Biochemistry & Biophysics , 590 :72 , 2016
Abstract : Oxons are bioactive metabolites of organophosphorus insecticides (OPs) that covalently inactivate serine hydrolases. KIAA1363 is one of the most abundant serine hydrolases in mouse brain. Although the physiological consequences related to the inhibition of KIAA1363 due to environmental exposures to OPs are poorly understood, the enzyme was previously shown to have a role in the detoxification of oxons. Here, we overexpressed human KIAA1363 and CES1 in COS7 cells and compared the potency of inhibition (IC50s, 15 min) of KIAA1363 and CES1 by chlorpyrifos oxon (CPO), paraoxon (PO), and methyl paraoxon (MPO). The order of potency was CPO > PO >> MPO for both enzymes. We also determined the bimolecular rate constants (kinact/Ki) for reactions of CPO and PO with KIAA1363 and CES1. KIAA1363 and CES1 were inactivated by CPO at comparable rates (4.4 x 10(6) s(-1) M(-1) and 6.7 x 10(6) s(-1) M(-1), respectively), whereas PO inactivated both enzymes at slower rates (0.4 x 10(6) s(-1) M(-1) and 1.5 x 10(6) s(-1) M(-1), respectively). Finally, the reactivation rate of KIAA1363 following inhibition by CPO was evaluated. Together, the results define the kinetics of inhibition of KIAA1363 by active metabolites of agrochemicals and indicate that KIAA1363 is highly sensitive to inhibition by these compounds.
ESTHER : Ross_2016_Arch.Biochem.Biophys_590_72
PubMedSearch : Ross_2016_Arch.Biochem.Biophys_590_72
PubMedID: 26617293
Gene_locus related to this paper: human-NCEH1

Title : Lipopolysaccharide suppresses carboxylesterase 2g activity and 2-arachidonoylglycerol hydrolysis: A possible mechanism to regulate inflammation - Szafran_2015_Prostaglandins.Other.Lipid.Mediat_121_199
Author(s) : Szafran B , Borazjani A , Lee JH , Ross MK , Kaplan BL
Ref : Prostaglandins Other Lipid Mediat , 121 :199 , 2015
Abstract : Inflammation is an important part of the innate immune response and is involved in the healing of many disease processes; however, chronic inflammation is a harmful component of many diseases. The regulatory mechanisms of inflammation are incompletely understood. One possible regulatory mechanism is the endocannabinoid system. Endocannabinoids such as 2-arachidonoylglycerol (2-AG) and anandamide (AEA) are generally anti-inflammatory via engagement of the cannabinoid receptor 2 (CB2) on innate cells; therefore, preventing the degradation of endocannabinoids by specific serine hydrolases such as fatty acid amide hydrolase (FAAH), monoacylglycerol lipase (MAGL), and carboxylesterases (CES) might decrease inflammation. We hypothesized that the activities of these catabolic enzymes would decrease with a subsequent increase in 2-AG and AEA in a model of inflammation. Mice were injected with lipopolysaccharide (LPS) for 6 or 24h, and inflammation was confirmed by an increase in interleukin-6 (il6) and il17 gene expression. Activity-based protein profiling (ABPP) of serine hydrolases showed no significant difference in various serine hydrolase activities in brain or liver, whereas a modest decrease in Ces activity in spleen after LPS administration was noted. 2-AG hydrolase activity in the spleen was also decreased at 6h post LPS, which was corroborated by LPS treatment of splenocytes ex vivo. ABPP-MudPIT proteomic analysis suggested that the decreased 2-AG hydrolysis in spleen was due to a reduction in Ces2g activity. These studies suggest that the endocannabinoid system could be activated via suppression of a 2-AG catabolic enzyme in response to inflammatory stimuli as one mechanism to limit inflammation.
