Programming effects of an early-life diet containing large phospholipid-coated lipid globules are transient under continuous exposure to a high-fat diet

Breastfeeding is associated with a lower risk of developing obesity during childhood and adulthood compared to feeding infant milk formula (IMF). Previous studies have shown that an experimental IMF (eIMF; comprising Nuturis®), programmed mouse pups for a lower body weight and fat mass gain in adulthood when challenged with a high-fat diet (HFD), compared to a control IMF (cIMF). Nuturis has a lipid composition and structure more similar to breastmilk. Here, the long-term effects were tested of a similar eIMF, but with an adapted lipid composition, and a cIMF, on body weight, glucose homeostasis, liver and adipose tissue. Nutrient composition was similar for the eIMF and cIMF; the lipid fractions comprised ∼50% milkfat. C57BL/6JOlaHsd mice were fed cIMF or eIMF from postnatal (PN) day 16-42 followed by a HFD until PN168. Feeding eIMF versus cIMF in early life resulted in a lower body weight (-9%) and body fat deposition (-14%) in adulthood (PN105). The effect appeared transient, as from PN126 onward, after 12 weeks HFD, eIMF-fed mice caught up on controls and body and fat weights became comparable between groups. Glucose and energy metabolism were similar between groups. At dissection (PN168), eIMF-fed mice showed larger (+27%) epididymal fat depots and a lower (-26%) liver weight without clear morphological aberrations. Our data suggest the size and coating but not the lipid composition of IMF fat globules underlies the programming effect observed. Prolonged exposure to a HFD challenge partly overrules the programming effect of early diet

An early-life diet containing large phospholipid-coated lipid globules programs later-life postabsorptive lipid trafficking in high-fat diet but not in low-fat dietfed mice

Feeding mice in early-life a diet containing an experimental infant milk formula (Nuturis®; eIMF), with a lipid structure similar to human milk, transiently lowered body weight and fat mass gain upon Western-style diet later in life, when compared to mice fed diets based on control IMF (cIMF). We tested the hypothesis that early-life eIMF feeding alters the absorption or the postabsorptive trafficking of dietary lipids in later-life. Male C57BL/6JOlaHsd mice were fed eIMF/cIMF from postnatal day 16-42, followed by low- (LFD, AIN-93G, 7wt% fat) or high-fat diet (HFD, D12451, 24wt% fat) until day 63-70. Lipid absorption rate and tissue concentrations were determined after intragastric administration of stable isotope (deuterium or 13C) labelled lipids in separate groups. Lipid enrichments in plasma and tissues were analysed using gas chromatography-mass spectrometry. The rate of triolein absorption was similar between eIMF and cIMF fed LFD: 3.2 SD 1.8 and 3.9 SD 2.1 and HFD: 2.6 SD 1.7 and 3.8 SD 3.0 %dose.ml-1.h-1. Postabsorptive lipid trafficking, i.e., concentrations of absorbed lipids in tissues, was similar in the eIMF and cIMF groups after LFD. Tissue levels of absorbed triglycerides after HFD-feeding were lower in heart (-42%) and liver (-46%), and higher in muscle (+81%, all p<0.05) in eIMF-fed mice. In conclusion, early-life IMF diet affected postabsorptive trafficking of absorbed lipids after HFD, but not LFD. Changes in postabsorptive lipid trafficking could underlie the observed lower body weight and body fat accumulation in later life upon a persistent long-term obesogenic challenge

Effects of an early life diet containing large phospholipid-coated lipid globules on hepatic lipid metabolism in mice

We recently reported that feeding mice in their early life a diet containing a lipid structure more similar to human milk (eIMF, Nuturis) results in lower body weights and fat mass gain upon high fat feeding in later life, compared to control (cIMF). To understand the underlying mechanisms, we now explored parameters possibly involved in this long-term effect. Male C57BL/6JOlaHsd mice, fed rodent diets containing eIMF or cIMF from postnatal (PN) day 16–42, were sacrificed at PN42. Hepatic proteins were measured using targeted proteomics. Lipids were assessed by LC–MS/MS (acylcarnitines) and GC-FID (fatty-acyl chain profiles). Early life growth and body composition, cytokines, and parameters of bile acid metabolism were similar between the groups. Hepatic concentrations of multiple proteins involved in β-oxidation (+ 17%) the TCA cycle (+ 15%) and mitochondrial antioxidative proteins (+ 28%) were significantly higher in eIMF versus cIMF-fed mice (p < 0.05). Hepatic l-carnitine levels, required for fatty acid uptake into the mitochondria, were higher (+ 33%, p < 0.01) in eIMF-fed mice. The present study indicates that eIMF-fed mice have higher hepatic levels of proteins involved in fatty acid metabolism and oxidation. We speculate that eIMF feeding programs the metabolic handling of dietary lipids

