What is arachidonic acid

Arachidonic acid is a 20 carbon chain polyunsaturated omega-6 essential fatty acid (PUFA) that is found in animal and human fat as well as in the liver, brain, and glandular organs. Arachidonic acid is present in all mammalian cells, typically esterified to membrane phospholipids (especially phosphatidylethanolamine, phosphatidylcholine, and phosphatidylinositides) and is one of the most abundant polyunsaturated fatty acids present in human tissue ¹. Arachidonic acid is formed by the synthesis from dietary linoleic acid (omega-6 fatty acid) ² and is a precursor in the biosynthesis of prostaglandins, thromboxanes, and leukotrienes, collectively known as eicosanoids ³. Arachidonic acid is a building block for molecules that can promote inflammation, blood clotting, and the constriction of blood vessels. But the body also converts arachidonic acid into molecules that calm inflammation and fight blood clots. It turns out that the body converts very little linolenic acid into arachidonic acid, even when linolenic acid is abundant in the diet. The latest nutrition guidelines call for consuming unsaturated fats like omega-6 fats in place of saturated fat. The American Heart Association along with the Institute of Medicine, recommends getting 5% to 10% of your daily calories from omega-6 fatty acids ⁴. For someone who usually takes in 2,000 calories a day, that translates into 11 to 22 grams. A salad dressing made with one tablespoon of safflower oil gives you 9 grams of omega-6 fatty acids; one ounce of sunflower seeds, 9 grams; one ounce of walnuts, 11 grams. Most Americans eat more omega-6 fats than omega-3 fatty acids, on average about 10 times more. A low intake of omega-3 fats is not good for cardiovascular health, so bringing the two into better balance is a good idea ⁴. But don’t do this by cutting back on healthy omega-6 fats. Instead, add some extra omega-3s. Nutritional scientists suggest the omega-6 fats to omega-3 fats ratio of 2:1 to 4:1, which indicates a high consumption of seafood ⁵. For optimal infant nutrition, the ratio of omega-6 fatty acids to omega-3 fatty acids must be not higher than 10 ⁶. High consumption of plant oils rich in omega-6 polyunsaturated fatty acid (PUFA) and consumption of relatively low marine foods (as source of omega-3 fatty acid) increases the omega-6 fats to omega-3 fats ratio ⁷. According to dietary recommendations of the American Heart Association Nutrition Committee ⁸, it is suggested to consume fish at least two times per week or omega-3 polyunsaturated fatty acid (PUFA) 500 mg/day to prevent and reduce the risk of heart disease. The European Food Safety Authority panel recommends taking 250 mg omega-3 fatty acids per day, in contrast to the Suggested Dietary Targets in Australia recommends for adult men 610 mg omega-3 fatty acids and women 430 mg omega-3 fatty acids per day (docosahexaenoic fatty acid [DHA] + eicosapentaenoic fatty acid [EPA]) necessary to reduce the risk of cardiovascular disease ⁹, ¹⁰. Infant formulae should contain at least 0.2% of total fatty acids as docosahexaenoic fatty acid (DHA) and 0.35% as arachidonic acid ¹¹.

Arachidonic acid chemical formula is C20H32O2, 20:4(omega-6), where 20:4 refers to its 20 carbon atom chain with four double bonds, and (omega-6) refers to the position of the first double bond from the last, omega carbon atom ¹². Arachidonic acid has an average mass of 304.467 g/mol and usually assumes a hairpin structure (Figure 1). Due to the presence of its four double bonds in the cis position (which means that all hydrogen atoms are on the same side of the double bonds), the compound has a certain degree of flexibility for interaction with proteins ¹³. Even at low temperature it helps in keeping the fluidity of cell membranes. The four double bonds also enable interaction with molecular oxygen giving rise to bioactive oxygenated molecules including eicosanoids (prostanoids and leukotrienes) and isoprostanes via enzymatic and non-enzymatic mechanisms, respectively ¹⁴.

Arachidonic acid is naturally found incorporated in the structural phospholipids of the cell membrane in the body or stored within lipid bodies in immune cells ¹⁵. Arachidonic acid is particularly abundant in skeletal muscle, brain, liver, spleen and retina phospholipids ¹⁶. The concentration of free arachidonic acid in the circulation is very low, owing the fact that in human plasma, albumin is highly abundant as its concentration reaches up to 35 mg/ml, which enables the binding of free fatty acids keeping their concentration below 0.1 µmol ¹⁷.

Arachidonic acid supplementation does not result in decreased docosahexaenoic fatty acid [DHA]/eicosapentaenoic fatty acid [EPA] ¹⁸. DHA/EPA composition was unchanged by 700 mg ¹⁹ or 838 mg ²⁰ of arachidonic acid per day. In the same manner, 240 and 720 mg ²¹ or 1500 mg ²² of arachidonic acid per day did not change DHA/EPA composition. In contrast, it is well known that arachidonic acid composition is decreased by DHA/EPA supplementation ²³. Interestingly, it is commonly observed that arachidonic acid supplementation results in large decreases in linoleic acid composition ²¹. It appears that the capacity for exchange or retention in the body is in the following order DHA/EPA > arachidonic acid > linoleic acid. Arachidonic acid intake from foods or supplementation is thought to have a great impact on long-chain polyunsaturated fatty acids metabolism. The continued accumulation of evidence from large and well-designed dietary surveys and clinical trials is expected to confirm this ¹⁸.

In humans, arachidonic acid plasma levels typically range from 0.1 µM to 50 µM, whereas under pathophysiological conditions, arachidonic acid plasma concentrations up to 500 µM can be found ²⁴. Interestingly, 50 µM to 100 µM of free arachidonic acid are cytotoxic in vitro, thus it is likely that plasma arachidonic acid concentrations found in the human body exert cytotoxic and apoptosis regulatory functions. Accordingly, arachidonic acid is essential in embryogenesis and infant development, but may also be a driving factor in cardiovascular, metabolic and inflammatory diseases ²⁵. In line with this, several studies have found an inverse relationship of plasma arachidonic acid concentrations and the risk of coronary heart disease ²⁶. Thus, it is important to elucidate any potential influence of arachidonic acid on various aspects in human health.

Endogenous arachidonic acid generation mainly occurs via the release of arachidonic acid from cell membrane phospholipids ²⁷. This process is catalyzed by enzymes of the phospholipase A2 (PLA2) superfamily and is induced by various cellular activation signals, including inflammation or infection driven tumor necrosis factor receptor (TNFR) and toll-like receptor 4 (TLR4) stimulation ²⁸. Among the phospholipase A2 (PLA2) enzymes superfamily, three members contribute to eicosanoid production and are involved in distinct functions of eicosanoid metabolism. The cytosolic Ca2+ dependent PLA2 (cPLA2) alpha mainly drives free fatty acids (FFAs) production and generation of arachidonic acid, which is involved in cellular signaling. The cytosolic Ca2+ independent PLA2 (iPLA2) alpha contributes to cellular homeostasis via the synthesis of specialized pro-resolving mediators and reacylation of free arachidonic acid, and the secretory PLA2 (sPLA2) controls free arachidonic acid release and induces the local inflammatory response in a paracrine manner ²⁹. In addition to PLA2, two other phospholipase families, namely phospholipase C (PLC) and phospholipase D (PLD), generate arachidonic acid via intermediate products such as diacylglycerol (DAG) ³⁰. Finally, endocannabinoids serve as an endogenous source for arachidonic acid, as demonstrated by the generation of arachidonic acid from anandamide ³¹.

