Fatty Acid Esters Of Hydroxy Fatty Acids Research Articles

Identification of short-chain fatty acid esters of hydroxy fatty acids (SFAHFAs) in a murine model by nontargeted analysis using ultra-high-performance liquid chromatography/linear ion trap quadrupole-Orbitrap mass spectrometry.

Fatty acid esters of hydroxy fatty acids (FAHFAs) are recently discovered endogenous lipids with outstanding health benefits. FAHFAs are known to exhibit antioxidant, antidiabetic and anti-inflammatory properties. The number of known long-chain FAHFAs in mammalian tissues and dietary resources increased recently because of the latest developments in high-resolution tandem mass spectrometry techniques. However, there are no reports on the identification of short-chain fatty acid esterified hydroxy fatty acids (SFAHFAs). Intestinal contents, tissues, and plasma of rats fed with high-fat diet (HFD) and normal diet (ND) were analyzed for fatty acids, hydroxy fatty acids, and FAHFAs using ultra-high-performance liquid chromatography (UHPLC) and linear trap quadrupole-Orbitrap mass spectrometry (LTQ Orbitrap MS) with negative heated electrospray ionization. Untargeted analysis of total lipid extracts from murine samples (male 13-week-old WKAH/HKmSlc rats) led to the identification of several new SFAHFAs of acetic acid or propanoic acid esterified long-chain (>C20)-hydroxy fatty acids. Furthermore, MS3 analysis revealed the position of the hydroxyl group in the long-chain fatty acid as C-2. The relative amounts of SFAHFAs were quantified in intestinal contents and their tissues (Cecum, small intestine, and large intestine), liver, and plasma of rats fed with HFD and ND. The large intestine showed the highest abundance of SFAHFAs with a concentration range from 0.84 to 57 pmol/mg followed by the cecum with a range of 0.66 to 28.6 pmol/mg. The SFAHFAs were significantly altered between the HFD and ND groups, with a strong decreasing tendency under HFD conditions. Identification of these novel SFAHFAs can contribute to a better understanding of the chemical and biological properties of individual SFAHFAs and their possible sources in the gut, which in turn helps us tackle the role of these lipids in various metabolic diseases.

Analytical Methods for the Determination of Fatty Acid Esters of Hydroxy Fatty Acids (FAHFAs) in Biological Samples, Plants and Foods.

Fatty acid esters of hydroxy fatty acids (FAHFAs) constitute a class of recently identified novel lipids exhibiting anti-diabetic and anti-inflammatory effects. Due to their high biological significance, a tremendous effort has been devoted to the development of analytical methods for the detection and quantitation of FAHFAs during the last five years. The analysis of FAHFAs is very challenging due to the great number of possible regio-isomers arising from the great number of possible combinations of FAs with HFAs, and the low abundancies of FAHFAs in biological samples. The aim of this review article is to summarize all the cutting-edge analytical methodologies for the determination of FAHFAs in biological samples, plant tissues and food matrices, with emphasis on extraction and analysis steps. All the analytical methodologies rely on the use of liquid chromatography–mass spectrometry (LC-MS), providing high sensitivity due to the MS detection. Powerful and robust analytical methodologies may highly contribute in studying FAHFAs levels under various biomedical conditions, and facilitate our understanding of the role of these lipid species in physiological and pathological conditions.

Charge-switch derivatization of fatty acid esters of hydroxy fatty acids via gas-phase ion/ion reactions