ESTHER : Szafran_2015_Prostaglandins.Other.Lipid.Mediat_121_199
PubMedSearch : Szafran_2015_Prostaglandins.Other.Lipid.Mediat_121_199
PubMedID: 26403860

Title : Effects of toxicologically relevant xenobiotics and the lipid-derived electrophile 4-hydroxynonenal on macrophage cholesterol efflux: silencing carboxylesterase 1 has paradoxical effects on cholesterol uptake and efflux - Ross_2014_Chem.Res.Toxicol_27_1743
Author(s) : Ross MK , Borazjani A , Mangum LC , Wang R , Crow JA
Ref : Chemical Research in Toxicology , 27 :1743 , 2014
Abstract : Cholesterol cycles between free cholesterol (unesterified) found predominantly in membranes and cholesteryl esters (CEs) stored in cytoplasmic lipid droplets. Only free cholesterol is effluxed from macrophages via ATP-binding cassette (ABC) transporters to extracellular acceptors. Carboxylesterase 1 (CES1), proposed to hydrolyze CEs, is inactivated by oxon metabolites of organophosphorus pesticides and by the lipid electrophile 4-hydroxynonenal (HNE). We assessed the ability of these compounds to reduce cholesterol efflux from foam cells. Human THP-1 macrophages were loaded with [(3)H]-cholesterol/acetylated LDL and then allowed to equilibrate to enable [(3)H]-cholesterol to distribute into its various cellular pools. The cholesterol-engorged cells were then treated with toxicants in the absence of cholesterol acceptors for 24 h, followed by a 24 h efflux period in the presence of toxicant. A concentration-dependent reduction in [(3)H]-cholesterol efflux via ABCA1 (up to 50%) was found for paraoxon (0.1-10 muM), whereas treatment with HNE had no effect. A modest reduction in [(3)H]-cholesterol efflux via ABCG1 (25%) was found after treatment with either paraoxon or chlorpyrifos oxon (10 muM each) but not HNE. No difference in efflux rates was found after treatments with either paraoxon or HNE when the universal cholesterol acceptor 10% (v/v) fetal bovine serum was used. When the re-esterification arm of the CE cycle was disabled in foam cells, paraoxon treatment increased CE levels, suggesting the neutral CE hydrolysis arm of the cycle had been inhibited by the toxicant. However, paraoxon also partially inhibited lysosomal acid lipase, which generates cholesterol for efflux, and reduced the expression of ABCA1 protein. Paradoxically, silencing CES1 expression in macrophages did not affect the percent of [(3)H]-cholesterol efflux. However, CES1 mRNA knockdown markedly reduced cholesterol uptake by macrophages, with SR-A and CD36 mRNA reduced 3- and 4-fold, respectively. Immunoblots confirmed SR-A and CD36 protein downregulation. Together, these results suggest that toxicants, e.g., oxons, may interfere with macrophage cholesterol homeostasis/metabolism.
ESTHER : Ross_2014_Chem.Res.Toxicol_27_1743
PubMedSearch : Ross_2014_Chem.Res.Toxicol_27_1743
PubMedID: 25250848

Title : Identification of palmitoyl protein thioesterase 1 in human THP1 monocytes and macrophages and characterization of unique biochemical activities for this enzyme - Wang_2013_Biochemistry_52_7559
Author(s) : Wang R , Borazjani A , Matthews AT , Mangum LC , Edelmann MJ , Ross MK
Ref : Biochemistry , 52 :7559 , 2013
Abstract : The profiles of serine hydrolases in human and mouse macrophages are similar yet different. For instance, human macrophages express high levels of carboxylesterase 1 (CES1), whereas mouse macrophages have minimal amounts of the orthologous murine CES1. On the other hand, macrophages from both species exhibit limited expression of the canonical 2-arachidonoylglycerol (2-AG) hydrolytic enzyme, MAGL. Our previous study showed CES1 was partly responsible for the hydrolysis of 2-AG (50%) and prostaglandin glyceryl esters (PG-Gs) (80-95%) in human THP1 monocytes and macrophages. However, MAGL and other endocannabinoid hydrolases, FAAH, ABHD6, and ABHD12, did not have a role because of limited expression or no expression. Thus, another enzyme was hypothesized to be responsible for the remaining 2-AG hydrolysis activity following chemical inhibition and immunodepletion of CES1 (previous study) or CES1 gene knockdown (this study). Here we identified two candidate serine hydrolases in THP1 cell lysates by activity-based protein profiling (ABPP)-MUDPIT and Western blotting: cathepsin G and palmitoyl protein thioesterase 1 (PPT1). Both proteins exhibited electrophoretic properties similar to those of a serine hydrolase in THP1 cells detected by gel-based ABPP at 31-32 kDa; however, only PPT1 exhibited lipolytic activity and hydrolyzed 2-AG in vitro. Interestingly, PPT1 was strongly expressed in THP1 cells but was significantly less reactive than cathepsin G toward the activity-based probe, fluorophosphonate-biotin. KIAA1363, another serine hydrolase, was also identified in THP1 cells but did not have significant lipolytic activity. On the basis of chemoproteomic profiling, immunodepletion studies, and chemical inhibitor profiles, we estimated that PPT1 contributed 32-40% of 2-AG hydrolysis activity in the THP1 cell line. In addition, pure recombinant PPT1 catalyzed the hydrolysis of 2-AG, PGE2-G, and PGF2alpha-G, although the catalytic efficiency of hydrolysis of 2-AG by PPT1 was ~10-fold lower than that of CES1. PPT1 was also insensitive to several chemical inhibitors that potently inhibit CES1, such as organophosphate poisons and JZL184. This is the first report to document the expression of PPT1 in a human monocyte and macrophage cell line and to show PPT1 can hydrolyze the natural substrates 2-AG and PG-Gs. These findings suggest that PPT1 may participate in endocannabinoid metabolism within specific cellular contexts and highlights the functional redundancy often exhibited by enzymes involved in lipid metabolism.
