Abstract Phosphatidylcholine (PC) is a major constituent of biological membranes and a component of serum lipoproteins and pulmonary surfactants. The PC and other glycerophospholipid compositions of membranes change dynamically through stimulus‐dependent and independent pathways, principally by the action of two different types of enzymes; phospholipase A2 [EC 3.1.1.4] and acyl‐CoA:lysophospholipid acyltransferase [EC 2.3.1.23]. Phospholipase A2 is a key enzyme that catalyzes deacylation of the sn‐2 position of glycerophospholipids. This enzyme is critical in the remodeling of membrane lipids and formation of two subclasses of lipid mediators, fatty acid derivatives and lysophospholipids. Among many different subtypes of phospholipase A2 enzymes, we found that cytosolic phospholipase A2α (cPLA2α) is important in various pathological and physiological responses. Here, we summarize the phenotypes resulting from genetic ablation of cPLA2α, and the properties of newly discovered enzymes in the cPLA2 family. Comprehensive analysis of lipid mediators using liquid chromatography‐tandem mass spectrometry (LC‐MS/MS) is useful for understanding the roles of individual mediators in physiological and pathological processes. iubmb Life, 58: 328‐333, 2006
In mammalian organisms, fatty acids (FAs) exist mostly in esterified forms, as building blocks of phospholipids, triglycerides, and cholesteryl esters, while some exist as non-esterified free FAs. The absolute quantification of FA species in total lipids or in a specific lipid class is critical in lipid-metabolism studies. To quantify FAs in biological samples, gas chromatography-hydrogen flame ionization detection (GC-FID)-based methods have been used as highly robust and reliable techniques. Prior to GC-FID analysis, FAs need to be derivatized to volatile FA methyl esters (FAMEs). The derivatization of unsaturated FAs using classical derivatization methods that rely on high reaction temperature requires skill; consequently, the quantification results are often unreliable. The recently available FA-methylation procedure rapidly and reliably derivatizes a variety of FA species, including poly-unsaturated FAs (PUFAs). To analyze FAs in mammalian tissue samples, lipid extraction and fractionation are also critical for robust analysis. In this report, we describe a whole protocol for the GC-FID-based FA quantification of mammalian tissue samples, including lipid extraction, fractionation, derivatization, and quantification. The protocol is useful when various FAs, especially unsaturated FAs, need to be reliably quantified.
Excess energy intake causes obesity, which leads to insulin resistance and various other complications of metabolic syndrome, including diabetes, atherosclerosis, dyslipidemia, and nonalcoholic fatty liver disease. Although recent studies have depicted altered lipid metabolism as an underlying feature, the detailed mechanisms are still unclear. Here we describe a possible role in high-fat diet (HFD)-induced obesity for monoacylglycerol lipase (MGL), an enzyme that is also known to hydrolyze the endocannabinoid 2-arachidonoylglycerol in brain. MGL-deficient [MGL-knockout (KO)] mice fed a HFD gained less body weight than wild-type mice and were protected from insulin resistance and hepatic steatosis. Food intake and energy expenditure were not altered in MGL-KO mice, but blood triglyceride levels after oral olive oil gavage were suppressed, indicating a role for MGL in intestinal fat absorption. Experiments with cannabinoid receptor type 1 (CB1)/MGL double-KO mice revealed that these phenotypes may include mechanisms that are independent of CB1-receptor–mediated endocannabinoid functions. We also noted that MGL-KO mice had less preference for HFD over normal chow diet. Oral but not intraperitoneal lipid administration strongly suppressed the appetites of MGL-KO and CB1/MGL double-KO mice, but not of wild-type and CB1-KO mice. Appetite suppression was reversed by vagotomy, suggesting involvement of MGL in the gut–brain axis regulation of appetite. Our results provide mechanistic insights of MGL's role in diet-induced obesity, lipid metabolic disorder, and regulation of appetite.—Yoshida, K., Kita, Y., Tokuoka, S. M., Hamano, F., Yamazaki, M., Sakimura, K., Kano, M., Shimizu, T. Monoacylglycerol lipase deficiency affects diet-induced obesity, fat absorption, and feeding behavior in CB1 cannabinoid receptor–deficient mice. FASEB J. 33, 2484–2497 (2019). www.fasebj.org
UV radiation induces systemic immunosuppression. Because nonsteroidal anti-inflammatory drugs suppress UV-induced immunosuppression, prostanoids have been suspected as a crucial mediator of this UV effect. However, the identity of the prostanoid involved and its mechanism of action remain unclear. Here, we addressed this issue by subjecting mice deficient in each prostanoid receptor individually or mice treated with a subtype-specific antagonist to UV irradiation. Mice treated with an antagonist for prostaglandin E receptor subtype 4 (EP4), but not those deficient in other prostanoid receptors, show impaired UV-induced immunosuppression, whereas administration of an EP4 agonist rescues the impairment of the UV-induced immunosuppression in indomethacin-treated mice. The EP4 antagonist treatment suppresses an increase in the number of CD4(+)/forkhead box P3-positive (Foxp3(+)) regulatory T cells (Treg cells) in the peripheral lymph nodes (LNs) and dendritic cells expressing DEC205 in the LNs and the skin after UV irradiation. Furthermore, the EP4 antagonist treatment down-regulates UV-induced expression of receptor activator of NF-κB ligand (RANKL) in skin keratinocytes. Finally, administration of anti-RANKL antibody abolishes the restoration of UV-induced immunosuppression by EP4 agonism in indomethacin-treated mice. Thus, prostaglandin E(2) (PGE(2))-EP4 signaling mediates UV-induced immunosuppression by elevating the number of Treg cells through regulation of RANKL expression in the epidermis.
