Free Fatty Acids

Test Description

Plasma fatty acids can be esterified to form triacylglycerols (triglycerides [TGs]) for storage, or metabolized as a high-energy fuel source of adenosine triphosphate (ATP). Plasma fatty acids that are not esterified to TGs are bound primarily to albumin and are termed “nonesterified fatty acids (NEFAs)” or “free fatty acids (FFAs).” FFAs are the primary form by which lipid stored in adipose tissue is transported to other sites for utilization.1

The FFAs are released from adipocytes via lipolysis of TGs, mediated by the combined actions of adipose triglyceride ligase (ATGL) and hormone sensitive lipase (HSL).2,3 Release and uptake of FFAs by tissues such as skeletal muscle and liver, is regulated by both insulin and catecholamines. FFA turnover is rapid, with a plasma half-life of ~2-4 min.4

Mitochondrial oxidation of FFAs (fatty acid β-oxidation) provides a significant energy source for organs and body tissues including myocardium, skeletal muscle, liver, and the kidney. Specific FFAs may also directly regulate cellular function, either by interactions with membrane phospholipid fatty acids, or in their role as natural ligands of cytoplasmic nuclear hormone receptors, such as peroxisome proliferator-activated receptors (PPARs).2

Following a meal, most lipids are transported as triglycerides packaged in chylomicrons and taken up primarily by adipose tissue and skeletal muscle.5,6 Both tissues can take up albumin-bound FFAs, but the majority of FFA uptake in adipose tissue is from chylomicrons after a meal. Of the postprandial FFAs taken up into adipocytes for storage, a proportion escapes and joins the plasma FFA pool (they may constitute 40-50% of circulating FFAs), in a process referred to as “spillover.”1,7 Hence, the plasma FFA pool changes in composition after a meal to reflect the fatty acid content of the meal.8

Secretion of insulin, stimulated by glucose in the postprandial state, results in suppression of lipolysis and a rapid drop in circulating FFAs. Circadian profiles of plasma FFA levels therefore show the highest concentrations after an overnight fast (when insulin is lowest and catecholamines high), with suppression after each meal.6,9,10

Typical plasma FFA concentrations range from around 0.3 to 0.6 mmol/L in the overnight fasting state (depending upon methodology and cohort) to 1.3 mmol/L after a 72-hour fast.11 Plasma FFA concentrations are usually found to be higher in women than in men,12-14 and are increased under physiological/psychological stress.15

Risk ranges for the Free Fatty Acids test are as follows:

Free Fatty Acids (mmol/L):

• High risk > 0.7
• Intermediate risk 0.6-0.7
• Optimal < 0.6

Clinical Interpretation

Elevated FFA and metabolic syndrome

Alterations in FFA metabolism are likely a major factor involved in the pathogenesis of hyperglycemia and dyslipidemia associated with the metabolic syndrome (MetS). MetS represents a constellation of metabolic abnormalities that are risk factors for cardiovascular disease [coronary heart disease (CAD), myocardial infarction, and stroke].16

The clinical criteria of Metabolic Syndrome include:

• abdominal obesity (high body mass index and/or large waist circumference)
• insulin-resistant glucose metabolism (hyperinsulinemia, impaired fasting glucose, impaired glucose tolerance, type 2 diabetes)
• dyslipidemia (high serum TG and low serum HDL-C concentrations)
• increased blood pressure.17

Metabolic Syndrome is also known as the insulin resistance syndrome, as insulin resistance is thought to be an underlying cause of the metabolic derangements.18

Elevated FFA levels are also associated with insulin resistance, leptin resistance, and decreased adiponectin levels. Decreased fatty acid oxidation may cause increased plasma FFA levels and evidence suggests that defects in fatty acid oxidation may precede the obese diabetic state.2,19,20 High plasma FFA levels are commonly observed in type 2 diabetes (T2DM), with an early increase indicating a shift from impaired glucose tolerance (IGT) to T2DM.10,21

Mechanisms underlying FFA-induced insulin resistance

Insulin, which inhibits lipolysis, is the major physiological regulator of basal adipose tissue lipolytic activity.22,23 Lipolysis of adipose tissue TGs is the major source of plasma FFA. Therefore, insulin resistance in adipose tissue stimulates an increase in lipolytic rate and FFA release into the bloodstream. The typical increase in plasma insulin levels associated with obesity does not completely compensate for adipose tissue insulin resistance, so insulin-resistant obese subjects have high basal lipolytic rates and plasma FFA levels.23

Increased plasma FFA levels can impair the ability of insulin to stimulate muscle glucose uptake and oxidation24,25 and suppress hepatic glucose production,26,27 leading to hyperglycemia. Increased delivery of FFAs to the liver can lead to atherogenic dyslipidemia by inducing hepatic very low-density lipoprotein (VLDL)-TG production,28,29 which results in elevated TG and apolipoprotein B (apoB) levels, and a subsequent decrease in HDL-C levels.30-33 This in turn increases the transfer of TG from VLDL to HDL, leading to increased HDL clearance and decreased plasma HDL levels.34

The resulting lipid profile, often observed with both T2DM and Metabolic Syndrome, is associated with significantly increased risk for cardiovascular disease (CVD).35-37 Furthermore, elevated FFA concentrations also impair endothelial function, stimulate vasoconstriction, and increase sodium absorption, possibly leading to increased blood pressure.38,39

Other factors related to intracellular fatty acid metabolism also contribute to insulin resistance. Relative impairments of mitochondrial oxidative phosphorylation in skeletal muscle have been identified in persons who have insulin resistance and are at increased risk for developing T2DM.40 Impaired mitochondrial fatty acid oxidation, in particular, may contribute to impaired insulin action by increasing the intracellular accumulation of fatty acids.