ESTHER : Wang_2013_Biochemistry_52_7559
PubMedSearch : Wang_2013_Biochemistry_52_7559
PubMedID: 24083319
Gene_locus related to this paper: human-PPT1

Title : Inhibition of recombinant human carboxylesterase 1 and 2 and monoacylglycerol lipase by chlorpyrifos oxon, paraoxon and methyl paraoxon - Crow_2012_Toxicol.Appl.Pharmacol_258_145
Author(s) : Crow JA , Bittles V , Herring KL , Borazjani A , Potter PM , Ross MK
Ref : Toxicol Appl Pharmacol , 258 :145 , 2012
Abstract : Oxons are the bioactivated metabolites of organophosphorus insecticides formed via cytochrome P450 monooxygenase-catalyzed desulfuration of the parent compound. Oxons react covalently with the active site serine residue of serine hydrolases, thereby inactivating the enzyme. A number of serine hydrolases other than acetylcholinesterase, the canonical target of oxons, have been reported to react with and be inhibited by oxons. These off-target serine hydrolases include carboxylesterase 1 (CES1), CES2, and monoacylglycerol lipase. Carboxylesterases (CES, EC 3.1.1.1) metabolize a number of xenobiotic and endobiotic compounds containing ester, amide, and thioester bonds and are important in the metabolism of many pharmaceuticals. Monoglyceride lipase (MGL, EC 3.1.1.23) hydrolyzes monoglycerides including the endocannabinoid, 2-arachidonoylglycerol (2-AG). The physiological consequences and toxicity related to the inhibition of off-target serine hydrolases by oxons due to chronic, low level environmental exposures are poorly understood. Here, we determined the potency of inhibition (IC(50) values; 15 min preincubation, enzyme and inhibitor) of recombinant CES1, CES2, and MGL by chlorpyrifos oxon, paraoxon and methyl paraoxon. The order of potency for these three oxons with CES1, CES2, and MGL was chlorpyrifos oxon>paraoxon>methyl paraoxon, although the difference in potency for chlorpyrifos oxon with CES1 and CES2 did not reach statistical significance. We also determined the bimolecular rate constants (k(inact)/K(I)) for the covalent reaction of chlorpyrifos oxon, paraoxon and methyl paraoxon with CES1 and CES2. Consistent with the results for the IC(50) values, the order of reactivity for each of the three oxons with CES1 and CES2 was chlorpyrifos oxon>paraoxon>methyl paraoxon. The bimolecular rate constant for the reaction of chlorpyrifos oxon with MGL was also determined and was less than the values determined for chlorpyrifos oxon with CES1 and CES2 respectively. Together, the results define the kinetics of inhibition of three important hydrolytic enzymes by activated metabolites of widely used agrochemicals.
ESTHER : Crow_2012_Toxicol.Appl.Pharmacol_258_145
PubMedSearch : Crow_2012_Toxicol.Appl.Pharmacol_258_145
PubMedID: 22100607

Title : Covalent inhibition of recombinant human carboxylesterase 1 and 2 and monoacylglycerol lipase by the carbamates JZL184 and URB597 - Crow_2012_Biochem.Pharmacol_84_1215
Author(s) : Crow JA , Bittles V , Borazjani A , Potter PM , Ross MK
Ref : Biochemical Pharmacology , 84 :1215 , 2012
Abstract : Carboxylesterase type 1 (CES1) and CES2 are serine hydrolases located in the liver and small intestine. CES1 and CES2 actively participate in the metabolism of several pharmaceuticals. Recently, carbamate compounds were developed to inhibit members of the serine hydrolase family via covalent modification of the active site serine. URB597 and JZL184 inhibit fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MAGL), respectively; however, carboxylesterases in liver have been identified as a major off-target. We report the kinetic rate constants for inhibition of human recombinant CES1 and CES2 by URB597 and JZL184. Bimolecular rate constants (k(inact)/K(i)) for inhibition of CES1 by JZL184 and URB597 were similar [3.9 (+/-0.2) x 10(3) M(-1) s(-1) and 4.5 (+/-1.3) x 10(3) M(-1) s(-1), respectively]. However, k(inact)/K(i) for inhibition of CES2 by JZL184 and URB597 were significantly different [2.3 (+/-1.3) x 10(2) M(-1) s(-1) and 3.9 (+/-1.0) x 10(3) M(-1) s(-1), respectively]. Rates of inhibition of CES1 and CES2 by URB597 were similar; however, CES1 and MAGL were more potently inhibited by JZL184 than CES2. We also determined kinetic constants for spontaneous reactivation of CES1 carbamoylated by either JZL184 or URB597 and CES1 diethylphosphorylated by paraoxon. The reactivation rate was significantly slower (4.5x) for CES1 inhibited by JZL184 than CES1 inhibited by URB597. Half-life of reactivation for CES1 carbamoylated by JZL184 was 49 +/- 15 h, which is faster than carboxylesterase turnover in HepG2 cells. Together, the results define the kinetics of inhibition for a class of drugs that target hydrolytic enzymes involved in drug and lipid metabolism.