Branched fatty acid esters of hydroxy fatty acids (FAHFAs) are a recently discovered class of endogenous bioactive lipids with anti-diabetic and anti-inflammatory effects. Identification of FAHFAs is challenging due to both the relatively low abundance of these metabolites in most biological samples and the significant structural diversity arising from the co-occurrence of numerous regioisomers. Ultimately, development of sensitive analytical techniques that enable rapid and unambiguous identification of FAHFAs is integral to understanding their diverse physiological functions in health and disease. While a battery of mass spectrometry (MS) based methods for complex lipid analysis has been developed, FAHFA identification presents specific challenges to conventional approaches. Notably, while the MS2 product ion spectra of [FAHFA – H]¯ anions afford the assignment of fatty acid (FA) and hydroxy fatty acid (HFA) constituents, FAHFA regioisomers are usually indistinguishable by this approach. Here, we report the development of a novel MS-based technique employing charge inversion ion/ion reactions with tris-phenanthroline magnesium complex dications, Mg(Phen)32+, to selectively and efficiently derivatize [FAHFA – H]¯ anions in the gas phase, yielding fixed-charge cations. Subsequent activation of [FAHFA – H + MgPhen2]+ cations yield product ions that facilitate the assignment of FA and HFA constituents, pinpoints unsaturation sites within the FA moiety, and elucidates ester linkage regiochemistry. Collectively, the presented approach represents a rapid, entirely gas-phase method for near-complete FAHFA structural elucidation and confident isomer discrimination without the requirement for authentic FAHFA standards.

Oxidation of polyunsaturated fatty acids to produce lipid mediators.

The chemistry, biochemistry, pharmacology and molecular biology of oxylipins (defined as a family of oxygenated natural products that are formed from unsaturated fatty acids by pathways involving at least one step of dioxygen-dependent oxidation) are complex and occasionally contradictory subjects that continue to develop at an extraordinarily rapid rate. The term includes docosanoids (e.g. protectins, resolvins and maresins, or specialized pro-resolving mediators), eicosanoids and octadecanoids and plant oxylipins, which are derived from either the omega-6 (n-6) or the omega-3 (n-3) families of polyunsaturated fatty acids. For example, the term eicosanoid is used to embrace those biologically active lipid mediators that are derived from C20 fatty acids, and include prostaglandins, thromboxanes, leukotrienes, hydroxyeicosatetraenoic acids and related oxygenated derivatives. The key enzymes for the production of prostanoids are prostaglandin endoperoxide H synthases (cyclo-oxygenases), while lipoxygenases and oxidases of the cytochrome P450 family produce numerous other metabolites. In plants, the lipoxygenase pathway from C18 polyunsaturated fatty acids yields a variety of important products, especially the jasmonates, which have some comparable structural features and functions. Related oxylipins are produced by non-enzymic means (isoprostanes), while fatty acid esters of hydroxy fatty acids (FAHFA) are now being considered together with the oxylipins from a functional perspective. In all kingdoms of life, oxylipins usually act as lipid mediators through specific receptors, have short half-lives and have functions in innumerable biological contexts.

Discovery of Eicosapentaenoic Acid Esters of Hydroxy Fatty Acids as Potent Nrf2 Activators.

Branched fatty acid esters of hydroxy fatty acids (FAHFAs) are a recently discovered class of biologically active lipids with anti-inflammatory and anti-diabetic properties. Despite the possible link between endogenous FAHFA levels and nuclear factor erythroid 2-related factor 2 (Nrf2), their possible function as antioxidants and the mechanisms involved in this are unknown. Here, we investigate FAHFAs’ plausible antioxidant potential with reference to their effect on the Nrf2 levels, oxidative stress, and lipid droplet oxidation in human hepatocytes (C3A). Six authentic FAHFAs were chemically synthesized and performed activity-based screening by reporter gene assay. Among them, eicosapentaenoic acid (EPA) esterified 12-hydroxy stearic acid (12-HSA) and 12-hydroxy oleic acid (12-HOA) FAHFAs showed less cytotoxicity compared to their free fatty acids and potent activators of Nrf2. To define their mode of action, relative levels of nuclear Nrf2 were determined, which found a higher amount of Nrf2 in nucleus of cells treated with 12-EPAHSA compared to the control. Furthermore, 12-EPAHSA increased the expression of Nrf2-dependent antioxidant enzyme genes (NQO1, GCLM, GCLC, SOD-1, and HO-1). Fluorescence imaging analysis of linoleic-acid-induced lipid droplets (LDs) in C3A cells treated with 12-EPAHSA revealed the strong inhibition of small-size LD oxidation. These results suggest that EPA-derived FAHFAs as a new class of lipids with less cytotoxicity, and strong Nrf2 activators with plausible antioxidant effects via the induction of cytoprotective proteins against oxidative stress, induced cellular damage.

