Fasting enriches liver triacylglycerol with n-3 polyunsaturated fatty acids: implications for understanding the adipose–liver axis in serum docosahexaenoic acid regulation

Genes Nutr (2015) 10:39 DOI 10.1007/s12263-015-0490-2 RESEARCH PAPER Fasting enriches liver triacylglycerol with n-3 polyunsaturated fatty acids: implications for understanding the adipose–liver axis in serum docosahexaenoic acid regulation Kristin A. Marks1,2 • Phillip M. Marvyn1 • Juan J. Aristizabal Henao2 • Ryan M. Bradley1 • Ken D. Stark2 • Robin E. Duncan1 Received: 29 May 2015 / Accepted: 29 August 2015 / Published online: 19 September 2015 Ó Springer-Verlag Berlin Heidelberg 2015 Abstract We investigated the effect of short-term fasting on coordinate changes in the fatty acid composition of adipose triacylglycerol (TAG), serum non-esterified fatty acids (NEFA), liver TAG, and serum TAG and phospholipids in mice fed ad libitum or fasted for 16 h overnight. In contrast to previous reports under conditions of maximal lipolysis, adipose tissue TAG was not preferentially depleted of n-3 PUFA or any specific fatty acids, nor were there any striking changes in the serum NEFA composition. Short-term fasting did, however, increase the hepatic proportion of n-3 PUFA, and almost all individual species of n-3 PUFA showed relative and absolute increases. The relative proportion of n-6 PUFA in liver TAG also increased but to a lesser extent, resulting in a significant decrease in the n-6:n-3 PUFA ratio (from 14.3 ± 2.54 to 9.6 ± 1.20), while the proportion of MUFA decreased significantly and SFA proportion did not change. Examination of genes involved in PUFA synthesis suggested that hepatic changes in the elongation and desaturation of precursor lipids could not explain this effect. Rather, an increase in the expression of fatty acid transporters specific for 22:6n-3 and other long-chain n-3 and n-6 PUFA Electronic supplementary material The online version of this article (doi:10.1007/s12263-015-0490-2) contains supplementary material, which is available to authorized users. & Robin E. Duncan 1 Lipid Enzyme Discovery Lab, Department of Kinesiology, University of Waterloo, 200 University Ave W, Waterloo, ON N2L 3G1, Canada 2 Laboratory of Nutritional and Nutraceutical Research, Department of Kinesiology, University of Waterloo, 200 University Ave W, Waterloo, ON N2L 3G1, Canada likely mediated the observed hepatic enrichment. Analysis of serum phospholipids indicated a specific increase in the concentration of 22:6n-3 and 16:0, suggesting increased specific synthesis of DHA-enriched phospholipid by the liver for recirculation. Given the importance of blood phospholipid in distributing DHA to neural tissue, these findings have implications for understanding the adipose– liver–brain axis in n-3 PUFA metabolism. Keywords n-3 Polyunsaturated fatty acids  Triacylglycerol  Adipose  Non-esterified fatty acids  Liver  Phospholipids  Fatty acid desaturases  Fatty acid elongases  Fatty acid transport proteins  Fatty acid binding proteins Introduction Adipose tissue lipolysis is activated during fasting [reviewed in (Raclot 2003; Duncan et al. 2007)]. The triacylglycerol (TAG) stored in adipose tissue is hydrolyzed at an increased rate by the lipases adipose triglyceride lipase (ATGL) and hormone-sensitive lipase (HSL) (Jaworski et al. 2007; Duncan et al. 2010; Ahmadian et al. 2009), releasing more non-esterified fatty acids (NEFA) into the circulation to provide lipid substrates for the body. HSL has been shown to display a preference for TAG containing long-chain polyunsaturated fatty acids (PUFA) (Raclot et al. 2001), and there is some evidence of selective mobilization of more unsaturated fatty acids from adipose tissue. Connor and colleagues maximally induced lipolysis in rabbits by injecting adrenocorticotropic hormone after an overnight fast and then calculated the relative mobilization of individual fatty acid species as a ratio of the percent abundance in plasma NEFA relative to the percent 123 39 Page 2 of 14 in adipose tissue TAG plus free fatty acid fractions combined. Under these conditions, the calculated relative mobilization tended to increase as the degree of unsaturation increased for fatty acids of a specific chain length (Conner et al. 1996). Although this finding may have been confounded by effects of selective uptake on serum NEFA concentrations, Raclot et al. also observed a highly similar effect using isolated human mammary adipocytes maximally stimulated to undergo lipolysis with isoprenaline and adenosine deaminase (Raclot et al. 1997). Both studies also agreed on the finding that 20:5n-3 and 20:4n-6 were proportionately the most highly mobilized fatty acyl species. Despite these reports, and others (Yli-Jama et al. 2001; Hellmuth et al. 2013), no studies have yet characterized the fatty acid composition of adipose TAG and serum NEFA in ad libitum fed animals versus animals undergoing a shortterm fast, when lipolysis is not maximally stimulated. Furthermore, downstream effects of fasting-mediated changes in adipocyte and serum fatty acids have also yet to be characterized. For example, it is unknown whether short-term fasting leads to selective changes in the composition of stored fatty acids in liver TAG, or to changes in liver-derived circulating complex lipids. Understanding the metabolic journey of adipose-derived fatty acids, and PUFA in particular, has a variety of implications for health. Most PUFA are either essential or conditionally essential for cellular processes (Cunnane 2000). Evidence of the selective mobilization of PUFA from adipose tissue, particularly under relatively common conditions such as an extended overnight fast, would constitute a ‘‘second chance’’ mechanism ensuring that essential fatty acids remain bioavailable for use by tissues, rather than locked in the core of adipocyte lipid droplets. Additionally, it has recently been found that the brain, which requires a constant supply of the very long chain n-3 PUFA docosahexaenoic acid (DHA, 22:6n-3) (Rahman et al. 2010; Polozova and Salem Jr 2007), can also uptake DHA as lysophosphatidylcholine via a Mfsd2a receptor (Nguyen et al. 2014) in addition to crossing as a NEFA (Domenichiello et al. 2015), indicating a potentially significant role for the adipose–liver axis in brain health. The aim of the present study, therefore, was to better understand the metabolism of adipose-derived NEFA by comparing fatty acid profiles in overnight fasted versus ad libitum fed mice, in the following pools: (1) adipose TAG, (2) blood NEFA that are primarily derived from adipose TAG, (3) hepatic TAG, which are synthesized primarily from circulating NEFA in the post-absorptive state, and (4) serum phospholipids and (5) serum TAG, which in fasting are derived predominantly from hepatic rather than intestinal synthesis. To better understand our findings in liver, we also determined the relative hepatic gene expression of desaturase and elongase genes involved 123 Genes Nutr (2015) 10:39 in PUFA biosynthesis, and the expression of genes involved in NEFA uptake. Methods Animals Al (...truncated)