ESTHER : Crow_2012_Biochem.Pharmacol_84_1215
PubMedSearch : Crow_2012_Biochem.Pharmacol_84_1215
PubMedID: 22943979

Title : Examination of the carboxylesterase phenotype in human liver - Ross_2012_Arch.Biochem.Biophys_522_44
Author(s) : Ross MK , Borazjani A , Wang R , Crow JA , Xie S
Ref : Archives of Biochemistry & Biophysics , 522 :44 , 2012
Abstract : Carboxylesterases (CES) metabolize esters. Two CES isoforms are expressed in human liver (CES1 and CES2) and liver extracts are used in reaction phenotyping studies to discern interindividual metabolic variation. We tested the hypothesis that an individual's CES phenotype can be characterized by reporter substrates/probes that interrogate native CES1 and CES2 activities in liver and immunoblotting methods. We obtained 25 livers and found that CES1 is the main hydrolytic enzyme. Moreover, although CES1 protein levels were similar, we observed large interindividual variation in bioresmethrin hydrolysis rates (17-fold), a pyrethroid metabolized by CES1 but not CES2. Bioresmethrin hydrolysis rates did not correlate with CES1 protein levels. In contrast, procaine hydrolysis rates, a drug metabolized by CES2 but not CES1, were much less variant (3-fold). Using activity-based fluorophosphonate probes (FP-biotin), which covalently reacts with active serine hydrolases, CES1 protein was the most active enzyme in the livers. Finally, using bioorthogonal probes and click chemistry methodology, the half-life of CES 1 and 2 in cultured HepG2 cells was estimated at 96 h. The cause of the differential CES1 activities is unknown, but the underlying factors will be important to understand because several carboxylic acid ester drugs and environmental toxicants are metabolized by this enzyme.
ESTHER : Ross_2012_Arch.Biochem.Biophys_522_44
PubMedSearch : Ross_2012_Arch.Biochem.Biophys_522_44
PubMedID: 22525521

Title : Catabolism of 4-hydroxy-2-trans-nonenal by THP1 monocytes\/macrophages and inactivation of carboxylesterases by this lipid electrophile - Borazjani_2011_Chem.Biol.Interact_194_1
Author(s) : Borazjani A , Edelmann MJ , Hardin KL , Herring KL , Allen Crow J , Ross MK
Ref : Chemico-Biological Interactions , 194 :1 , 2011
Abstract : Oxidative stress in cells and tissues leads to the formation of an assortment of lipid electrophiles, such as the quantitatively important 4-hydroxy-2-trans-nonenal (HNE). Although this cytotoxic aldehyde is atherogenic the mechanisms involved are unclear. We hypothesize that elevated HNE levels can directly inactivate esterase and lipase activities in macrophages via protein adduction, thus generating a biochemical lesion that accelerates foam cell formation and subsequent atherosclerosis. In the present study we examined the effects of HNE treatment on esterase and lipase activities in human THP1 monocytes/macrophages at various physiological scales (i.e., pure recombinant enzymes, cell lysate, and intact living cells). The hydrolytic activities of bacterial and human carboxylesterase enzymes (pnbCE and CES1, respectively) were inactivated by HNE in vitro in a time- and concentration-dependent manner. In addition, so were the hydrolytic activities of THP1 cell lysates and intact THP1 monocytes and macrophages. A single lysine residue (Lys105) in recombinant CES1 was modified by HNE via a Michael addition reaction, whereas the lone reduced cysteine residue (Cys389) was found unmodified. The lipolytic activity of cell lysates and intact cells was more sensitive to the inhibitory effects of HNE than the esterolytic activity. Moreover, immunoblotting analysis using HNE antibodies confirmed that several cellular proteins were adducted by HNE following treatment of intact THP1 monocytes, albeit at relatively high HNE concentrations (>50muM). Unexpectedly, in contrast to CES1, the treatment of a recombinant human CES2 with HNE enhanced its enzymatic activity approximately 3-fold compared to untreated enzyme. In addition, THP1 monocytes/macrophages can efficiently metabolize HNE, and glutathione conjugation of HNE is responsible for approximately 43% of its catabolism. The functional importance of HNE-mediated inactivation of cellular hydrolytic enzymes with respect to atherogenesis remains obscure, although this study has taken a first step toward addressing this important issue by examining the potential of HNE to inhibit this biochemical activity in a human monocyte/macrophage cell line.