Arachidonic acid metabolites are implicated in immune surveillance, the establishment of allergy, the initiation and resolution of an inflammatory response, platelet aggregation and the coagulation cascade, glucose metabolism, neuronal signaling, female fertility, and the adaptation of mood and appetite ¹². Arachidonic acid metabolites are important factors in the initiation and resolution of inflammation and have been linked to the pathophysiology of obesity, diabetes mellitus, nonalcoholic fatty liver disease (NAFLD), nonalcoholic steatohepatitis (NASH) and cardiovascular diseases ²⁷. Interestingly, available enzyme cassettes to process arachidonic acid and G-protein coupled receptors are different for each cell and tissue type, ensuring a cell/tissue-specific regulation and function of arachidonic acid metabolism ³². The functional state of cells (e.g., activated vs. non-activated macrophages) and tissue environment impact on the synthesis of arachidonic acid derivatives, and the necessity for cell-cell interactions to produce specific eicosanoids, also termed transcellular biosynthesis, adds further complexity to arachidonic acid metabolism ³³. Targeted lipidomic strategies using mass spectrometry-based technology platforms revealed that the compound effect of local lipid metabolites, and their time-dependent generation, determine the biological function of the arachidonic acid metabolome, rather than the specific concentration of a single eicosanoid at a given site and time point ³⁴. This was also demonstrated by the partial inhibition of the immunomodulatory effects of free fatty acids by omega-3 PUFAs, such as eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), which display anti-inflammatory properties and are precursors for pro-resolving specialized pro-resolving mediators ³⁵. Specialized pro-resolving mediators, including resolvin D1, enhance phagosome formation in macrophages and are capable of reverssing the phagocytic defect of macrophages derived from diabetic mice. They may thereby counter-balance the effects of pro-inflammatory arachidonic acid metabolites ³⁶.

Lastly the plethora of arachidonic acid derivatives and their multitudinous implications, and the differential expression and functionality of arachidonic acid metabolites in various cells and tissues, as well as their transcellular biosynthesis, render the arachidonic acid metabolism one of the most complex regulatory systems within the human body. It is thus not surprising that many aspects of the arachidonic acid metabolome still need further investigation, despite extensive scientific progress in the field ²⁷.

Figure 1. Arachidonic acid chemical structure (arachidonic acid hairpin conformation)

Footnote: Arachidonic acid is incorporated in phospholipids in the cells’ cytosol, adjacent to the endoplasmic reticulum membrane that is studded with the proteins necessary for phospholipid synthesis and their allocation to the diverse biological membranes ³⁷. Of note, glycerophospholipids are composed of a glycerol backbone esterified to two hydrophobic fatty acids tails at sn- (stereospecifically numbered) 1 and 2 position and a hydrophilic head-group at sn 3 ³⁸.

Figure 2. Formation of arachidonic acid from linoleic acid metabolism

[Source ¹² ]

Arachidonic acid foods

Arachidonic acid can be provided to humans by an exogenous source supplied either by the direct consumption of dietary food that contains high level of arachidonic acid, whole eggs, poultry, animal organs, salmon, tuna ³⁹, ⁴⁰, a wide range of lean meat ⁴¹ and visible meat fats ⁴² or through the parent molecule, linoleic acid (omega-6 fatty acid) (see Figure 2 above). Linoleic acid is considered to be an essential fatty acid since humans and some mammals lack the enzymes required for its synthesis ⁴³. Linoleic acid is abundant in vegetable oils such as soya, corn, sunflower and safflower and also found in walnuts ⁴⁴. In human body, linoleic acid is subjected to series of desaturation enzymes (delta-6 fatty acid desaturase and delta-5 fatty acid desaturase), and elongation enzymes that carry out their action in the endoplasmic reticulum (ER) membrane ⁴⁵. Beside the exogenous source, endocannabinoids such as N-arachidonoyl ethanolamine (anandamide) serve as an endogenous source of arachidonic acid ¹². An integrated membrane protein enzyme, fatty acid amide hydrolase, is responsible for the catalysis of anandamide into arachidonic acid and ethanolamine to eliminate the anandamide signal in the nervous system ⁴⁶.

Another arachidonic acid source, that is important for herbivores and vegetarians, is linoleic acid, also an omega-6 PUFA that contains only two cis- double bonds (18:2ω-6). Linoleic acid is an essential fatty acid for animals because they cannot synthesize it, in contrast to plants, which can synthesize it from oleic acid ³⁸. Linoleic acid is abundant in many nuts, walnuts, fatty seeds and their derived vegetable oils ⁷. Linoleic acid is converted in animals cells cytosol to arachidonic acid, docosatetraenoic acid (22:4ω-6) and other fatty acids by step-wise desaturation and chain elongation (see Figure 2). Linoleic acid conversion to arachidonic acid is, however, low. Linoleic acid is readily oxidized by delta 6-desaturase to gamma-linolenic acid (18:3-n6), but several factors such as aging, nutrition, smoking impair the activity of the enzyme ³⁸. Gamma linolenic elongation step to dihomo-gamma-linolenic acid (20:3-n6) is rapid; yet, it is oxidized by delta-5 desaturase to yield arachidonic acid at a small percentage because delta-5 desaturase prefers the n-3 to n-6 fatty acids ⁴⁷.

Table 1. Content of arachidonic acid and the other fatty acids per 100 g edible portion of animal foods

Food group	Total fat (mg)	Fatty acids (mg)
		Palmitic acid	Oleic acid	Linoleic acid	Arachidonic acid	Eicosapentaenoic acid (EPA)	Docosahexaenoic acid (DHA)
Meats and poultry
Pork, loin, whole, lean and fat, raw	12580	2720	5140	1110	80	0	0
Pork, medium type breed, loin, lean and fat, raw	22600	5600	9100	1900	68	0	12
Chicken, broiler, thigh, meat and skin, raw	16610	3511	5832	3096	104	4	7
Chicken, broiler, thigh, meat with skin, raw	14200	3300	5800	1600	79	1	7
Beef, hip, inside (top) round steak, boneless, lean, raw	2210	520	910	120	40	0	0
Beef, inside round, lean, raw	4300	890	1500	120	24	4	1
Eggs
Chicken, whole, fresh or frozen, raw	10010	2218	3810	1109	156	2	72
Hen, whole, raw	10300	2100	3500	1300	150	0	120
Fishes and seafoods
Salmon, pink (humpback), raw	6700	1044	1108	102	127	547	859
Pink salmon, raw	6600	790	920	81	31	400	690
Flatfish (flounder or sole or plaice), raw	1930	282	358	45	30	137	108
Righteye flounder, brown sole, raw	1300	150	140	10	50	100	72
Sardine, pacific, canned in tomato sauce, drained with bones	10450	1738	1851	123	190	532	864
Sardine, Japanese pilchard, canned products, in tomato sauce	10800	1900	1200	140	160	1300	1100
Milk and dairy products
Cheese, cream	34240	8497	7923	1032	50	0	0
Cheese, cream	33000	8700	6400	570	38	20	6