AIG1 and ADTRP are endogenous hydrolases of fatty acid esters of hydroxy fatty acids (FAHFAs) in mice

Fatty acid esters of hydroxy fatty acids (FAHFAs) are a newly discovered class of signaling lipids with anti-inflammatory and anti-diabetic properties. However, the endogenous regulation of FAHFAs remains a pressing but unanswered question. Here, using MS-based FAHFA hydrolysis assays, LC-MS-based lipidomics analyses, and activity-based protein profiling, we found that androgen-induced gene 1 (AIG1) and androgen-dependent TFPI-regulating protein (ADTRP), two threonine hydrolases, control FAHFA levels in vivo in both genetic and pharmacologic mouse models. Tissues from mice lacking ADTRP (Adtrp-KO), or both AIG1 and ADTRP (DKO) had higher concentrations of FAHFAs particularly isomers with the ester bond at the 9th carbon due to decreased FAHFA hydrolysis activity. The levels of other lipid classes were unaltered indicating that AIG1 and ADTRP specifically hydrolyze FAHFAs. Complementing these genetic studies, we also identified a dual AIG1/ADTRP inhibitor, ABD-110207, which is active in vivo Acute treatment of WT mice with ABD-110207 resulted in elevated FAHFA levels, further supporting the notion that AIG1 and ADTRP activity control endogenous FAHFA levels. However, loss of AIG1/ADTRP did not mimic the changes associated with pharmacologically administered FAHFAs on extent of upregulation of FAHFA levels, glucose tolerance, or insulin sensitivity in mice, indicating that therapeutic strategies should weigh more on FAHFA administration. Together, these findings identify AIG1 and ADTRP as the first endogenous FAHFA hydrolases identified and provide critical genetic and chemical tools for further characterization of these enzymes and endogenous FAHFAs to unravel their physiological functions and roles in health and disease.

Triacylglycerol-Rich Oils of Marine Origin are Optimal Nutrients for Induction of Polyunsaturated Docosahexaenoic Acid Ester of Hydroxy Linoleic Acid (13-DHAHLA) with Anti-Inflammatory Properties in Mice.

The docosahexaenoic acid ester of hydroxy linoleic acid (13-DHAHLA) is a bioactive lipid with anti-inflammatory properties from the family of fatty acid esters of hydroxy fatty acids (FAHFA). To explore the biosynthesis of 13-DHAHLA from dietary oils, C57BL/6N mice are gavaged for 8 days with various corn oil/marine oil mixtures containing the same amount of DHA. Plasma levels of omega-3 FAHFAs are influenced by the lipid composition of the mixtures but do not reflect the changes in bioavailability of polyunsaturated fatty acids in plasma. Triacylglycerol-bound DHA and linoleic acid serve as more effective precursors for 13-DHAHLA synthesis than DHA bound in phospholipids or wax esters. Both 13(S)- and 13(R)-DHAHLA inhibit antigen and PGE2 -induced chemotaxis and degranulation of mast cells to a comparable extent and 13(S)-DHAHLA is identified as the predominant isomer in mouse adipose tissue. Here, the optimal nutritional source of DHA is identified, which supports production of anti-inflammatory FAHFAs, as triacylglycerol-based marine oil and also reveals a possible role of triacylglycerols in the synthesis of FAHFA lipokines.

In-Silico-Generated Library for Sensitive Detection of 2-Dimethylaminoethylamine Derivatized FAHFA Lipids Using High-Resolution Tandem Mass Spectrometry.