ESTHER : Borazjani_2011_Chem.Biol.Interact_194_1
PubMedSearch : Borazjani_2011_Chem.Biol.Interact_194_1
PubMedID: 21878322

Title : Effect of developmental chlorpyrifos exposure, on endocannabinoid metabolizing enzymes, in the brain of juvenile rats - Carr_2011_Toxicol.Sci_122_112
Author(s) : Carr RL , Borazjani A , Ross MK
Ref : Toxicol Sci , 122 :112 , 2011
Abstract : The endogenous cannabinoids 2-arachidonoylglycerol (2-AG) and arachidonoyl ethanolamide (AEA or anandamide) play vital roles during nervous system development including regulating axonal guidance and synaptogenesis. The enzymatic degradation of 2-AG and AEA is highly susceptible to inhibition by organophosphate compounds in vitro. Furthermore, acute in vivo exposure of adult animals to the agricultural insecticide chlorpyrifos (CPS) caused moderate inhibition of both 2-AG and AEA hydrolysis. However, the effects of repeated exposure to lower levels of CPS, especially during development, on endocannabinoid metabolism in the brain is not known. To examine this, rat pups were orally exposed daily from postnatal days 10-16 to either 1.0, 2.5, or 5.0 mg/kg CPS. Body weight gain was reduced by 5.0 mg/kg on all days of treatment whereas 2.5 mg/kg reduced the weight gain only on the last two days of treatment. At 4-h postexposure on day 16, forebrain cholinesterase (ChE) activity and hydrolysis of 2-AG and AEA were inhibited in a dose-related manner, and the extent of inhibition from highest to lowest level was AEA hydrolysis > ChE activity > 2-AG hydrolysis. The extent of inhibition of AEA hydrolysis was approximately twice than that of ChE activity with AEA hydrolysis being virtually eliminated by 2.5 and 5.0 mg/kg and 1.0 mg/kg causing 40% inhibition. The sensitivity of AEA hydrolysis, compared with canonical targets such as ChE activity, suggests a potential alternative developmental target for CPS. Inhibition of AEA hydrolysis could result in accumulation of endocannabinoids, which could alter normal endocannabinoid transmission during brain maturation.
ESTHER : Carr_2011_Toxicol.Sci_122_112
PubMedSearch : Carr_2011_Toxicol.Sci_122_112
PubMedID: 21507991

Title : Inactivation of lipid glyceryl ester metabolism in human THP1 monocytes\/macrophages by activated organophosphorus insecticides: role of carboxylesterases 1 and 2 - Xie_2010_Chem.Res.Toxicol_23_1890
Author(s) : Xie S , Borazjani A , Hatfield MJ , Edwards CC , Potter PM , Ross MK
Ref : Chemical Research in Toxicology , 23 :1890 , 2010
Abstract : Carboxylesterases (CES) have important roles in pesticide and drug metabolism and contribute to the clearance of ester-containing xenobiotics in mammals. Tissues with the highest levels of CES expression are the liver and small intestine. In addition to xenobiotics, CES also harness their broad substrate specificity to hydrolyze endobiotics, such as cholesteryl esters and triacylglycerols. Here, we determined if two human CES isoforms, CES1 and CES2, hydrolyze the endocannabinoids 2-arachidonoylglycerol (2AG) and anandamide (AEA), and two prostaglandin glyceryl esters (PG-Gs), which are formed by COX-mediated oxygenation of 2AG. We show that recombinant CES1 and CES2 efficiently hydrolyze 2AG to arachidonic acid (AA) but not amide-containing AEA. Steady-state kinetic parameters for CES1- and CES2-mediated 2AG hydrolysis were, respectively, kcat, 59 and 43 min(-1); Km, 49 and 46 muM; and kcat/Km, 1.2 and 0.93 muM(-1) min(-1). kcat/Km values are comparable to published values for rat monoacylglycerol lipase (MAGL)-catalyzed 2AG hydrolysis. Furthermore, we show that CES1 and CES2 also efficiently hydrolyze PGE2-G and PGF2alpha-G. In addition, when cultured human THP1 macrophages were treated with exogenous 2AG or PG-G (10 muM, 1 h), significant quantities of AA or PGs were detected in the culture medium; however, the ability of macrophages to metabolize these compounds was inhibited (60-80%) following treatment with paraoxon, the toxic metabolite of the insecticide parathion. Incubation of THP1 cell lysates with small-molecule