Evidence indicates that principal features of the membrane involve structural organization of lipids in the form of a bilayer with functional proteins either bound to the bilayer surface or inserted into the bilayer and interacting within specific domains in the lipid milieux. In homeotherms, intrinsic and extrinsic factors apparently form the basis for determination of membrane lipid composition and thus membrane physicochemical properties. Moreover, many intrinsic metabolic controls, such as fatty acid desaturation and phospholipid biosynthesis, may be attenuated by change in the nature of the extrinsic or dietary influence. This review will focus on the role of dietary fat as a determinant of subcellular structural constituents to illustrate that feeding nutritionally adequate diets differing in fatty acid composition can induce physiological transitions in membrane function involving the activity of enzymes responsible for synthesis of membrane constituents, hormone-activated functions and expression of activity in the cell nucleus.

Lipids are important biomolecules in all biological systems and serve numerous essential cellular functions. The global analysis of complex lipids is very challenging due to the extreme diversity in lipid structures. Variation in linkages and positions of fatty acyl chain(s) on the lipid backbone, functional group modification, occurrence of the molecular species as isomers or isobars are among some of the greatest challenges to resolve in lipidomics. In this work, we describe a routine analytical approach combining two liquid chromatography platforms: hydrophilic interaction (HILIC) and C30 reversed-phase chromatography (C30RP) coupled to high resolution mass spectrometry (HRMS) as complementary high throughput platforms to analyze complex lipid mixtures. Vascular plants (kale leaves and corn roots), rat brain and soil microbes were used as proxies to evaluate the efficiency of the enhanced approach to resolve traditional, as well as, modified lipids during routine lipidomics analysis. We report for the first time, the observation of a modified class of acylphosphatidylglycerol (acylPG) in corn roots by HILIC, and further resolution of the isomers using C30RP chromatography. We also used this approach to demonstrate the presence of high levels of N-monomethyl phosphatidylethanolamine (MMPE) in soil microbes, as well as to determine the regioisomers of lysophospholipids in kale leaves. Additionally, neutral lipids were demonstrated using C30RP chromatography in positive ion mode to resolve triacylglycerol isomers in rat brain. The work presented here demonstrates how the enhanced approach can more routinely permit novel biomarker discovery, or lipid metabolism in a wide range of biological samples.

We determined if fatty acids can regulate the murine Cyp7a1 and human CYP7A1 gene promoters via peroxisome proliferator-activated receptor ␣ (PPAR␣)/9-cisretinoic acid receptor ␣ (RXR␣). In transfected cells, the murine Cyp7a1 gene promoter displayed markedly lower basal activity, but greater sensitivity to fatty acidor WY 14,643-activated PPAR␣/RXR␣ when compared with the human CYP7A1 gene promoter. PPAR␣/RXR␣ can bind to a site (Site II) located within the region at nucleotides ؊158 to ؊132 of both promoters. Mutagenesis of the human CYP7A1 Site II element abolished the response to activated PPAR␣/RXR␣. The murine Cyp7a1 gene promoter contains an additional PPAR␣/RXR␣-binding site (Site I) located within nucleotides ؊72 to ؊57. Replacement of a single residue in human CYP7A1 Site I with that found in the murine Cyp7a1 Site I sequence enabled PPAR␣/RXR␣ binding, and this mutation resulted in reduced basal activity, but substantially improved the response to activated PPAR␣/RXR␣ in transfected cells. We conclude that fatty acids can regulate the cyp7a gene promoter via PPAR␣/RXR␣. The differential response of the murine Cyp7a1 and human CYP7A1 gene promoters to PPAR␣ activators is attributable to the additional PPAR␣/RXR␣-binding site in the murine Cyp7a1 gene promoter.

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Dietary cholesterol has been shown to have a stimulatory effect on the murine cholesterol 7␣-hydroxylase gene (Cyp7a1), but its effect on human cholesterol 7␣-hydroxylase gene (CYP7A1) expression in vivo is not known. A transgenic mouse strain harboring the human CYP7A1 gene and homozygous for the disrupted murine Cyp7a1 gene was created. Cholesterol feeding increased the expression of the endogenous modified Cyp7a1 allele but failed to stimulate the human CYP7A1 transgene. In transfected hepatoma cells, 25-hydroxycholesterol increased murine Cyp7a1 gene promoter activity, whereas the human CYP7A1 gene promoter was unresponsive. Electrophoretic mobility shift assays demonstrated the interaction of the liver X receptor ␣ (LXR␣): retinoid X receptor (RXR) heterodimer, a transcription factor complex that is activated by oxysterols, with the murine Cyp7a1 gene promoter, whereas no binding to the human CYP7A1 gene promoter was detected. The results demonstrate that the human CYP7A1 gene is not stimulated by dietary cholesterol in the intact animal, and this is attributable to the inability of the CYP7A1 gene promoter to interact with LXR␣:RXR.

We have previously shown the anti-diabetic effects of swertiamarin; however, pharmacokinetic analysis showed that swertiamarin had a plasma half-life of 1.3 h. Gentianine is an active metabolite of swertiamarin that possesses a pharmacophoric moiety. The aim of this study was to explore the possibility whether the anti-diabetic effect of swertiamarin is due to gentianine. Swertiamarin treatment had no significant effect on adipogenesis, or the mRNA expression of PPAR-γ and GLUT-4; however, there was a significant increase in the mRNA expression of adiponectin. On the other hand, treatment with gentianine significantly increased adipogenesis, which was associated with a significant increase in the mRNA expression of PPAR-γ, GLUT-4 and adiponectin. These findings suggest, for the first time, that the anti-diabetic effect of swertiamarin is due to gentianine, an active metabolite of swertiamarin.

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