[Source ¹⁸ ]

Arachidonic acid function

Arachidonic acid is an integral constituent of biological cell membrane phospholipids, conferring it with fluidity and flexibility, so necessary for the function of all cells, especially in nervous system, skeletal muscle, and immune system ³⁸. The four double bonds of arachidonic acid predispose it to oxygenation that leads to a plethora of metabolites of considerable importance for the proper function of the immune system, promotion of allergies and inflammation, resolving of inflammation, mood, and appetite ¹². Platelets, mononuclear cells, neutrophils, liver, brain and muscle have up to 25% phospholipid fatty acids as arachidonic acid ⁴⁸. Arachidonic acid participates in the Lands cycle, a membrane phospholipids’ reacylation/deacylation cycle, which serves to keep the concentration of free arachidonic acid in cells at a very low level ⁴⁹. Since arachidonic acid is a fundamental constituent of cell structure, it will particularly be needed for during development and growth and upon severe or widespread cell damage and injury ³⁸.

Cell membrane fluidity

Arachidonic acid four cis double bonds endow it with mobility and flexibility conferring flexibility, fluidity and selective permeability to membranes ⁵⁰. Arachidonic acid control of membrane fluidity influences the function of specific membrane proteins involved in cellular signaling ⁵¹ and plays a fundamental role in maintenance of cell and organelle integrity and vascular permeability ⁵². These properties might explain arachidonic acid critical role in neuron function, brain synaptic plasticity, and long-term potentiation in the hippocampus ⁵³.

Ion channels

Non-esterified, free arachidonic acid affects neuronal excitability and synaptic transmission via acting on most voltage-gated ion (sodium [NaV], potassium [KV], calcium [CaV], and proton [HV]) channels, responsible for regulating the electric activity of excitable tissues, such as the brain, heart and muscles ³⁸. Ion channels are large families of integral membrane proteins that form a selective pore for ions to cross the lipid bilayer, via undergoing conformational changes in response to alteration in the cell transmembrane electrical potential. These channels gate passage of specific ions and thus control the propagation of nerve impulses, muscle contraction, and hormone secretion ⁵⁴. The homologous mon-, di- or tetrameric subunits of arachidonic acid-sensitive voltage-gated channels are composed of four transmembrane helices spanning the cell membrane lipid bilayer (S1-S4) making up the voltage-sensor domain, and/or 2 transmembrane segments constituting the central ion-conducting pore ⁵⁵. The gating charges are situated on helix S4, a positively charged voltage sensor, which responds to changes in voltage across the membrane by inducing movements of the helix relative to the remainder of the protein or the movement of the positive charges through the membrane toward the extracellular side ⁵⁵. Since S4 is in contact with the lipid bilayer, the arachidonic acid lipophilic, flexible acyl chain can position its carboxylate negative charge onto the voltage sensor, and modulate its activity, likely shifting the voltage dependency of activation via channel-activating electrostatic interactions ⁵⁴.

Free arachidonic acid evoked K+ channel opening in neurons of the rat visual cortex, thus suggesting the existence of an arachidonic acid-activated type of K+ channel, which may play a critical role in modulating cortical neuronal excitability ⁵⁶. Arachidonic acid was previously reported to directly activate K+ channels in gastric, pulmonary artery, and vascular smooth muscle cells, and cardiac atrial cells likely via interacting with the ion channel protein itself ⁵⁷. Conversely, arachidonic acid is known to suppress the Kv4 family of voltage-dependent K+ channels, in a direct, fast, potent, and partially reversible mode ⁵⁸. The activity of the large-conductance Ca2+- dependent K+ (BK) channels, which control diverse functions in the central nervous system such as sleep and neural regulation of the heart, is increased up to 4 folds by arachidonic acid, consequent to direct interaction with the channel protein ⁵⁹. Conversely, arachidonic acid inhibited intermediate conductance, Ca++-activated K+ channels, which play crucial roles in agonist-mediated transepithelial Cl− secretion across airway and intestinal epithelia, via interacting with the pore-lining amino acids (aa) threonine (aa 250) and valine (aa 275) ⁶⁰. Background, non-voltage-dependent two pore domain K(+) channels, which play an essential role in setting the neuronal membrane potential and potential duration are opened by arachidonic acid, and not its metabolites, provided the carboxyl end is not substituted with an alcohol or methyl ester ⁶¹. Additionally, arachidonic acid was reported to inhibit the ATP- sensitive K+ channel in cardiac myocytes almost completely, while activated the ATP-insensitive K+ channel ⁶².

Free, non-esterified arachidonic acid prevents ischemia-induced heart arrhythmia, a major cause of sudden cardiac death in humans, by modulating the activity of cardiac Na+ channels, the major class of ion channels that determine cardiac excitability, causing a reduction in the electrical excitability and/or automaticity of cardiac myocytes ⁶³. Sodium channels consist of a large functional subunit and a smaller subunit, which interacts with a regulatory segment. Arachidonic acid, but not its metabolic products, was shown to voltage-dependent modulate muscle Na+ channel currents, displaying both activation and inhibitory effects depending on the depolarizing potential ⁶⁴. Arachidonic acid also displayed both activation and inhibitory effects on different Cl- channels, widely distributed especially in epithelial tissues, and thus, mediate increase or block of Cl- ions permeation ⁶⁵. The double bonds and hydrophilic head were recently reported to be responsible for the arachidonic acid mediated dramatic increases in proton current amplitude through the voltage-gated proton (Hv) channel. The latter lacks a pore domain but allows passage of proton through the center of each voltage sensor domain ⁵⁴ and supports the rapid production of reactive oxygen species (ROS) in phagocytes through regulation of pH and membrane potential ⁶⁶.

Receptors and enzymes

Exogenous or endogenously produced arachidonic acid was discovered to greatly enhance the functional activity of ligand-gated ion channels, namely the gamma-amino butyric acid receptor (GABA-R) located on the neuronal membrane, via modulating the GABA-R interaction characteristics with its ligands ⁶⁷. Free arachidonic acid exposure essentially led to inhibiting the muscle and neuronal nicotinic acetylcholine receptor (nAChR), an integral membrane protein deeply embedded in the postsynaptic region, with two agonist binding sites and a central ion pore. The receptor inhibition resulted from arachidonic acid displacing lipids from their sites in the plasma membrane and direct acting as antagonist at the PUFA-protein interface ⁶⁷.