inhibitors targeting CES1 (thieno[3,2-e][1]benzothiophene-4,5-dione or JZL184) significantly reduced lipid glyceryl ester hydrolase activities (40-50% for 2AG and 80-95% for PG-Gs). Immunodepletion of CES1 also markedly reduced 2AG and PG-G hydrolase activities. These results suggested that CES1 is in part responsible for the hydrolysis of 2AG and PG-Gs in THP1 cells, although it did not rule out a role for other hydrolases, especially with regard to 2AG metabolism since a substantial portion of its hydrolysis was not inactivated by the inhibitors. An enzyme (Mr 31-32 kDa) of unknown function was detected by serine hydrolase activity profiling of THP1 cells and may be a candidate. Finally, the amounts of in situ generated 2AG and PG-Gs in macrophages were enhanced by treating the cells with bioactive metabolites of OP insecticides. Collectively, the results suggest that in addition to MAGL and fatty-acid amide hydrolase (FAAH), which have both been documented to terminate endocannabinoid signaling, CES may also have a role. Furthermore, since PG-Gs have been shown to possess biological activities in their own right, CES may represent an important enzyme class that regulates their in vivo levels.
ESTHER : Xie_2010_Chem.Res.Toxicol_23_1890
PubMedSearch : Xie_2010_Chem.Res.Toxicol_23_1890
PubMedID: 21049984

Title : Inhibition of carboxylesterase activity of THP1 monocytes\/macrophages and recombinant human carboxylesterase 1 by oxysterols and fatty acids - Crow_2010_Biochim.Biophys.Acta_1801_31
Author(s) : Crow JA , Herring KL , Xie S , Borazjani A , Potter PM , Ross MK
Ref : Biochimica & Biophysica Acta , 1801 :31 , 2010
Abstract : Two major isoforms of human carboxylesterases (CEs) are found in metabolically active tissues, CES1 and CES2. These hydrolytic enzymes are involved in xenobiotic and endobiotic metabolism. CES1 is abundantly expressed in human liver and monocytes/macrophages, including the THP1 cell line; CES2 is expressed in liver but not in monocytes/macrophages. The cholesteryl ester hydrolysis activity in human macrophages has been attributed to CES1. Here, we report the direct inhibitory effects of several endogenous oxysterols and fatty acids on the CE activity of THP1 monocytes/macrophages and recombinant human CES1 and CES2. Using THP1 whole-cell lysates we found: (1) 27-hydroxycholesterol (27-HC) is a potent inhibitor of carboxylesterase activity (IC50=33 nM); (2) 24(S),25-epoxycholesterol had moderate inhibitory activity (IC(50)=8.1 microM); and (3) cholesterol, 7-ketocholesterol, 22(R)-hydroxycholesterol, 24(S)-hydroxycholesterol, and 25-hydroxycholesterol each had little inhibitory activity. 27-HC was a partially noncompetitive inhibitor of recombinant CES1 (K(iapp)=10 nM) and impaired intracellular CES1 activity following treatment of intact THP1 cells. In contrast, recombinant CES2 activity was not inhibited by 27-HC, suggesting isoform-selective inhibition by 27-HC. Furthermore, unsaturated fatty acids were better inhibitors of CES1 activity than saturated fatty acids, while CES2 activity was unaffected by any fatty acid. Arachidonic acid (AA) was the most potent fatty acid inhibitor of recombinant CES1 and acted by a noncompetitive mechanism (K(iapp)=1.7 microM); when not complexed to albumin, exogenous AA penetrated intact THP1 cells and inhibited CES1. Inhibition results are discussed in light of recent structural models for CES1 that describe ligand binding sites separate from the active site. In addition, oxysterol-mediated inhibition of CES1 activity was demonstrated by pretreatment of human liver homogenates or intact THP1 cells with exogenous 27-HC, which resulted in significantly reduced hydrolysis of the pyrethroid insecticide bioresmethrin, a CES1-specific xenobiotic substrate. Collectively, these findings suggest that CE activity of recombinant CES1, cell lysates, and intact cells can be impaired by naturally occurring lipids, which may compromise the ability of CES1 to both detoxify environmental pollutants and metabolize endogenous compounds in vivo.