Fatty acid esters of hydroxy fatty acids (FAHFAs) are a family of recently discovered lipids with important physiological functions in mammals and plants. However, low detection sensitivity in negative ionization mode mass spectrometry makes low-abundance FAHFA challenging to analyze. A 2-dimethylaminoethylamine (DMED) based chemical derivatization strategy was recently reported to improve the MS sensitivity of FAHFAs by labeling FAHFAs with a positively ionizable tertiary amine group. To facilitate reliable, high-throughput, and automatic annotation of these compounds, a DMED-FAHFA in silico library containing 4290 high-resolution tandem mass spectra covering 264 different FAHFA classes was developed. The construction of the library was based on the heuristic information from MS/MS fragmentation patterns of DMED-FAHFA authentic standards, and then, the patterns were applied to computer-generated DMED-FAHFAs. The developed DMED-FAHFA in silico library was demonstrated to be compatible with library search software NIST MS Search and the LC-MS/MS data processing tool MS-DIAL to guarantee high-throughput and automatic annotations. Applying the in silico library in Arabidopsis thaliana samples for profiling FAHFAs by high-resolution LC-MS/MS enabled the annotation of 19 DMED-FAHFAs from 16 families, including 3 novel compounds. Using the in silico library largely decreased the false-positive annotation rate in comparison to low-resolution LC-MS/MS. The developed library, MS/MS spectra, and development templates are freely available for commercial and noncommercial use at https://zenodo.org/record/3606905.

Long-term high intake of 9-PAHPA or 9-OAHPA increases basal metabolism and insulin sensitivity but disrupts liver homeostasis in healthy mice

Branched fatty acid esters of hydroxy fatty acids (FAHFAs) are a new family of endogenous lipids recently discovered. Several studies reported that some FAHFAs have antidiabetic and anti-inflammatory effects. The objective of this study was to explore the impact of two FAHFAs, 9-PAHPA or 9-OAHPA, on the metabolism of mice. C57Bl/6J male mice, 6 weeks old, were divided into 3 groups of 10 mice each. One group received a control diet and the two others groups received the control diet supplemented with 9-PAHPA or 9-OAHPA for 12 weeks. Mouse weight and body composition were monitored throughout the study. Some days before euthanasia, energy expenditure, glucose tolerance and insulin sensitivity were also determined. After sacrifice, blood and organs were collected for relevant molecular, biochemical and histological analyses. Although high intake of 9-PAHPA or 9-OAHPA increased basal metabolism, it had no direct effect on body weight. Interestingly, the 9-PAHPA or 9-OAHPA intake increased insulin sensitivity but without modifying glucose tolerance. Nevertheless, 9-PAHPA intake induced a loss of glucose-stimulated insulin secretion. Surprisingly, both studied FAHFAs induced hepatic steatosis and fibrosis in some mice, which were more marked with 9-PAHPA. Finally, a slight remodeling of white adipose tissue was also observed with 9-PAHPA intake. In conclusion, the long-term high intake of 9-PAHPA or 9-OAHPA increased basal metabolism and insulin sensitivity in healthy mice. However, this effect, highly likely beneficial in a diabetic state, was accompanied by manifest liver damage in certain mice that should deserve special attention in both healthy and pathological studies.

FAHFA footprint in the visceral fat of mice across their lifespan

Fatty acid esters of hydroxy fatty acids (FAHFAs) are a new class of endogenous lipids with anti-inflammatory and anti-diabetic effects, having the potential to treat type 2 diabetes (T2D). In view of the important regulatory and therapeutic actions of FAHFAs on age-related diseases such as T2D, we hypothesized that they may also play crucial roles in the growth, development, and aging process. Here, we investigated the FAHFA footprint in the visceral adipose tissue (VAT) of mice across lifespan to attain potential clues for understanding the roles of FAHFAs in growth, development, and aging using chemical isotope labeling assisted liquid chromatography-mass spectrometry. VAT samples were harvested from 80 C57BL/6J male mice of nine different ages (1, 2, 3, 6, 9, 12, 15, 18, and 24 months). The results showed that a total of 51 FAHFA families, including 301 regioisomers, were detected in the VAT of mice of all ages, and the number of FAHFAs (both family and regioisomer) in VAT increased with age, from 35 families (186 ± 0 regioisomers) at 1-month-old mice to 46 families (278 ± 6 regioisomers) in 18-month-old mice. Furthermore, the content of 12 FAHFA families per 100 mg of VAT of mice was highly correlated with age, and was usually low in the middle-age (3–15 months). However, because the VAT mass was 4–5 fold higher in middle-aged mice compared to younger or older mice, the total amount of most of the FAHFA regioisomers in VAT was increased in middle-aged mice. To the best of our knowledge, this is the first study to show that the number of regioisomers from 51 FAHFA families and abundance of 15 FAHFA families are strongly dependent on age, which would be helpful for understanding the mechanisms underlying the effects of FAHFAs on growth, development, and aging.