Activation of eosinophils, neutrophils, and macrophages elicits powerful respiratory burst associated with reduction of molecular oxygen to superoxide via activation of the NADPH oxidase complex, which consists of five proteins residing in resting cells in the cytosol or membrane of intracellular vesicles, and in activated cells are assembled on the cell membrane ⁶⁸. Generated reactive oxyegn species (ROS) induce membrane depolarization and cytoplasm pH decrease, thus restricting NADPH activity. Concentrations of arachidonic acid of 5–10 µM added to neutrophils enhanced NADPH oxidase stimulation due to arachidonic acid-mediated activation of the Hv channel, modulation of the membrane potential and pH, and efflux of the H+ ions generated together with the superoxide anion, O2− ⁶⁹.

PUFA, especially arachidonic acid, are documented activators of membrane-associated, magnesium-dependent, neutral sphingomyelinases (nSMase) ⁷⁰. Arachidonic acid was recently documented as activator of Schistosoma mansoni and Schistosoma haematobium tegument-associated neutral sphingomyelinase in a dose-dependent manner, eventually leading to their attrition in vitro and in vivo ⁷¹.

Cell death

Free arachidonic acid levels are kept very low in cells as uncontrolled accumulation of unesterified arachidonic acid decisively impaired cell survival via induction of apoptosis ⁴⁹. The apoptotic effect was attributed to free arachidonic acid and not its metabolites as it was recorded in the absence of lipoxygenase or cyclooxygenase enzyme activity, and was speculated to be associated with oxidative stress and/or changes in membrane fluidity ⁷². Pompeia et al. ⁷³ reported that the cytotoxicity of arachidonic acid is undeniable, but may well be one of its fundamental functions in vivo. arachidonic acid concentrations of 50–100 µM are cytotoxic to most cell lines in vitro. In the majority of models 1–10 µM arachidonic acid is necessary to elicit any biological response but some activities require 100–300 µM ⁵¹. This indicates that arachidonic acid apoptotic and physiological levels overlap and it is very possible that arachidonic acid cytotoxicity occurs in vivo because under some pathologic conditions, human plasma arachidonic acid levels can increase from 0.1–50 μM to 100 and up to 500 μM ⁷³.

Arachidonic acid metabolism

Four enzymatic pathways process free arachidonic acid, resulting in the generation of a plethora of arachidonic acid derivatives with multitudinous functions (Figures 3 and 4).

First, cyclooxygenase 1 (COX1) and cyclooxygenase-2 (COX-2), also known as Prostaglandin G/H synthases, foster the production of thromboxane A2 (TXA2), prostacyclin (PGI2) and several prostaglandins (PGs) ⁷⁴. Cyclooxygenase 1 (COX1) is constitutively expressed and found in all tissues, which are prone to lipopolysaccharide (LPS) induced inflammation. Thromboxane A2 (TXA2) and prostaglandins (PGs), such as prostaglandin D2 (PGD2), are metabolites of the COX1 pathway, and alter the vessel tone, mediate platelet aggregation and are implicated in immune surveillance ⁷⁵. Cyclooxygenase 2 (COX2) is found in macrophages and endothelial cells, demonstrates high expression levels in the kidney and brain, and is induced by bacterial endotoxins, growth factors, hormones, and several cytokines ⁷⁶. Its major metabolites prostaglandin E2 (PGE2), prostaglandin I2 (PGI2, aslo known as prostacyclin), prostaglandin D2 (PGD2) and prostaglandin F2alpha (PGF2α) adapt host immune response, vessel tone regulation, thrombus formation, pain reception, and female fertility, and are involved in neurodegeneration and cancer ⁷⁷. Additionally, cyclooxygenase 3 (COX3), a splice variant of COX1, facilitates prostaglandin synthesis in the human brain and heart ⁷⁸.

Second, four lipoxygenases (LOX), namely 5-LOX, 8-LOX, 12-LOX and 15-LOX, process arachidonic acid. 5-LOX produces 5-hydroperoxyicosatetraenoic acid (5-HPETE), 5-hydroxyicosatetraenoic acid (5-HETE), and 5-oxo-eicosatetraenoic acid (5-oxo-ETE), as well as various leukotrienes (LTs) ⁷⁹. These metabolites affect neutrophil recruitment and diapedesis, epithelial barrier function, vascular permeability and bronchoconstriction ⁸⁰. 8-LOX and 15-LOX lipoxygenases convert free arachidonic acid to 8- and 15-hydroperoxyicosatetraenoic acid (8-HPETE and 15-HPETE) and foster the production of 15-HPETE derivatives, such as 15-hydroxyicosatetraenoic acid (15-HETE), lipoxins and eoxins ⁸¹. 12-LOX metabolize arachidonic acid to 12-hydroperoxyeicosatetraenoic acid (12-HPETE), which itself serves as a precursor for 12-hydroxyeicosatetraenoic acid (12-HETE) and to hepoxilins ⁸². Arachidonic acid derivatives from the 8-, 15-, and 12-LOX pathways are involved in the establishment of hyperalgesia and induce the expression of fatty acid translocase ⁸³. Lipoxins exert mainly anti-inflammatory properties. Interestingly, their synthesis is induced by the COX inhibitor aspirin, an antithrombotic agent widely used in the treatment and for secondary prevention of cardiovascular diseases ⁸⁴.

Third, the cytochrome P450 (CYP450) pathway, which is mainly restricted to the liver and is well known for its function in detoxification, includes epoxygenase and ω-hydroxylase, which facilitate the production of epoxy-eicosatrienoic acids (EETs) and hydroxy-eicosatetraenoic acids (HETEs), respectively. These arachidonic acid derivatives regulate vessel constriction and dilation, hamper hyperalgesia, and inhibit COX2 expression ⁸⁵.

Fourth, anandamide, an endocannabinoid, is generated from arachidonic acid via the fatty acid amide hydrolase (FAAH) in a reversible reaction ⁸⁶, whereas the processing of anandamide to arachidonic acid and ethanolamine takes place when tissue damage is associated with high levels of free arachidonic acid ⁸⁷. In this situation, anandamide supports tissue regeneration, and cellular proliferation via interaction with cannabinoid type 1 (CB1) receptors ⁸⁸.

Finally, free arachidonic acid is also processed by non-enzymatic reactions. The four double bonds of arachidonic acid are readily oxygenated to form bioactive molecules. Thus, oxidative stress and/or exposure of arachidonic acid to reactive oxygen species (ROS) and reactive nitrogen species (RNS) result in oxidation of arachidonic acid and generate isoprostanes and nitroeicosatetraenoic acids ⁸⁹. These eicosanoids were reported to inhibit COX1, whereas isoprostanes have been linked to platelet aggregation, vasoconstriction, smooth muscle cell proliferation, and cardiomyocyte hypertrophy ⁹⁰. To conclude, arachidonic acid and its derivatives can enter numerous metabolic pathways that interconnect lipid metabolism with immunity. A synopsis of key enzymes, metabolites, and biological functions of the arachidonic acid metabolism is depicted in Figure 3.