ESTHER : Crow_2010_Biochim.Biophys.Acta_1801_31
PubMedSearch : Crow_2010_Biochim.Biophys.Acta_1801_31
PubMedID: 19761868

Title : Inhibition of carboxylesterase 1 is associated with cholesteryl ester retention in human THP-1 monocyte\/macrophages - Crow_2008_Biochim.Biophys.Acta_1781_643
Author(s) : Crow JA , Middleton BL , Borazjani A , Hatfield MJ , Potter PM , Ross MK
Ref : Biochimica & Biophysica Acta , 1781 :643 , 2008
Abstract : Cholesteryl esters are hydrolyzed by cholesteryl ester hydrolase (CEH) yielding free cholesterol for export from macrophages. Hence, CEH has an important regulatory role in macrophage reverse cholesterol transport (RCT). CEH and human carboxylesterase 1 (CES1) appear to be the same enzyme. CES1 is inhibited by oxons, the bioactive metabolites of organophosphate (OP) pesticides. Here, we show that CES1 protein is robustly expressed in human THP-1 monocytes/macrophages and its biochemical activity inhibited following treatment of cell lysates and intact cells with chlorpyrifos oxon, paraoxon, or methyl paraoxon (with nanomolar IC(50) values) or after immunodepletion of CES1 protein. CES1 protein expression in cells is unaffected by a 24-h paraoxon treatment, suggesting that the reduced hydrolytic activity is due to covalent inhibition of CES1 by oxons and not down-regulation of expression. Most significantly, treatment of cholesterol-loaded macrophages with either paraoxon (a non-specific CES inhibitor) or benzil (a specific CES inhibitor) caused enhanced retention of intracellular cholesteryl esters and a "foamy" phenotype, consistent with reduced cholesteryl ester mobilization. Thus, exposure to OP pesticides, which results in the inhibition of CES1, may also inhibit macrophage RCT, an important process in the regression of atherosclerosis.
ESTHER : Crow_2008_Biochim.Biophys.Acta_1781_643
PubMedSearch : Crow_2008_Biochim.Biophys.Acta_1781_643
PubMedID: 18762277
Gene_locus related to this paper: human-CES1

Title : Evaluation of the 'side door' in carboxylesterase-mediated catalysis and inhibition - Streit_2008_Biol.Chem_389_149
Author(s) : Streit TM , Borazjani A , Lentz SE , Wierdl M , Potter PM , Gwaltney SR , Ross MK
Ref : Biol Chem , 389 :149 , 2008
Abstract : Abstract Structures of mammalian carboxylesterases (CEs) reveal the presence of a 'side door' that is proposed to act as an alternative pore for the trafficking of substrates and products. p-Nitrobenzyl esterase (pnb CE) from Bacillus subtilis exhibits close structural homology and a similar side-door domain as mammalian CEs. We investigated the role of a specific 'gate' residue at the side door (i.e., Leu 362) during pnb CE-catalyzed hydrolysis of model esters, pesticides, and lipids. Recombinant pnb CE proteins containing mutations at position 362 demonstrated markedly lower kcat and kcat/Km values. The mutation with the most significant impact on catalysis was the L362R mutant (kcat/Km was 22-fold lower). Moreover, the ability of the L362R mutant to be inhibited by organophosphates (OP) was also lower. Investigation into the altered catalytic proficiency using pH-activity studies indicated that the catalytic triad of the mutant enzyme was preserved. Furthermore, viscosity variation and carbamate inhibition experiments indicated that rates of substrate association and acylation/deacylation were lower. Finally, recombinant CEs were found to possess lipolytic activity toward cholesteryl oleate and 2-arachidonylglycerol. In summary, the L362R mutant CE markedly slowed the rate of ester hydrolysis and was less sensitive to OP inhibition. The apparent causes of the diminished catalysis are discussed.
