The Δ9 Fatty Acid Desaturation Index as a Predictor of Metabolic Disease

There is a tremendous need for improved biomarkers that can be used in the clinical setting to aid in the early detection of metabolic diseases. Of particular interest is the metabolic syndrome, or insulin resistance syndrome, a condition that is often clinically diagnosed as a cluster of risk factors including abdominal obesity, hypertriglyceridemia, low HDL cholesterol, hypertension, impaired glucose tolerance, and increased fasting glucose (1)(2). This constellation of phenotypes greatly increases the risk for the development of chronic conditions such as diabetes and several cardiovascular diseases. The root cause of the metabolic syndrome is insulin resistance, a condition in which the pancreas increases insulin secretion to compensate for the decreased ability of insulin-sensitive tissues to perform insulin-stimulated glucose uptake to maintain euglycemia (1). Therefore, the development of new tests that focus on the early detection of insulin resistance could help identify at-risk patients so that the benefits of lifestyle and pharmacological interventions can be maximized and progression to overt metabolic disease can be prevented. Insulin resistance is not limited to defects in carbohydrate metabolism, but is also associated with major perturbations in lipid metabolism (2). For example, adipocytes increase the release of free fatty acids (FFA)1 into the circulation, resulting in subsequent FFA uptake by the liver and other peripheral tissues, processes that may further exacerbate insulin resistance (2)(3). Although hepatic insulin resistance contributes to hyperglycemia by causing an inappropriate elevation in hepatic glucose production, the branch of insulin signaling responsible for stimulating de novo fatty acid biosynthesis in the liver remains intact (2)(4). Additionally, high-carbohydrate diets, especially those enriched in rapidly absorbed refined carbohydrates such as glucose and fructose, are able to potently stimulate de novo lipogenesis (DNL) (5). The effects of insulin and dietary carbohydrate occur via the transcriptional activation of lipogenic transcription factors such as sterol regulatory element binding protein (SREBP) and carbohydrate response-element binding protein, which increase the transcription of genes encoding enzymes of fatty acid synthesis such as acetyl–coenzyme A (CoA) carboxylase, fatty acid synthase, and the elongase Elovl6 (4)(5). These enzymes promote the conversion of acetyl-CoA into the 16-carbon saturated fatty acid, palmitic acid (16:0) and its 18-carbon elongation product stearic acid (18:0). Another enzyme that is upregulated under these conditions is stearoyl-CoA desaturase-1 (SCD1), a Δ9 fatty acid desaturase that catalyzes the conversion of 16:0 and 18:0 into the monounsaturated fatty acids palmitoleic (16:1n7) and oleic acid (18:1n9), respectively (6). During conditions of increased hepatic lipogenesis there is often a concomitant increase in the SCD1 product/substrate ratio, reflecting increased hepatic SCD1 activity. Thus, fatty acid profiling of biological samples holds the potential to be used in the clinical diagnosis of insulin resistance and related metabolic diseases. Hepatic fatty acids derived from both de novo synthesis and adipose-derived FFA accumulate in the liver primarily as triacylglycerides (TG) and result in hepatic steatosis during insulin-resistant states (2). These intracellular TG stores are used by the liver to assemble a TG-rich VLDL particle, which is secreted into the circulation during both the fed and fasted states (7). In this issue of Clinical Chemistry, Peter et al. report the results of simultaneous analysis of the fatty acid composition of defined lipid fractions in human liver and plasma VLDL, as well as the determination of liver lipogenic gene expression (8). The authors first set out to determine whether hepatic stearoyl-CoA desaturase (delta-9-desaturase) (SCD)2 gene expression correlates with the fatty acid composition of hepatic TG, diacylglycerols (DG), FFA, cholesterol ester (CE), and phospolipid (PL). Peter et al. report a strong correlation between SCD mRNA levels and the 16:1n7/16:0 ratios of liver TG, FFA, CE, and PL, suggesting that the fatty acid composition of the liver can be predictive of hepatic SCD1 activity. These authors then investigated whether Δ9 fatty acid indices in plasma VLDL can be used as a surrogate to a liver sample to predict hepatic SCD gene expression and hepatic fatty acid composition. Peter et al. identified a significant association between total plasma VLDL or VLDL-TG 16:1n7/16:0 and the 16:1n7/16:0 ratios in hepatic TG, DG, FFA, CE, and PL. A much weaker association was identified among hepatic SCD mRNA and the 18:1n9/18:0 ratios of hepatic lipids and plasma VLDL. This weaker association may be explained by the high dietary abundance of 18:1n9 relative to the scarce amount of 16:1n7 in the diet, which may results in the 18:1n9/18:0 ratios being more susceptible to dietary fat intake. Alternatively, the authors suggest that 16:1n7 may be a more sensitive indicator of SCD activity relative to 18:1n9, owing to the large accumulation of previously synthesized 18:1n9 relative to 16:1n7 in hepatic TG, which dilute the signal from newly synthesized fatty acids. Because liver biopsies are not available for routine clinical tests, these findings validate the analysis of plasma VLDL-TG fatty acids as a means to provide insight into liver lipid metabolism. The plasma compartment is a complex mixture of lipids primarily associated with VLDL, LDL, HDL, and chylomicrons (in the postprandial state) that include mostly