Interventional effects of the direct application of \u201cSanse powder\u201d on knee osteoarthritis in rats as determined from lipidomics via UPLC-Q-Exactive Orbitrap MS

BackgroundOur previous clinical evidence suggested that the direct application of “Sanse powder” the main ingredient of “Yiceng” might represent an alternative treatment for knee osteoarthritis. However, the mechanism underlying its effect is poorly understood. In this study, we investigated the mechanism of the effect of direct “Sanse powder” application for the treatment of knee osteoarthritis (KOA) in rats by using lipidomics.MethodsKOA rats were established by cutting the anterior cruciate ligament, and the cold pain threshold and mechanical withdrawal threshold (MWT) of seven rats from each group were measured before modelling (0 days) and at 7, 14, 21 and 28 days after modelling. Histopathological evaluation of the synovial tissue was performed by haematoxylin and eosin (H&E) staining after modelling for 28 days. Interleukin-1β (IL-1β), pro-interleukin-1β (pro-IL-1β) and tumor necrosis factor-α (TNF-α) proteins in synovial tissue were measured by western blot, and the mRNA expression levels of IL-1β and TNF-α in synovial tissue were measured using Real-time reverse transcription polymerase chain reaction (qRT-PCR), the levels of IL-1β and TNF-α in rat serum were measured by enzyme-linked immunosorbent assay (ELISA), Serum lipid profiles were obtained by using ultra-performance liquid chromatography combined with quadrupole-Exactive Orbitrap mass spectrometry (UPLC-Q-Exactive Orbitrap MS).ResultsThe results confirmed that the direct application of “Sanse powder” had a significant protective effect against KOA in rats. Treatment with “Sanse powder” not only attenuated synovial tissue inflammation but also increased the levels of the cold pain threshold and MWT. In addition, the lipidomics results showed that the levels of diacylglycerol (DAG), triacylglycerols (TAGs), lysophosphatidylcholine (LPC), phosphatidylcholine (PC), fatty acid esters of hydroxy fatty acids (FAHFAs), and phosphatidylethanolamine (PE) were restored almost to control levels following treatment.ConclusionsLipidomics provides a better understanding of the actions of direct application “Sanse powder” therapy for KOA.

Sensitive analysis of fatty acid esters of hydroxy fatty acids in biological lipid extracts by shotgun lipidomics after one-step derivatization.

Branched fatty acid esters of hydroxy fatty acids (FAHFAs) are an important family of endogenous lipids, possessing antidiabetic and anti-inflammatory functions. Therefore, analysis of FAHFAs in biological samples obtained under healthy and disease states can uncover underlying mechanisms of various relevant disorders (e.g., diabetes and autoimmune diseases). Up to now, due to their extremely low abundance, the determination of the changed levels of these species is still a huge challenge, even though great efforts have been made by utilizing liquid chromatography-tandem mass spectrometry with or without derivatization. Herein, we described a novel method for analysis of FAHFAs present in lipid extracts of biological examples after solid-phase extraction and chemical derivatization with one authentic FAHFA specie as an internal standard based on the principles of multi-dimensional mass spectrometry-based shotgun lipidomics. The approach possessed marked sensitivity, high specificity, and broad linear dynamic range of over 3 orders without obvious matrix effects. Moreover, after chemical derivatization, the molecular masses of FAHFAs shift from an overlapped region with ceramide species to a new region without overlaps, removing these contaminating signals from ceramides, and thereby reducing the false results of FAHFAs. Finally, this novel method was successfully applied for determining FAHFAs levels in varieties of representative biological samples, including plasma from lean and overweight/obese individuals of normoglycemia, and tissue samples (such as liver and white adipose tissue from diabetic (db/db) mice). We revealed significant alterations of FAHFAs in samples under patho(physio)logical conditions compared to their respective controls. Taken together, the developed method could greatly contribute to studying altered FAHFA levels under a variety of biological/biomedical conditions, and facilitate the understanding of these lipid species in the patho(physio)logical process.