The expression and activation of phospholipase A2 (PLA2) enzyme can be a response to a wide range of cellular activation signals from receptor dependent events requiring a G coupled transducing protein as Toll-like receptor 4 (TLR4), purinergic receptors and inflammation stimulation to calcium ionophores, melittin (bee venom) and tumor promoting agents [⁹¹. Three fates wait for the liberated, free functional arachidonic acid: 1) it may diffuse to other cells, 2) reincorporated into the phospholipids, or 3) metabolized.

Furthermore, the activation of PLA2 enzyme can be through the binding of tumor necrosis factor alpha (TNF-α) to its receptor, P75 and P55, inducing the release of arachidonic acid from phosphatidylcholine and phosphatidylethanolamine. Free arachidonic acid can have an important role in cell apoptosis as its accumulation that occurs as a result of arachidonyl CoA transferase inhibition, can promote the activation of sphingomyelinase, enzymes that trigger the degradation of sphingolipids (known to play an important role in cell regulation and cell cycle) to phosphocholine and ceramide ⁹².

Free arachidonic acid can be metabolized via enzymatic reactions. Free arachidonic acid can undergo four possible enzymatic pathways: Cyclooxygenase, Lipoxygenase, Cytochrome p450 (CYP 450) and Anandamide pathways to create bioactive oxygenated PUFA containing 20 carbon (eicosanoids) acting as local hormones and other compounds acting as signaling molecules ¹². Enzymes involved in the cyclooxygenase pathway are cyclooxygenase-1 (COX-1) and cyclooxygenase-2 (COX-2) also called prostaglandin H synthase, along with downstream enzymes that mediate the production of prostaglandins (PGH2, an unstable intermediate, PGE2, PGD2 and PGF2alpha, prostacyclins (PGI2), and thromboxanes (TXA2, TXB2) ¹². Lipoxygenase pathway consists of LOX-5, LOX-8, LOX-12, and LOX-15 enzymes and their products, leukotrienes (LTA4, an unstable intermediate, LTB4, LTC4, LTD4 and LTE4), lipoxins (LXA4 and LXB4 formed upon LXA4 degradation) and 8–12- 15- hydroperoxyeicosatetraenoic acid (HPETE) ¹². The CYP 450 pathway involves two enzymes, CYP450 epoxygenase and CYP450 ω-hydroxylase giving rise to epoxyeicosatrienoic acid (EETs) and 20-hydroxyeicosatetraenoic acid (20-HETE) respectively. Anandamide pathway comprises the fatty acid amide hydrolase (FAAH) to produce the endocannabinoid, anandamide ⁹³.

Arachidonic acid may additionally undergo non-enzymatic reactions. Studies proved that the exposure of carbon tetrachloride (CCL4) to rats to mimic the oxidative stress and as an induction of lipid peroxidation state in vivo, leads to the formation of PGF2-like compounds called isoprostanes and other compounds such as nitroeicosatetraenoic acids. Arachidonic acid autoxidation by reactive oxygen species (ROS) and reactive nitrogen species (RNS) are also examples of non-enzymatic oxidative metabolism ⁹⁴.

Arachidonic acid metabolism and enzymes expression usually vary from cell to cell and from tissue to another according to various factors; consequently, the level and type of biosynthesized eicosanoids will differ in each case. It was reported that bone marrow macrophages differ from peritoneal macrophage responses regarding generated eicosanoids quantities and specificity. One more factor that causes this variation is the state of the cell whether it was stimulated or in resting phase. In normal cell state, eicosanoids are generated in very minute amounts and subsequent up regulation can only occur following an inflammatory stimulation ⁹⁵.

The complexity of eicosanoid biosynthesis lies in the cell–cell interaction, where a donor cell has to transfer its unstable intermediate e.g. PGH2, LTA4 to another recipient cell to trigger the latter for eicosanoids biosynthesis. The single donor cell should have all the necessary enzymes to produce eicosanoids while the recipient cell has not to have all the required enzymes for arachidonic acid release. Hence, for initiation inflammation or tissue injury, at least two cells in the injured tissue must have the complete enzyme cassette to initiate eicosanoids production. Accordingly, eicosanoids are described, as mentioned above, as local hormones due to their autocrine and paracrine action. Adding to the complexity of the trans cellular interactions, the arachidonic acid intermediate metabolites are lipophilic with short half-life time (90–100 s) and require some other mechanisms to be translocated ⁹⁶. These facts are in line with studies since 1985 by Dahinden et al. ⁹⁷ who revealed that exogenous LTA4 stabilized by albumin is uptaken by bone-marrow mast cells to produce sulfidopeptide LTC4. Later on, it was found by Dickinson et al. ⁹⁸ that a group of special proteins called fatty acid binding proteins (FABP), specific for each cell type, are responsible for increase of arachidonic acid intermediates export via their stabilization and lengthening their half-life time by up to 30 minutes at 4 °C.

Though eicosanoids have a short half-life time, they contribute to many biological activities in paracrine or autocrine manner. They have crucial dual role, regulating innate immunity and inflammation resolution through binding to G protein coupled receptors located on other cells. First, once tissues are inflamed or infected, beside pain and swelling, arachidonic acid metabolites amplify the inflammatory signals to recruit leukocytes, pro-inflammatory cytokines, and immune cells to help in pathogens resistance and clearance. Second, they balance the induced inflammatory signals by producing resolving metabolites to act as host protection, since inflammation exerts a threatening action if it is not controlled in an appropriate time manner ⁹¹. COX-1 induced eicosanoids were shown to be lethal when produced continuously. Von Moltke et al. ⁹⁹ was able to prove that eicosanoids can be lethal, as COX-1-mediated prostaglandins generation were shown to be responsible for vascular leakage and mortality in mice model. Another example of homeostasis regulation, COX-1-mediated TXA2 production regulates platelets aggregation while COX-2-mediated PGI2 release inhibits platelets aggregation and promotes vasodilation. Imbalance between levels of PGI2 and TXA2 could elevate the risk of cardiovascular diseases. On these terms, physicians prescribe aspirin as a cardio protectant to inhibit COX-1 enzyme and subsequent inhibition of TXA2 ⁹¹.

PLA2 is up-regulated once triggered by same receptor that leads to inflammasome activation, prompting the class switching of eicosanoids from prostaglandins to lipoxins to reprogram cells from the pro-inflammation and inflammasome activation to resolution ¹⁰⁰. This mechanism can be enhanced by acetylated COX-2 and FLAP (5-lipoxygenase-activating protein, is a constitutively expressed protein that serves the docking of arachidonic acid with 5-LOX active site as well as the co-localization of both 5-LOX – cPLA2 to perinuclear membrane) to produce lipoxins ¹⁰¹.

Figure 3. Arachidonic acid metabolism and their biological functions in metabolic and cardiovascular diseases

Footnote: Metabolites and enzymes involved in arachidonic acid metabolism and their biological functions in metabolic and cardiovascular diseases. Endogenous arachidonic acid is mainly derived from cell membrane phospholipids, which are processed by phospholipase A2 (PLA2), phospholipase C (PLC), and phospholipase D (PLD). Free arachidonic acid serves as a precursor for a plethora of metabolites, including prostaglandins (PGs), prostacyclin, thromboxane, HPETE, leukotrienes, lipoxins, hypoxins, anandamide, and epoxyeicosatrienoic acids (EETs). In addition to this enzymatic processing of arachidonic acid, there is also a non-enzymatic metabolization. The latter is important for the production of isoprostanes and nitroeicosatetraenoic acid. Many arachidonic acid metabolites are highly bioactive and involved in various crucial vital processes. Relevant biological functions in metabolic and cardiovascular diseases are summarized in the blue boxes at the bottom.