TG, CE, PL, and unesterified cholesterol, as well as non–lipoprotein-associated FFA. In their study, Peter et al. also compared the Δ9 desaturation indices in total VLDL fatty acids to those specifically in the VLDL-TG fraction and found them to be nearly identical for 16:1n7/16:0 and also significantly associated for 18:1n9/18:0 (8). This high correlation is likely attributable to the large neutral lipid core of VLDL being composed of primarily liver-derived TG, allowing for the analysis of total VLDL fatty acids or VLDL-TG (7). Thus, the application of thin-layer chromatography to isolate TG is apparently unnecessary. Most of the plasma TG in a fasting blood sample is associated with VLDL, and this situation may allow the circumvention of VLDL isolation by ultracentrifugation by enabling analysis of whole plasma TG. Either of these modifications could make this assay more amenable for large-scale population analyses. Many previous studies in humans have relied on total lipid extracts from whole plasma or tissues to obtain fatty acid indices and infer SCD1 activity (9)(10)(11)(12). As highlighted by Karpe and Hodson, the analysis of whole plasma fatty acid ratios can yield misleading results because each plasma lipid class (TG, CE, FFA, and PL) has a unique fatty acid enrichment profile and there is not a concerted elevation in all lipid classes during the accumulation of TG in liver or plasma (13)(14). Thus, when whole plasma fatty acid ratios from hyperlipidemic patients are analyzed, the increased TG content, and not necessarily a change in SCD activity, may yield an apparently higher 16:1n7/16:0 or 18:1n9/18:0 ratio simply because of the higher molar percentage of 16:1n7 and 18:1n9 found in the TG fraction relative to other lipid classes (14). This salient point is also conveyed by Peter et al., who show that the 16:1n7/16:0 and 18:1n9/18:0 ratios differ remarkably between liver lipid classes (8). One effect of hepatic insulin resistance is an increase in VLDL-TG production (2)(7). Therefore, it is of extreme importance that future studies aimed at using fatty acid ratios to infer SCD activity adhere to the recommendation to use defined lipid fractions to avoid confusing changes in SCD activity with changes in lipid composition. Fatty acid ratios also hold the potential to be used to identify individuals with abnormally high liver fatty acid synthesis who are at risk for nonalcoholic fatty liver disease. The 16:0/18:2n6 ratio, which is a ratio of the main product of DNL and an essential dietary fatty acid, is often used as an index of DNL. Chong et al. have previously reported a parallel activation of DNL and SCD activity after short-term high-carbohydrate feeding by using fatty acid ratios as well as conversion of intravenously infused [2H2]-16:0 to [2 H2]-16:1n7 in VLDL-TG (15). Although Chong et al. reported a good association (P = 0.06) between the VLDL-TG 16:1n7/16:0 and 16:0/18:2n6 ratios, Peter et al. analyzed this 16:0/18:2n6 DNL index in both VLDL and liver TG and found these 2 indices to be highly correlated (8)(15). In the study by Peter et al., the hepatic-TG DNL index was also associated with expression of the hepatic lipogenic genes acetyl-Coenzyme A carboxylase alpha (ACACA), fatty acid synthase (FASN), and sterol regulatory element binding transcription factor 1 (SREBP-1), and changes in the expression of these genes were also closely reflected by the VLDL-TG DNL index. Together, these 2 studies suggest that hepatic DNL can be estimated from either the VLDL-TG DNL index (16:0/18:2n6) or the SCD activity index (16:1n7/16:0). Another study by Kotronen et al. found no relationship between the hepatic DNL and SCD indices, but this study analyzed total lipids and used the 18:1n9/18:0 ratio instead of the 16:1n7/16:0 ratio (16). Interestingly, Stefan et al. reported that in obese but not lean individuals the VLDL-TG 18:1n9/18:0 ratio is inversely correlated with liver fat quantified by 1H-magnetic resonance (17). The incongruent conclusions of the above studies emphasize the need for further validation of these fatty acid indices before clinical application. SCD1 has been implicated in a variety of metabolic disorders, including obesity, insulin resistance, diabetes, fatty liver, hyperlipidemia, inflammation, atherosclerosis, and cancer (6). Plasma fatty acid ratios such as the SCD desaturation index hold the potential to serve as noninvasive and sensitive predictors for the early stages of these chronic diseases and may be the newest members of the metabolic syndrome phenotypic cluster. Additionally, monitoring of the plasma VLDL-TG SCD desaturation index could potentially measure the efficacy of dietary and lifestyle interventions on reducing hepatic insulin resistance. It is noteworthy that the hepatic activity of SCD and other lipogenic enzymes can be acutely affected by dietary composition (5). Therefore, future applications of fatty acid ratios must be interpreted in the context of both acute and chronic changes in metabolism. Author Contributions:All authors confirmed they have contributed to the intellectual content of this paper and have met the following 3 requirements: (a) significant contributions to the conception and design, acquisition of data, or analysis and interpretation of data; (b) drafting or revising the article for intellectual content; and (c) final approval of the published article. Authors’ Disclosures of Potential Conflicts of Interest:No authors declared any potential conflicts of interest. Role of Sponsor: The funding organizations played no role in the design of study, choice of enrolled patients, review and interpretation of data, or preparation or approval of manuscript.