Petra Tollet-Egnell

Metabolite profiles of body fluids or tissue extracts can be regarded as important indicators of physiological or pathological states. Whether hormone‐specific alterations of the serum metabolome can be identified using this technique has not been tested yet. The aim of this study was to investigate metabolic responses during hormone therapy in postmenopausal women by a nontargeted metabolomics approach. Sixty naturally postmenopausal women were randomly assigned to treatment with testosterone undecanoate 40 mg every second day; estradiol valerate 2 mg daily; or the combination of both. Serum metabolites were determined by gas chromatography‐mass spectrometry (GC‐MS) before and after 3 months of treatment. Metabolites affected by the treatment were identified and correlated with changes in insulin sensitivity and lipid profiles. Treatment‐dependent and hormone‐specific effects on serum metabolites were observed, ranging between 69% reduction and 184% increase, but the metabolites that best explained the differences could not be structurally identified. Effects on annotated metabolites were less associated with clinical parameters as compared to established serum markers for adverse lipid and carbohydrate metabolism, such as cholesterol and triglycerides. However, cystine, lysine and tyrosine were shown to change in correlation with insulin sensitivity and high‐density lipoprotein cholesterol levels in response to testosterone, indicating that those responses were somehow related to each other. Oestrogen‐ and androgen‐specific alterations in the serum metabolome could be identified using GC‐MS, reflecting hormone‐specific effects on whole body metabolism. New knowledge regarding steroid‐mediated metabolic responses within different tissues might be obtained using a similar approach on tissue extracts.

Male Zucker diabetic fatty (mZDF) rats spontaneously develop type 2 diabetes, whereas females only become diabetic when fed a diabetogenic high-fat diet (high-fat-fed female ZDF rat, HF-fZDF). The aim of this study was to investigate if differences in liver functions could provide clues to this sex difference. Non-diabetic obese fZDF rats were compared with either mZDF or HF-fZDF regarding hepatic molecular profiles, to single out those components that might be protective in the females. High-fat feeding in fZDF led to enhanced weight gain, increased blood glucose and insulin levels, reduced insulin sensitivity and a trend towards reduced glucose tolerance, indicative of a prediabetic state. mZDF rats were diabetic, with low levels of insulin, high levels of glucose, reduced insulin sensitivity and impaired glucose tolerance. Transcript profiling and capillary electrophoresis time-of-flight mass spectrometry were used to indentify hepatic transcripts and metabolites that might be related to this. Many diet-induced alterations in transcript and metabolite levels in female rats were towards a ‘male-like’ phenotype, including reduced lipogenesis, increased fatty acid (FA) oxidation and increased oxidative stress responses. Alterations detected at the level of hepatic metabolites, indicated lower capacity for glutathione (GSH) production in male rats, and higher GSH turnover in females. Taken together, this could be interpreted as if anabolic pathways involving lipogenesis and lipid output might limit the degree of FA oxidation and oxidative stress in female rats. Together with a greater capacity to produce GSH, these hepatic sex differences might contribute to the sex-different development of diabetes in ZDF rats.

Background: Genes involved in hepatic metabolism have a sex-different expression in rodents. To test whether male and female rat livers differ regarding lipid and carbohydrate metabolism, whole-genome transcript profiles were generated and these were complemented by measurements of hepatic lipid and glycogen content, fatty acid (FA) oxidation rates and hepatic glucose output (HGO). The latter was determined in perfusates from in situ perfusion of male and female rat livers. These perfusates were also analysed using nuclear magnetic resonance (NMR) spectroscopy to identify putative sex-differences in other liver-derived metabolites. Effects of insulin were monitored by analysis of Akt-phosphorylation, gene expression and HGO after s.c. insulin injections. Results: Out of approximately 3 500 gene products being detected in liver, 11% were significantly higher in females, and 11% were higher in males. Many transcripts for the production of triglycerides (TG), cholesterol and VLDL particles were female-predominant, whereas genes for FA oxidation, gluconeogenesis and glycogen synthesis were male-predominant. Sex-differences in mRNA levels related to metabolism were more pronounced during mild starvation (12 h fasting), as compared to the postabsorptive state (4 h fasting). No sex-differences were observed regarding hepatic TG content, FA oxidation rates or blood levels of ketone bodies or glucose. However, males had higher hepatic glycogen content and higher HGO, as well as higher ratios of insulin to glucagon levels. Based on NMR spectroscopy, liver-derived lactate was also higher in males. HGO was inhibited by insulin in parallel with increased phosphorylation of Akt, without any sex-differences in insulin sensitivity. However, the degree of Thr172-phosphorylated AMP kinase (AMPK) was higher in females, indicating a higher degree of AMPK-dependent actions. Conclusions: Taken together, males had higher ratios of insulin to glucagon levels, higher levels of glycogen, lower degree of AMPK phosphorylation, higher expression of gluconeogenic genes and higher hepatic glucose output. Possibly these sex-differences reflect a higher ability for the healthy male rat liver to respond to increased energy demands.

The aim of this study was to compare the effects of cocoa butter and safflower oil on hepatic transcript profiles, lipid metabolism and insulin sensitivity in healthy rats. Cocoa butter‐based high‐fat feeding for 3 days did not affect plasma total triglyceride (TG) levels or TG‐rich VLDL particles or hepatic insulin sensitivity, but changes in hepatic gene expression were induced that might lead to increased lipid synthesis, lipotoxicity, inflammation and insulin resistance if maintained. Safflower oil increased hepatic β‐oxidation, was beneficial in terms of circulating TG‐rich VLDL particles, but led to reduced hepatic insulin sensitivity. The effects of safflower oil on hepatic gene expression were partly overlapping with those exerted by cocoa butter, but fewer transcripts from anabolic pathways were altered. Increased hepatic cholesterol levels and increased expression of hepatic CYP7A1 and ABCG5 mRNA, important gene products in bile acid production and cholesterol excretion, were specific effects elicited by safflower oil only. Common effects on gene expression included increased levels of p8, DIG‐1 IGFBP‐1 and FGF21, and reduced levels of SCD‐1 and SCD‐2. This indicates that a lipid‐induced program for hepatic lipid disposal and cell survival was induced by 3 days of high‐fat feeding, independent on the lipid source. Based on the results, we speculate that hepatic TG infiltration leads to reduced expression of SCD‐1, which might mediate either neutral, beneficial or unfavorable effects on hepatic metabolism upon high‐fat feeding, depending on which fatty acids were provided by the diet.

MicroRNAs (miRNAs) are short non-coding RNAs playing an important role in post-transcriptional regulation of gene expression. We have previously shown that hepatic transcript profiles are different between males and females; that some of these differences are under the regulation of growth hormone (GH); and that mild starvation diminishes some of the differences. In this study, we tested if hepatic miRNAs are regulated in a similar manner.

CD36 is a multiligand receptor involved in various metabolic pathways, including cellular uptake of long-chain fatty acids. Defect function or expression of CD36 can result in dyslipidemia or insulin resistance. We have previously shown that CD36 expression is female-predominant in rat liver. In the present study, hormonal and nutritional regulation of hepatic CD36 expression was examined in male and female rats. Since alternative transcription start sites have been described in murine and human Cd36, we investigated whether alternative CD36 transcripts are differentially regulated in rat liver during these conditions.