Linoleic acid esters of hydroxy linoleic acids are anti-inflammatory lipids found in plants and mammals

Fatty acid esters of hydroxy fatty acids (FAHFAs) are a recently discovered class of biologically active lipids. Here we identify the linoleic acid ester of 13-hydroxy linoleic acid (13-LAHLA) as an anti-inflammatory lipid. An oat oil fraction and FAHFA-enriched extract from this fraction showed anti-inflammatory activity in a lipopolysaccharide-induced cytokine secretion assay. Structural studies identified three LAHLA isomers (15-, 13-, and 9-LAHLA) as being the most abundant FAHFAs in the oat oil fraction. Of these LAHLAs, 13-LAHLA is the most abundant LAHLA isomer in human serum after ingestion of liposomes made of fractionated oat oil, and it is also the most abundant endogenous LAHLA in mouse and human adipose tissue. As a result, we chemically synthesized 13-LAHLA for biological assays. 13-LAHLA suppresses lipopolysaccharide-stimulated secretion of cytokines and expression of pro-inflammatory genes. These studies identify LAHLAs as an evolutionarily conserved lipid with anti-inflammatory activity in mammalian cells.

Analysis of Fatty Acid Esters of Hydroxyl Fatty Acid in Selected Plant Food.

Metabolic syndrome, characterized by obesity, low-grade inflammation, insulin resistance, hyperglycemia, dyslipidemia and hypertension, is a major risk factor for cardiovascular mortality. Preclinical studies on recently discovered classes of lipids - fatty acid esters of hydroxy fatty acids (FAHFA) have revealed their anti-inflammatory and insulin-sensitizing potential. The FAHFA levels are significantly decreased in insulin-resistant individuals, their application exhibited anti-inflammatory effects and restoring the glucose-insulin homeostasis. The aim of our research was to analyze the overall FAHFA composition in a common diet, as only a partial FAHFA composition has been revealed so far (only the PAHSA subclass was analyzed in a few foods). A new approach to the FAHFAs analysis includes nano-LC and post-column modifier followed by negative ion mass spectrometry, in order to obtain maximum sensitivity. Analysis of different foods - oat (whole grain, coarse flakes and fine flakes), apple, clementine, lemon, strawberry, blueberry, mango, kiwi, avocado, pineapple, banana, onion, garlic, cherry tomato, carrot, parsley root, pepper and radish - exhibited wide inter-food variation in the FAHFA profiles. Sixteen analyzed FAHFAs (palmitic, oleic, palmitoleic and stearic hydroxy-esters) showed microgram to low nanogram levels (0.165ng/g - 32μg/g FW), with the highest abundancy in oat, clementine, garlic and pineapple. Stearic acid hydroxy stearic acid (SAHSA) was the most abundant FAHFA, especially in the food with antioxidative, anti-inflammatory and beneficial metabolic effects. In contrary, the PAHSA - previously proven to have the strongest antihyperglycemic and insulin-sensitizing effects, was not present in some foods (radish, avocado, mango, lemon, cherry tomato, kiwi). Our study proves the importance of overall FAHFA analysis in food (especially in a functional food), because of their potential metabolic benefits and possible future incorporation in special diets.

Synthesis of chemically edited derivatives of the endogenous regulator of inflammation 9-PAHSA.

