Inactivation of lipid glyceryl ester metabolism in human THP1 monocytes/macrophages by activated organophosphorus insecticides: role of carboxylesterases 1 and 2.
Inactivation of lipid glyceryl ester metabolism in human THP1 monocytes/macrophages by activated organophosphorus insecticides: role of carboxylesterases 1 and 2.
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活化的有机磷杀虫剂对人 THP1 单核细胞/巨噬细胞中脂质甘油酯代谢的灭活：羧酸酯酶 1 和 2 的作用。
DOI：
10.1021/tx1002194
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发表时间：
2010-12-20
影响因子：
4.1
通讯作者：
Ross MK
中科院分区：
文献类型：
--
作者：
Xie S;Borazjani A;Hatfield MJ;Edwards CC;Potter PM;Ross MK
羧酸酯酶（Carboxylesterases，CES）在农药和药物代谢中具有重要作用，并参与含酯异生物质在哺乳动物体内的清除。CES表达水平最高的组织是肝脏和小肠。除了外源性物质，CES还利用其广泛的底物特异性来水解内源性物质，如胆固醇酯和三酰甘油。在这里，我们确定了两种人CES亚型，CES 1和CES 2，是否水解内源性大麻素2-花生四烯酰甘油（2AG）和花生四烯酰胺（AEA），以及两种前列腺素甘油酯（PG-Gs），这是由COX介导的2AG氧化形成的。我们表明，重组CES 1和CES 2有效地水解2AG花生四烯酸（AA），但不含酰胺的AEA。CES 1和CES 2介导的2AG水解的稳态动力学参数分别为：kcat，59和43 min −1; Km，49和46μM; kcat/Km，1.2和0.93μM−1 min−1。kcat/Km值与大鼠单酰基甘油脂酶（MAGL）催化的2AG水解的公开值相当。此外，我们发现CES 1和CES 2也有效地水解PGE 2-G和PGF 2 α-G。此外，当用外源性2AG或PG-G（10μM，1 h）处理培养的人THP 1巨噬细胞时，在培养基中检测到大量AA或PGs;然而，用对氧磷处理后，巨噬细胞代谢这些化合物的能力受到抑制（60-80%），对氧磷是杀虫剂敌百虫的毒性代谢产物。将THP 1细胞裂解物与靶向CES 1的小分子抑制剂（噻吩并[3，2-e][1]苯并噻吩-4，5-二酮或JZL 184）一起孵育显著降低了脂质甘油酯水解酶活性（2AG为40-50%，PG-Gs为80-95%）。免疫耗竭CES 1也显著降低2AG和PG-G水解酶活性。这些结果表明，CES 1在一定程度上负责THP 1细胞中2AG和PG-Gs的水解，尽管它不排除其他水解酶的作用，特别是关于2AG代谢，因为其水解的大部分没有被抑制剂灭活。通过THP 1细胞的丝氨酸水解酶活性分析检测到一种未知功能的酶（Mr 31- 32 kDa），可能是候选酶。最后，用OP杀虫剂的生物活性代谢产物处理巨噬细胞，增加了巨噬细胞中原位产生的2AG和PG-Gs的量。总的来说，结果表明，除了MAGL和脂肪酸酰胺水解酶（FAAH），这两个都被证明终止内源性大麻素信号，CES也可能有一个作用。此外，由于PG-G已被证明具有自身的生物活性，CES可能代表调节其体内水平的重要酶类。
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 43min−1; Km, 49 and 46μM; kcat/Km, 1.2 and 0.93μM−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 PGF2α-G. In addition, when cultured human THP1 macrophages were treated with exogenous 2AG or PG-G (10μM, 1h), 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 31-32kDa) 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.
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影响因子：
4.8
作者：
Kozak, KR;Crews, BC;Marnett, LJ
通讯作者：
Marnett, LJ
影响因子：
5.6
作者：
Bencharit, Sompop;Edwards, Carol C.;Redinbo, Matthew R.
通讯作者：
Redinbo, Matthew R.
影响因子：
--
作者：
Blankman, Jacqueline L.;Simon, Gabriel M.;Cravatt, Benjamin F.
通讯作者：
Cravatt, Benjamin F.
影响因子：
4.8
作者：
Kozak, KR;Rowlinson, SW;Marnett, LJ
通讯作者：
Marnett, LJ
影响因子：
2.9
作者：
Kidd, D;Liu, YS;Cravatt, BF
通讯作者：
Cravatt, BF