ESTHER : Streit_2008_Biol.Chem_389_149
PubMedSearch : Streit_2008_Biol.Chem_389_149
PubMedID: 18163883

Title : Hydrolysis of pyrethroids by human and rat tissues: examination of intestinal, liver and serum carboxylesterases - Crow_2007_Toxicol.Appl.Pharmacol_221_1
Author(s) : Crow JA , Borazjani A , Potter PM , Ross MK
Ref : Toxicol Appl Pharmacol , 221 :1 , 2007
Abstract : Hydrolytic metabolism of pyrethroid insecticides in humans is one of the major catabolic pathways that clear these compounds from the body. Rodent models are often used to determine the disposition and clearance rates of these esterified compounds. In this study the distribution and activities of esterases that catalyze pyrethroid metabolism have been investigated in vitro using several human and rat tissues, including small intestine, liver and serum. The major esterase in human intestine is carboxylesterase 2 (hCE2). We found that the pyrethroid trans-permethrin is effectively hydrolyzed by a sample of pooled human intestinal microsomes (5 individuals), while deltamethrin and bioresmethrin are not. This result correlates well with the substrate specificity of recombinant hCE2 enzyme. In contrast, a sample of pooled rat intestinal microsomes (5 animals) hydrolyze trans-permethrin 4.5-fold slower than the sample of human intestinal microsomes. Furthermore, it is demonstrated that pooled samples of cytosol from human or rat liver are approximately 2-fold less hydrolytically active (normalized per mg protein) than the corresponding microsomal fraction toward pyrethroid substrates; however, the cytosolic fractions do have significant amounts (approximately 40%) of the total esteratic activity. Moreover, a 6-fold interindividual variation in carboxylesterase 1 protein expression in human hepatic cytosols was observed. Human serum was shown to lack pyrethroid hydrolytic activity, but rat serum has hydrolytic activity that is attributed to a single CE isozyme. We purified the serum CE enzyme to homogeneity to determine its contribution to pyrethroid metabolism in the rat. Both trans-permethrin and bioresmethrin were effectively cleaved by this serum CE, but deltamethrin, esfenvalerate, alpha-cypermethrin and cis-permethrin were slowly hydrolyzed. Lastly, two model lipase enzymes were examined for their ability to hydrolyze pyrethroids. However, no hydrolysis products could be detected. Together, these results demonstrate that extrahepatic esterolytic metabolism of specific pyrethroids may be significant. Moreover, hepatic cytosolic and microsomal hydrolytic metabolism should each be considered during the development of pharmacokinetic models that predict the disposition of pyrethroids and other esterified compounds.
ESTHER : Crow_2007_Toxicol.Appl.Pharmacol_221_1
PubMedSearch : Crow_2007_Toxicol.Appl.Pharmacol_221_1
PubMedID: 17442360

Title : Hydrolytic metabolism of pyrethroids by human and other mammalian carboxylesterases - Ross_2006_Biochem.Pharmacol_71_657
Author(s) : Ross MK , Borazjani A , Edwards CC , Potter PM
Ref : Biochemical Pharmacology , 71 :657 , 2006
Abstract : Pyrethroid chemicals are attractive alternatives to the organophosphates (OPs) because of their selective toxicity against pests rather than mammals. The carboxylesterases (CEs) are hepatic enzymes that metabolize ester-containing xenobiotics such as pyrethroids. The primary aim of this study was to gain insight into the catalytic properties of the CE enzymes in humans that metabolize pyrethroids, while a secondary aim was to investigate pyrethroid metabolism using CEs from other mammalian species. Pure human CEs (hCE-1 and hCE-2), a rabbit CE (rCE), and two rat CEs (Hydrolases A and B) were used to study the hydrolytic metabolism of the following pyrethroids: 1Rtrans-resmethrin (bioresmethrin), 1RStrans-permethrin, and 1RScis-permethrin. hCE-1 and hCE-2 hydrolyzed trans-permethrin 8- and 28-fold more efficiently than cis-permethrin (when k(cat)/K(m) values were compared), respectively. In contrast, hydrolysis of bioresmethrin was catalyzed efficiently by hCE-1, but not by hCE-2. The kinetic parameters for the pure rat and rabbit CEs were qualitatively similar to the human CEs when hydrolysis rates of the investigated pyrethroids were evaluated. Further, a comparison of pyrethroid hydrolysis by hepatic microsomes from rats, mice, and humans indicated that the rates for each compound were similar between species, which further supports the use of rodent models for pyrethroid metabolism studies. An eight-fold range in hydrolytic rates for 11 individual human liver samples toward trans-permethrin was also found, although this variability was not related to the levels of hCE-1 protein in each sample. We also determined that the CE inhibitor 2-chloro-3,4-dimethoxybenzil blocked hCE-2-catalyzed trans-permethrin hydrolysis 36 times more potently than hCE-1. Thus, this inhibitor will be useful in future studies that examine CE-mediated metabolism of pyrethroids. While there are likely other esterases in human liver that hydrolyze pyrethroids, the results of this study clearly demonstrate that hCE-1 and hCE-2 are human pyrethroid-hydrolyzing CEs.
ESTHER : Ross_2006_Biochem.Pharmacol_71_657
PubMedSearch : Ross_2006_Biochem.Pharmacol_71_657
PubMedID: 16387282