Reducing the Dietary Omega 6 Omega 3 Utilizing alpha Linolenic Acid; Not a Sufficient Therapy for Attenuating High Fat Diet Induced Obesity Development Nor Related Detrimental Metabolic and Adipose Tissue Inflammatory Outcomes

Aims  To examine the effect of manipulating the omega 6 omega 3 ratio utilizing only a alpha linolenic and linoleic acid within a clinically relevant high fat diet (HFD) composed of up to seven sources of fat and designed to be similar to the standard American diet (MUFA PUFA of 2 1, 12 per cent and 40 per cent of calories from saturated and total fat, respectively) on body composition, macrophage polarization, inflammation, and metabolic dysfunction in mice. Methods  Diets were administered for 20 weeks. Body composition and metabolism (HOMA index and lipid profile) were examined monthly. GC  MS was utilized to determine the eicosapentaenoic acid (EPA) arachidonic acid (AA) and the docosahexaenoic acid (DHA) AA in AT phospholipids. Adipose tissue (AT) mRNA expression of chemokines (MCP  1, Fetui  A, CXCL14), marker genes for M1 and M2 macrophages (CD11c and CD206, respectively) and inflammatory markers were measured along with activation of NFkB, JNK, and STAT3. Macrophage infiltration into AT was examined using F4 per 80 immunohistochemistry. Results  Any therapeutic benefit produced by reducing the omega 6 omega 3 was evident only when comparing the 1 1 to 20 1 HFD; the 1 1 HFD resulted in a lower TC HDL  C and decreased AT CXCL14 gene expression and AT macrophage infiltration, which was linked to a higher EPA AA and DHA AA in AT phospholipids. However, despite these effects, and independent of the omega 6 omega 3, all HFDs, in general, led to similar levels of adiposity, insulin resistance, and AT inflammation. Conclusion  Reducing the omega 6 omega 3 using alpha linolenic acid is not an effective therapy for attenuating obesity and type II diabetes mellitus development. (Authors abstract)

While the absolute intake of dietary fat has undoubtedly contributed to the international expansion of obesity, the fatty acid (FA) composition of a high fat diets (HFD), independent of total dietary fat, can greatly impact metabolic and inflammatory processes.  Recently, significant attention has surrounded the dietary ratio of omega 6 polyunsaturated fatty acids (PUFAs) to omega 3 PUFAs (omega 6 omega 3) as it relates to chronic disease development. A high omega 6 omega 3 consumption has been associated with the promotion of many chronic diseases. Given this, it has been recommended that the omega 6 omega 3 consumption should be 4 to 11 to 1 as is commonly seen within the populations of wild animals and our human ancestors. Conversely, in the United States, the average consumption of omega 6 omega 3 is 10to 20 to 1. This study examined the influence of four tightly controlled, clinically applicable, HFDs (40 per cent  of total calories from dietary fat) differing solely in the omega 6 omega 3 (1 1, 5 1, 10 1, and 20 1) using only parent chain, plant based, omega 6 and omega 3 FAs (linoleic (LA; C18 2) and a linolenic acid (ALA; C18 3), respectively) on body composition, macrophage polarization, inflammation, and metabolic dysfunction in mice. Evidence indicates that omega 3 FAs have the potential to offset the detrimental effects of a HFD, including reduced adiposity and inflammation, and improve insulin sensitivity. The manipulation of the omega6 omega3 was in a diet that contained 40 per cent of total calories from fat, 12 per cent calories from saturated fat and a MUFA PUFA of 2 1. It utilized up to seven different sources of fat so that the total consumption of omega3 PUFAs in the diet fell within a clinically appropriate dose (4.7 per cent, 1.6 per cent , .86 per cent , and .46 per cent  of total calories from omega 3 PUFAs for the 1 1, 5 1, 10 1, and 20 1 diets, respectively; it has been reported that the typical American’s diet is composed of up to .2 to .7 per cent of total calories from omega 3 PUFAs ), and the only FA ratio which changed in the diet was the omega 6 omega 3 .  Although there are several means by which omega 3 FAs are thought to elicit their therapeutic effects on attenuating obesity development, the three primary mechanisms are as follows  1) enhanced ability of omega3 FAs to be oxidized rather than stored, 2) modulation of the phospholipid composition of the cellular membrane influencing the biosynthesis of inflammatory eicosanoids, and 3) ability to bind GPR120 and inhibit proinflammatory pathways. Of all unsaturated FAs, ALA is reported to be one of the most efficiently oxidized. After 20 weeks of HFD consumption, there was no difference in adipocyte size, total visceral AT mass, or body composition between any of the HFD groups. This may be due to the fact that LA is also an efficiently metabolized FA and any difference in the proficiency to metabolize ALA vs. LA or the absolute discrepancy in the consumption of ALA vs. LA was not sufficient enough to elicit a difference in adiposity. However, metabolic flux experiments would need to be performed in order to conclusively corroborate these findings. Independent of the omega 6 omega 3 and the percentage of total fat in the diet, all HFDs upregulated MCP1 and CXCL14 gene expression compared to the control diet.  However, CXCL14 mRNA was found to be lower in the 1 to 1 HFD compared to all other HFDs. Regarding macrophage polarization, all HFDs upregulated the gene expression of the general macrophage marker, F4/80, and the M1 (CD11c) and M2 (CD206) pro and anti inflammatory macrophage markers, respectively. However, in both the F4 per 80 and CD11c analyses, consumption of the 1 to 1 HFD resulted in significantly less mRNA of these markers than the 20 to 1 HFD. These findings should be substantiated using fluorescence activated cell sorting methodology to firmly establish the influence of manipulating the omega 6 omega 3 with ALA on macrophage infiltration and polarization during HFD induced obesity. Taken together, the findings regarding AT phospholipids composition and the outcomes related to macrophage polarization, inflammation, and insulin resistance suggest that changes in phospholipid composition leading to an increase in the EPA AA and/or DHA AA may lead to reduced macrophage infiltration, possibly via regulation of CXCL14, but does not necessarily reduce AT inflammation, as assessed by the inflammatory markers measured, nor hinder the development of insulin resistance. CXCL14 has been shown to be an efficient chemokine for PGE2  treated monocytes. However, despite the decreased infiltration of macrophages, there were no differences with respect to the inflammatory markers measured or insulin resistance. The results of this study do not suggest that ALA does not hold important physiological functions or therapeutic properties. In fact, ALA is an essential FA required for the endogenous synthesis of EPA and DHA. Further, studies
have shown that ALA may possess cardioprotective and antiinflammatory qualities. It is likely that ALA exhibits such qualities most distinctly when it is consumed in place of long chain saturated FAs, largely characterized as proinflammatory. In conclusion, this study suggests that reducing the omega6 omega3 solely with ALA is not an effective therapy for mitigating obesity nor type II diabetes mellitus development. It is likely that reducing the omega6 omega3 utilizing EPA and DHA may be a more effective strategy for attenuating obesity and related health complications, as EPA and DHA, in addition to being potent activators of GPR120, have the capability to be directly stored in phospholipids rather than having to compete for enzymes to be synthesized and then incorporated into the cellular membrane. However, an appropriately designed study needs to be conducted in order to substantiate such a hypothesis. (Editors comments)