Abbreviations: COX = cyclooxygenase; CYP = cytochrome; ROS = Reactive oxygen species; RNS = Reactive nitrogen species; FAAH = fatty acid amide hydrolase; HX = hypoxin; LOX = lipoxygenase; LT = leukotriene; and LX = lipoxin.

[Source ²⁷ ]

Figure 4. Arachidonic acid metabolism

Footnote: Sites of hydrolysis for each phospholipase [phospholipase A1 (PLA1), phospholipase A2 (PLA2), phospholipase C (PLC) and phospholipase D (PLD)].

[Source ¹² ]

Arachidonic acid benefits

Resolution of inflammation

Free arachidonic acid and its metabolites, prostaglandins (PG), namely prostaglandin F2alpha (PGF2a, also known as dinoprost), prostaglandin E2 (PGE2, also known as dinoprostone), and prostaglandin I2 (PGI2, also known as prostacyclin) display essential roles in skeletal muscle development and growth by controlling proliferation, differentiation, migration, fusion and survival of myoblasts ¹⁰². Eicosanoids (prostanoids and leukotrienes) produced from arachidonic acid tend to promote muscle growth during and after physical activity in healthy humans ³⁸. Yet, the major action of arachidonic acid metabolites is promotion of acute inflammatory response, characterized by the production of pro-inflammatory mediators such as prostaglandin E2 (PGE2) and prostaglandin I2 (PGI2), followed by a second phase in which lipid mediators with pro-resolution activities may be generated. Resolution of inflammation is no more considered a passive process, but rather an active programmed response regulated by mediators with pro-resolving capacity, prominent among which is arachidonic acid-derived lipoxin A4 ¹⁰³. Lipoxin A4 stimulates cessation of neutrophil infiltration, enhances macrophage uptake of apoptotic cells in pre-clinical animal models ¹⁰⁴, attenuates leukotriene C4-induced bronchoconstriction in asthmatic subjects, decreases eczema severity and duration and improves patients’ quality of life via inhibiting the activity of innate lymphoid cells type 2 ¹⁰⁵.

Lipoxin A4 (1 nM) was also reported to attenuate adipose inflammation, decreasing interleukin (IL)- 6 and increasing IL-10 expression in aged mice ¹⁰⁶. Recently, lipoxin A4 encapsulated in poly-lactic-co-glycolic acid microparticles displayed considerable healing effects in topical treatment of dorsal rat skin lesions, provided interaction with its specific receptor on skin cells ¹⁰⁴. Other arachidonic acid metabolites, notably PGE2, PGI2 and leukotriene B4 and leukotriene D4 readily promote wound healing via regulating the production of angiogenic factors and endothelial cell functions ¹⁰⁷ and inducing stem cells’ proliferation and angiogenic potential ¹⁰⁸. Furthermore, lipoxin A4 was reported to have anti-diabetic potential via inhibiting IL-6, tumor necrosis factor and ROS generation ¹⁰⁹.

Newborns health

Polyunsaturated fatty acids (PUFAs), especially arachidonic acid, affect the function of numerous ion channels, the activity of various enzymes and are implicated in cell apoptosis, necrosis and death, events of critical importance during embryogenesis, thereby have significant physiological and pharmacological impact on the health of newborns ⁵⁴. Arachidonic acid and docosahexaenoic acid (DHA, 22:6omega3) are important components of human milk but are lacking in cow milk and most commercial infant formula in developing countries ¹¹⁰. Due to its importance in development especially of the central nervous system and retina ¹¹⁰, the Food and Agricultural Organization (FAO)/World Health Organization (WHO) recommended that infant formula, unless specifically added, should be supplemented with arachidonic acid ¹¹¹. Decreased postnatal arachidonic acid and docosahexaenoic fatty acid (DHA) blood levels in premature infants were found to be associated with neonatal morbidities, while adding DHA and arachidonic acid to preterm-infant formulas led to improved visual acuity, visual attention and cognitive development ¹¹⁰. The arachidonic acid levels in human milk and arachidonic acid requirements, essentiality in pre- and neonatal life and during development, and inclusion in infant formulas have recently been reviewed ¹¹², challenged and discussed ¹¹³.

Neurological disorders

Arachidonic acid does not only influence cell membrane fluidity and the activity of ion channels, especially in the brain, it constitutes together with docosahexaenoic fatty acid (DHA) 20% of the human brain dry weight, concentrated in the neurons outer membrane and in the myelin sheath ¹¹⁴. Additionally, positron emission tomography was used to show that the brain of human healthy volunteers consumes arachidonic acid at a rate of 17.8 mg/d ¹¹⁵. Accordingly, arachidonic acid was recommended for management of central nervous system, visual and auditory damage in preterm infants via supporting neurovascular membrane integrity ¹¹⁶. Children with autism had lower levels of blood PUFA, especially arachidonic acid, than normal children ¹¹⁷ and showed notable improvement after dietary PUFA intake ¹¹⁸. In the elderly too, arachidonic acid supplementation improved cognitive functions ⁵³, perhaps via increasing the proliferation of neural stem/progenitor cells or newborn neurons and general hippocampal neurogenesis ¹¹⁹. The charged arachidonic acid displayed beneficial effects on epileptic seizures and cardiac arrhythmia by electrostatically affecting the kV channel’s voltage sensor, thus regulating neuronal excitability ¹²⁰.

Bodybuilding

In skeletal muscles, arachidonic acid has been found to make up to 15–17% of total fatty acids, thus explaining why arachidonic acid supplementation affected body composition, muscle function and power output in strength-training individuals ¹²¹. It is also possible that arachidonic acid modulates neuromuscular signaling through its incorporation into cell membranes, and/or increases neurotransmitter firing from nerve cells ¹²².

A 2007 clinical study examining the effects of 1,000 mg/day of arachidonic acid for 50 days found supplementation to enhance anaerobic capacity and performance in exercising men ¹²³. Arachidonic acid supplementation during resistance-training may enhance anaerobic capacity and lessen the inflammatory response to training. However, arachidonic acid supplementation did not promote statistically greater gains in strength, muscle mass, or influence markers of muscle hypertrophy ¹²³.