Adverse effects of elevated FFA in the liver and vasculature

Like those in skeletal muscle, cellular events responsible for fatty acid-induced insulin resistance in the liver are fairly complicated. In addition to the impairment of insulin-mediated glucose uptake and utilization by the liver,17,37 the effect of elevated FFAs in the liver may indirectly lead to increased small dense (sd)LDL particle concentration, by stimulating secretion of apoB and raising numbers of apoB-containing lipoprotein particles which eventually become remodeled to sdLDL particles. The presence of such atherogenic particles has been associated with an increased risk for myocardial infarction and worsened severity of coronary artery disease (CAD).34 In insulin resistance, arteries are also exposed to high chronic levels of albumin-FFA complexes and

FFAs are locally released by lipases from the lipids of apoB-lipoproteins entrapped by proteoglycans of the intima.41,42 Therefore, in insulin resistance and diabetes, the main organ responsible for apoB-lipoprotein generation and metabolism, the liver, and the endothelium, where these liver products can initiate atherosclerotic lesions, are both exposed to potentially deleterious effects of excess circulating FFAs.

Seminal experiments by Hennig et al. showed that exposure of endothelial cells to high levels of albumin-bound fatty acids altered the amount and composition of the basement membrane proteoglycans, causing endothelial cell activation, an early step in the atherosclerotic process.43 Fatty acids can increase permeability of the endothelial basement membrane by reduction of heparan proteoglycans,44 while cellular exposure to high FFA levels increases intracellular diacylglycerol (DAG) concentration, activating protein kinase C (PKC) isoforms and resulting in compromised insulin signaling. By this mechanism, FFAs reduce insulin’s control of proteoglycan expression, which may contribute to increase retention of LDL in the intima, a key step in atherogenesis.45,46

FFAs are an important precursor for the synthesis of ceramides (a type of sphingolipid) that exert multiple injurious effects on the vasculature, including induction of reactive oxygen species and induction of proinflammatory nuclear factor kappa B (NFkB)-mediated signaling.47 Multiple studies have implicated the toll-like receptor-4 (TLR4) as a FFA “receptor” that activates NFkB signaling in endothelium and impairs insulin action in other tissues.48 It has been proposed that the chronic inflammatory state induced by FFAs and ceramides not only increases cardiovascular risk but further worsens insulin-glucose homeostasis in a positive feedback manner.49,50

In summary, recent studies support the increasing body of literature demonstrating that high FFA levels have deleterious effects and promote the development of Metabolic Syndrome, Type-2 diabetes, and cardiovascular disease. FFA levels are highly predictive of cardiovascular death even after adjustment for all of the classic cardiovascular risk factors. Clinical studies have shown that high FFA levels are associated with impairment of endothelium-dependent vasodilation, high blood pressure, myocardial infarction, stroke, atrial fibrillation, and sudden death.1-3,38,52-53

Treatment Considerations

Whether FFAs are only an independent risk marker of specific underlying pathophysiology, or also directly impact cardiovascular risk, interventions to reduce FFA levels should target the underlying cardiometabolic stress (e.g., at the level of the adipocyte). Reducing circulating FFA levels should be considered an important target for any therapeutic intervention intended to decrease CVD, especially with concomitant insulin resistance.

A combination of basic lifestyle measures such as

• smoking cessation
• greater physical activity
• modest dietary change (e.g., reduced calories, consumption of whole grains in place of refined carbohydrates, reduced trans fat)
• modest weight loss
• drugs that reduce VLDL (i.e., niacin, fenofibrate) and reduce FFA levels
• drugs that improve insulin sensitivity (e.g., metformin).54
• PPAR gamma receptor agonist, pioglitazone, may also inhibit FFA release from adipose tissue via enhanced insulin activity, as well as by its ability to promote lipogenesis and lipid storage over fatty acid release.

Although insulin resistance is the sole cause of elevated FFA, women and the elderly have higher levels than men or younger individuals. Therefore, it is important to evaluate the overall cardiometabolic risk profile of each individual with elevated FFA levels.

Lifestyle55-61

• Limit carbohydrates (especially simple sugars and processed carbohydrates) while maintaining moderate fat intake
• Regular exercise (150 minutes/week combining cardiovascular activity at a moderate-to-vigorous pace with resistance training)

Medication choices may include:

• Metformin (e.g., Glucophage®, Glumetza®)
• Pioglitazone (Actos®)
• Pioglitazone-metformin combination

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