Fatty acid esters of hydroxy fatty acids (FAHFAs) are a growing class of natural products found in organisms ranging from plants to humans. The roles these endogenous derivatives of fatty acids play in biology and their novel pathways for controlling inflammation have increased our understanding of basic human physiology. FAHFAs incorporate diverse fatty acids into their structures, however, given their recent discovery non-natural derivatives have not been a focus and as a result structure-activity relationships remain unknown. The importance of the long chain hydrocarbons extending from the ester linkage as they relate to anti-inflammatory activity is unknown. Herein the systematic removal of carbons from either the hydroxy fatty acid or fatty acid regions of the most studied FAHFA, palmitic acid ester of 9-hydroxystearic acid (9-PAHSA), was achieved and these synthetic, abridged analogs were tested for their ability to attenuate IL-6 production. Reduction of the carbon chain lengths of the 9-hydroxystearic acid portion or palmitic acid hydrocarbon chain resulted in lower molecular weight analogs that maintained anti-inflammatory activity or in one case enhanced activity.

Sequestration of 9-Hydroxystearic Acid in FAHFA (Fatty Acid Esters of Hydroxy Fatty Acids) as a Protective Mechanism for Colon Carcinoma Cells to Avoid Apoptotic Cell Death.

Hydroxy fatty acids are known to cause cell cycle arrest and apoptosis. The best studied of them, 9-hydroxystearic acid (9-HSA), induces apoptosis in cell lines by acting through mechanisms involving different targets. Using mass spectrometry-based lipidomic approaches, we show in this study that 9-HSA levels in human colorectal tumors are diminished when compared with normal adjacent tissue. Since this decrease could be compatible with an escape mechanism of tumors from 9-HSA-induced apoptosis, we investigated different features of the utilization of this hydroxyfatty acid in colon. We show that in colorectal tumors and related cell lines such as HT-29 and HCT-116, 9-HSA is the only hydroxyfatty acid constituent of branched fatty acid esters of hydroxyfatty acids (FAHFA), a novel family of lipids with anti-inflammatory properties. Importantly, FAHFA levels in tumors are elevated compared with normal tissue and, unlike 9-HSA, they do not induce apoptosis of colorectal cell lines over a wide range of concentrations. Further, the addition of 9-HSA to colon cancer cell lines augments the synthesis of different FAHFA before the cells commit to apoptosis, suggesting that FAHFA formation may function as a buffer system that sequesters the hydroxyacid into an inactive form, thereby restricting apoptosis.

De novo Lipogenesis in Adipocytes Results in the Production of Structurally Novel Signaling Lipids with Beneficial Metabolic and Anti‐inflammatory Effects

Increased de novo lipogenesis in adipose tissue is associated with enhanced insulin sensitivity in humans and rodents. Mice overexpressing the Glut4 glucose transporter in adipose tissue have elevated adipose tissue lipogenesis and increased glucose tolerance in spite of obesity and elevated circulating fatty acids. Untargeted lipidomic analysis of adipose tissue from these mice revealed marked elevation of a structurally novel class of lipids, branched Fatty Acid esters of Hydroxy Fatty Acids (FAHFAs). More than 25 FAHFA family members, which are distinguished by different acyl chain composition, have been identified in mammalian tissues and plants. Levels of some family members such as Palmitic Acid esters of Hydroxy Stearic Acids (PAHSAs) are highly regulated in altered nutritional and metabolic states. PAHSA levels are reduced in serum and subcutaneous adipose tissue of insulin‐resistant humans and serum PAHSA levels correlate very strongly with insulin sensitivity in humans. Chronic PAHSA treatment improves glucose tolerance and insulin sensitivity in insulin‐resistant mice. PAHSAs also have anti‐inflammatory effects in states such as obesity, colitis and auto‐immune Type 1 diabetes in mice. Many effects of PAHSA are mediated through G‐protein coupled receptors. Thus, PAHSAs exert multiple beneficial metabolic and anti‐inflammatory effects in diabetes and immune‐mediated diseases. The pathways that regulate these lipids provide new opportunities for disease prevention and treatment.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Asymmetric Synthesis of Saturated Hydroxy Fatty Acids and Fatty Acid Esters of Hydroxy Fatty Acids