Schistosomicidal action

The first evidence relating PUFA to schistosomes came from the ability of corn oil to exposed to surface membrane antigens of Schistosoma mansoni lung-stage larvae to specific antibody binding, thus allowing serologic visualization ¹²⁴. Further studies indicated that among PUFA, arachidonic acid (10 µM for 30 minutes) was the most effective in allowing specific antibody binding to otherwise hidden surface membrane antigens of Schistosoma mansoni and Schistosoma haematobium lung-stage schistosomula ¹²⁵. Of importance, exposure to 20 µM arachidonic acid for 30 minutes elicited surface membrane disintegration and attrition of the schistosomula ¹²⁵. Studies aiming at clarifying these observations led to identification of surface membrane sphingomyelin instrumental role in schistosome immune evasion. Controlled sphingomyelin hydrolysis by parasite tegument-associated neutral sphingomyelinase (nSMase) allows entry of nutrients but not host molecules >600 Da or antibodies. Excessive nSMase activation and consequent SM hydrolysis elicits exposure of surface membrane antigens and eventual larval death. arachidonic acid is a major nSMase activator. Accordingly, it was straightforward to predict that arachidonic acid possesses potentially potent schistosomicidal activity ¹²⁶.

All adult worms of Schistosoma mansoni and Schistosoma haematobium exposed to 2.5 mM arachidonic acid in the presence of 100% fetal calf serum showed extensive damage, disorganization, and degeneration of the tegument and the subtegumental musculature followed by death of all worms within 5 hours ¹²⁷. Pure arachidonic acid and different arachidonic acid formulations elicited notable, reproducible, and safe schistosomicidal activity against larval, juvenile and adult Schistosoma mansoni and Schistosoma haematobium infection of inbred and outbred mice and hamsters ¹²⁸. The ability of arachidonic acid to control infection with Schistosoma mansoni was demonstrated in Egyptian children. The chemotherapeutic activity of arachidonic acid and praziquantel was equally high in low infection settings and equally low in children with heavy infection living in high endemicity areas. The highest cure was consistently achieved in children with light or heavy infection when arachidonic acid was combined with praziquantel ¹²⁹.

A breakthrough regarding the usefulness of arachidonic acid in defense against schistosomes came from the demonstration of association between resistance of the water-rat, Nectomys squamipes to repeated infection with Schistosoma mansoni and accumulation of arachidonic acid in liver cells ¹³⁰. This pioneering study prompted us to examine the relation between susceptibility and resistance of rodents to Schistosoma mansoni or Schistosoma haematobium infection and arachidonic acid levels in serum and lung and liver cells before and weekly after infection. The results strongly suggested that arachidonic acid is a potent “natural” schistosomicide, and may be considered an endoschistosomicide ¹³¹.

The schistosomicidal action of arachidonic acid is based on excessive hydrolysis of parasite surface membrane sphingomyelin. Interestingly, Miltefosine, a hexa-decyl-phosphocholine, which interferes with proper biosynthesis of sphingomyelin, was recently documented as a potent schistosomicide in vitro and in vivo ¹³².

Anti-cancer potential

Reports decades earlier indicated that PUFA, and especially free unesterified arachidonic acid possess tumoricidal activity in vitro and in vivo ¹³³. The most important and consistent studies documenting the tumoricidal action of PUFA, namely arachidonic acid were reported by Undurti Das and Colleagues ¹³⁴, who advocated arachidonic acid as a potential anti-cancer drug ¹³⁵. Thus, arachidonic acid was reported to kill tumor cells selectively in vitro via eliciting cell surface membrane lipid peroxidation, which can be blocked by vitamin E, uric acid, glutathione peroxidase and superoxide dismutase ¹³⁶. Free arachidonic acid was found to inhibit the in vitro growth of human cervical carcinoma cells and methyl cholanthrene-induced sarcoma cells. Free arachidonic acid augmented the generation of superoxide anion and lipid peroxidation in the tumor cells indicating a possible correlation between the ability of unesterified PUFA to augment free radicals and their tumoricidal action ¹³⁷. Moreover, free unesterified arachidonic acid, independently of its metabolites, displayed cytotoxic action on both vincristine-sensitive (KB-3-1) and resistant (KB-Ch(R)-8-5) cancer cells in vitro that appeared to be a free-radical dependent process ¹³⁸. At concentrations of 100–200 µM, arachidonic acid was more effective than mesotrexate in in vitro suppression of gastric carcinoma cells, as a result of lipid peroxidation processes ¹³⁹ and inhibited proliferation of human prostate cancer and human prostate epithelial cells, independently of free radicals generation ¹⁴⁰. Arachidonic acid-mediated apoptosis of colon cancer cells appeared to be essentially due to loss of mitochondrial membrane, accumulation of ROS, and caspase-3 and caspase-9 activation ¹⁴¹. Accordingly, it was concluded that arachidonic acid suppresses proliferation of normal and tumor cells by a variety of mechanisms that may partly depend on the type(s) of cell(s) being tested and the way arachidonic acid is handled by the cells ¹³⁸. A contradictory effect of arachidonic acid on tumorigenesis was observed in mice with a germline mutation in the adenomatous polyposis coli gene ¹⁴².

Tallima and El Ridi ¹⁴³ have recently proposed that arachidonic acid may inhibit proliferation and elicit death of tumor cells via its activating impact on cell membrane-associated neutral sphingomyelinase (nSMase) and increased outer leaflet sphingomyelin hydrolysis. Disruption of the tight sphingomyelin-based hydrogen bond network around cancer cells may allow contact inhibition processes to proceed and cell proliferation to stop ⁷¹, whereby the primary sphingomyelin catabolite, ceramide released following sphingomyelin hydrolysis is a renowned secondary messenger involved in programmed cell death ¹⁴⁴. It is of importance to recall that Miltefosine, which has been approved for the treatment of breast cancer metastasis, and is currently used for the treatment of cutaneous metastases of mammary carcinoma significantly inhibits sphingomyelin biosynthesis in human hepatoma and other tumor cells ¹⁴⁵. The likely mechanism of action of this phospholipid analogue is inhibition of phosphatidylcholine biosynthesis, thus hindering sphingomyelin metabolism, and substantially increasing the levels of ceramide ¹⁴⁵.

Roles in type 2 immune responses

Allergens, cysteine peptidases and numerous helminth-derived excretory-secretory products disrupt the epithelial or endothelial barriers, eliciting release of the type 2 immunity master cytokines and alarmins, TSLP (thymic stromal lymphopoietin), interleukin (IL)-25 and IL-33 ¹²⁶. These cytokines bind to receptors on innate lymphoid cells 2 (ILC2), tissue-resident sentinels, mainly found in the skin and at mucosal surfaces of intestine and lungs. Cytokine-receptors’ interactions result into signals that induce ILC2 recruitment, proliferation and activation. The activated ILC2 produce type 2 cytokines, principally IL-5 and IL-13, which are instrumental in the recruitment and activation of eosinophils, basophils, and mast cells ¹⁴⁶. Major basic proteins, proteases, histamine, heparin, type 2 cytokines, and reactive ROS are not only produced inducing the various signs of inflammation, arachidonic acid is furthermore released from the activated cell membrane and oxidized to inflammatory metabolites ¹². The arachidonic acid-derived metabolites are the road to generation of resolvins that help in resolving inflammation and wound and lesion healing ¹⁰⁹. Of considerable importance is the discovery that ILC2 share with airway and gut smooth muscle cells, and/or epithelial cells, eosinophils, mast cells, macrophages, dendritic cells, and T helper 2 (Th2) lymphocytes surface membrane receptors for arachidonic acid-derived metabolites. Chemoattractant receptor-homologous molecule expressed on Th2 cells (CRTH2) is a receptor for prostaglandin D2, CysLTR for cysteinyl leukotrienes D4 and E4 and ALXR for lipoxin A4. Prostaglandin D2, leukotriene D4 and E4 stimulate, while lipoxin A4 inhibits ILC-2 expansion and effector functions ¹⁴⁷, thus, implicating arachidonic acid metabolites as secondary inducers of type 2 immune responses’ amplification, regulation and memory in airway and gut hyperresponsiveness and repair, and resistance to parasites ¹⁴⁸.