The recent discovery of a novel class of endogenous lipids, named Fatty Acid esters of Hydroxy Fatty Acids (FAHFAs), that present antidiabetic and anti‐inflammatory activities, has attracted the interest on hydroxy fatty acids (HFAs) and their derivatives. We present herein the development of a convenient and general methodology for the asymmetric synthesis of HFAs and FAHFAs. The enantioselective organocatalytic synthesis of asymmetric terminal epoxides starting from monoprotected α,ω‐diols and their subsequent ring opening by a Grignard reagent are the key‐steps for our methodology, allowing the introduction of the hydroxy group at different positions of the long chain by proper selection of the starting diol and the Grignard reagent. MacMillan's third generation imidazolidinone organocatalyst has been employed for the epoxide formation, ensuring products in high enantiomeric purity. Furthermore, an approach for the synthesis of deuterated HFAs and FAHFAs was developed, leading to deuterated derivatives, which can be useful in biological studies and in mass spectrometry studies as internal standards.

Moose and Caribou as Novel Sources of Functional Lipids: Fatty Acid Esters of Hydroxy Fatty Acids, Diglycerides and Monoacetyldiglycerides.

Fatty acid esters of hydroxy fatty acids (FAHFA), diglycerides (DG) and monoacetyldiglycerides (MAcDG) are gaining interest as functional lipids in pharmaceuticals and functional food formulations for managing and treating metabolic or inflammatory diseases. Herein, we investigated whether the antler and/or meat of two Cervids (moose and caribou) are novel sources of FAHFA, DG and MAcDG. We observed FAHFA present in moose and caribou composed mainly of polyunsaturated families, and that the esterification occurred frequently at the C5-hydroxy fatty acid moiety, most noticeably arachidonic acid 5-hydroxyeicosatrienoic acid (ARA-5-HERA). Moose antler, caribou and moose meat also contained significant levels of both 1,2-DG and 1,3-DG lipids. The 1,3-DG molecular species consisted mainly of 16:0/18:1, 18:0/16:0, and 18:0/18:1. On the other hand, major 1,2-DG species consisted of DG 18:0/18:0, 16:0/16:0 and 18:1/18:1 molecular species with higher levels in the antler compared to the meat. The molecular species composition of MAcDG was very simple and consisted of 14:2/18:2/2:0, 16:0/18:2/2:0, 16:0/18:1/2:0 and 18:0/18:1/2:0 with the first species 14:2/18:2/2:0 predominating in the tip of moose antlers. Increasing access to and knowledge of the presence of these functional lipids in foods will enhance their intake in the diet with potential implications in improving personal and population health.

Loss of Transcriptional Repression by BCL6 Confers Insulin Sensitivity in the Setting of Obesity

SUMMARYAccumulation of visceral adiposity is directly linked to the morbidity of obesity, while subcutaneous body fat is considered more benign. We have identified an unexpected role for B cell lymphoma 6 (BCL6), a critical regulator of immunity, in the developmental expansion of subcutaneous adipose tissue. In adipocyte-specific knockout mice (Bcl6AKO), we found that Bcl6 deletion results in strikingly increased inguinal, but not perigonadal, adipocyte size and tissue mass in addition to marked insulin sensitivity. Genome-wide RNA expression and DNA binding analyses revealed that BCL6 controls gene networks involved in cell growth and fatty acid biosynthesis. Using deuterium label incorporation and comprehensive adipokine and lipid profiling, we discovered that ablation of adipocyte Bcl6 enhances subcutaneous adipocyte lipogenesis, increases levels of adiponectin and fatty acid esters of hydroxy fatty acids (FAHFAs), and prevents steatosis. Thus, our studies identify BCL6 as a negative regulator of subcutaneous adipose tissue expansion and metabolic health.