Endocannabinoids

Endocannabinoids are so termed because they activate the same G protein-coupled, cannabinoid receptors (CB1 and CB2) as delta-9-tetrahydrocannabinol, the active component of marijuana (Cannabis sativa). Endocannabinoids are important modulators of brain reward signaling, motivational processes, emotion, stress responses, pain and energy balance ¹⁴⁹. The endocannabinoids, N-arachidonoyl-ethanolamine and 2-arachidonoylglycerol, are arachidonic acid-derived. arachidonic acid Trans-acylase-catalyzed transfer of arachidonic acid from the sn-1 position of phospholipids to the nitrogen atom of phosphatidylethanolamine generates N-arachidonoyl-phosphatidylethanolamine (NAPE). NAPE can be hydrolyzed to arachidonylethanolamine (anandamide) in a one-step reaction catalyzed by NAPE-specific phospholipase D, or two-steps reaction catalyzed by a phospholipase C and a phosphatase. NAPE can be converted to anandamide via two further synthetic pathways ¹⁵⁰. The importance of anandamide can be inferred from the redundancy of its precursor conversion pathways. 2-arachidonoylglycerol (2-AG) is produced from the hydrolysis of diacylglycerols (DAGs) containing arachidonate in the 2 position, catalyzed by a DAG lipase that is selective for the sn-1 position ¹⁵⁰.

Interaction of arachidonic acid-derived endocannabinoids with their specific receptors generate signals, which control neural processes that underpin key aspects of social behavior whereby endocannabinoid signaling dysregulation is associated with social impairment related to neuropsychiatric disorders ¹⁴⁹. Endocannabinoid-mediated signaling, especially in the brain, modulates a variety of pathophysiological processes, including appetite, pain and mood, whereby inhibition of endocannabinoid degradation is predicted to be instrumental in reducing pain and anxiety ¹⁵¹. Additionally, anandamide appeared to modulate human sperm motility ¹⁵² and improve renal functions and chronic inflammatory disorders of the gastrointestinal tract by regulating gut homeostasis, gastrointestinal motility, visceral sensation, and inflammation ¹⁵³.

Arachidonic acid side effects

Arachidonic acid supplementation in daily doses of 1,000–1,500 mg for 50 days has been well tolerated during several clinical studies, with no significant side effects reported. All common markers of health, including kidney and liver function ¹⁵⁴, serum lipids ¹⁵⁵, immunity ¹⁵⁶ and platelet aggregation ¹⁵⁷ appear to be unaffected with this level and duration of use. Furthermore, higher concentrations of arachidonic acid in muscle tissue may be correlated with improved insulin sensitivity ¹⁵⁸. However a study by Thomas and colleagues showed in mice study that dietary arachidonic acid could amplify beta-amyloid peptide oligomer neurotoxicity ¹⁵⁹. Thomas and colleagues found that the administration of an arachidonic acid-enriched diet in mice for 12 weeks induced short-term memory impairment and increased deleterious effects of beta-amyloid oligomers on learning abilities ¹⁵⁹. Arachidonic acid consumption could constitute a risk factor for Alzheimer’s disease in humans and should be taken into account in future preventive strategies ¹⁶⁰. Arachidonic acid deleterious effect on cognitive capacity could be linked to the balance between arachidonic acid-mobilizing enzymes.

Several studies have suggested that increase of dietary arachidonic acid intake or tissue arachidonic acid amounts favors the occurrence of systemic chronic inflammation ¹⁶¹ and the severity of pathologies with a strong inflammatory component, such as chronic inflammatory intestinal diseases ¹⁶², rheumatoid arthritis ¹⁶³ and atherosclerosis ¹⁶⁴. This effect would be mediated by the increase of cellular free arachidonic acid and its conversion into proinflammatory eicosanoids, especially in individuals with some genetic backgrounds and in animal models ¹⁶⁵. Furthermore, neuroinflammation is one of the pathological components of Alzheimer’s disease, and researchers in many studies have reported higher expression of enzymes using arachidonic acid as a substrate, such as cyclooxygenases and prostaglandin synthases ¹⁶⁶ or brain production of proinflammatory eicosanoids in an Alzheimer’s disease mice model ¹⁶⁷. However, despite the fact that neuroinflammation can induce the first synaptic dysfunction and cognitive impairment in Alzheimer’s disease, before beta-amyloid peptide accumulation and plaque formation ¹⁶⁸, it is not known if higher arachidonic acid levels in the brain facilitate neuroinflammation. Additional experiments on inflammation kinetics in animal model and using various diets would be necessary to clarify this point.

While studies looking at arachidonic acid supplementation in sedentary subjects have failed to find changes in resting inflammatory markers in doses up to 1,500 mg daily, strength-trained subjects may respond differently. One study reported a significant reduction in resting inflammation (via marker IL-6) in young men supplementing 1,000 mg/day of arachidonic acid for 50 days in combination with resistance training ¹²³. This suggests that rather being pro-inflammatory, supplementation of arachidonic acid while undergoing resistance training may actually improve the regulation of systemic inflammation ¹⁶⁹.

A meta-analysis looking for associations between heart disease risk and individual fatty acids reported a significantly reduced risk of heart disease with higher levels of EPA and DHA (omega-3 fats), as well as the omega-6 arachidonic acid ¹⁷⁰. A scientific advisory from the American Heart Association has also favorably evaluated the health impact of dietary omega-6 fats, including arachidonic acid ¹⁷¹. The group does not recommend limiting this essential fatty acid. In fact, the paper recommends individuals follow a diet that consists of at least 5–10% of calories coming from omega-6 fats, including arachidonic acid. It suggests dietary arachidonic acid is not a risk factor for heart disease, and may play a role in maintaining optimal metabolism and reduced heart disease risk. Maintaining sufficient intake levels of both omega-3 and omega-6 fatty acids, therefore, is recommended for optimal health.

Arachidonic acid is not carcinogenic, and studies show dietary level is not associated (positively or negatively) with risk of cancers  ¹⁷², ¹⁷³. Arachidonic acid remains integral to the inflammatory and cell growth process, however, which is disturbed in many types of disease including cancer. Therefore, the safety of arachidonic acid supplementation in patients suffering from cancer, inflammatory, or other diseased states is unknown, and supplementation is not recommended.

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