Common arterial trunk and ventricular non-compaction in Lrp2 knockout mice indicate a crucial role of LRP2 in cardiac development

Lipoprotein-related receptor protein 2 (LRP2) is important for development of the embryonic neural crest and brain in both mice and humans. Although a role in cardiovascular development can be expected, the hearts of Lrp2 knockout (KO) mice have not yet been investigated. We studied the cardiovascular development of Lrp2 KO mice between embryonic day 10.5 (E10.5) and E15.5, applying morphometry and immunohistochemistry, using antibodies against Tfap2α (neural crest cells), Nkx2.5 (second heart field), WT1 (epicardium derived cells), tropomyosin (myocardium) and LRP2. The Lrp2 KO mice display a range of severe cardiovascular abnormalities, including aortic arch anomalies, common arterial trunk (persistent truncus arteriosus) with coronary artery anomalies, ventricular septal defects, overriding of the tricuspid valve and marked thinning of the ventricular myocardium. Both the neural crest cells and second heart field, which are essential for the lengthening and growth of the right ventricular outflow tract, are abnormally positioned in the Lrp2 KO. T hi s explains the absence of the aorto-pulmonary septum, which leads to common arterial trunk and ventricular septal defects. Severe blebbing of the epicardial cells covering the ventricles is seen. Epithelial-mesenchymal transition does occur; however, there are fewer WT1-positive epicardium-derived cells in the ventricular wall as compared to normal, coinciding with the myocardial thinning and deep intertrabecular spaces. LRP2 plays a crucial role in cardiovascular development in mice. This corroborates findings of cardiac anomalies in humans with LRP2 mutations. Future studies should reveal the underlying signaling mechanisms in which LRP2 is involved during cardiogenesis

Spontaneous liver disease in wild-type C57BL/6JOlaHsd mice fed semisynthetic diet

Mouse models are frequently used to study mechanisms of human diseases. Recently, we observed a spontaneous bimodal variation in liver weight in C57BL/6JOlaHsd mice fed a semisynthetic diet. We now characterized the spontaneous variation in liver weight and its relationship with parameters of hepatic lipid and bile acid (BA) metabolism. In male C57BL/6JOlaHsd mice fed AIN-93G from birth to postnatal day (PN)70, we measured plasma BA, lipids, Very low-density lipoprotein (VLDL)-triglyceride (TG) secretion, and hepatic mRNA expression patterns. Mice were sacrificed at PN21, PN42, PN63 and PN70. Liver weight distribution was bimodal at PN70. Mice could be subdivided into two nonoverlapping groups based on liver weight: 0.6 SD 0.1 g (approximately one-third of mice, small liver; SL), and 1.0 SD 0.1 g (normal liver; NL; p<0.05). Liver histology showed a higher steatosis grade, inflammation score, more mitotic figures and more fibrosis in the SL versus the NL group. Plasma BA concentration was 14-fold higher in SL (p<0.001). VLDL-TG secretion rate was lower in SL mice, both absolutely (-66%, p<0.001) and upon correction for liver weight (-44%, p<0.001). Mice that would later have the SL-phenotype showed lower food efficiency ratios during PN21-28, suggesting the cause of the SL phenotype is present at weaning (PN21). Our data show that approximately one-third of C57BL/6JOlaHsd mice fed semisynthetic diet develop spontaneous liver disease with aberrant histology and parameters of hepatic lipid, bile acid and lipoprotein metabolism. Study designs involving this mouse strain on semisynthetic diets need to take the SL phenotype into account. Plasma lipids may serve as markers for the identification of the SL phenotype

The Origin of Follicular Bile Acids in the Human Ovary

Bile acids (BAs) are present in ovarian follicular fluid (FF) and are linked to embryo development. However, information on the source of ovarian BA is scarce. Therefore, we aimed to explore local ovarian synthesis and BA transport from blood into FF. BA levels were determined in matching FF and serum from women undergoing in vitro fertilization. In vitro BA production by human mural granulosa cells (MGCs) and cumulus granulosa cells (CGCs) was measured by mass spectrometry. Gene and protein expression were quantified in MGC and CGC and in human ovarian tissue by quantitative PCR and Western blot/immunohistochemistry, respectively. BA levels in blood and FF were significantly correlated (r(s) = 0.186, P = 0.027) but were almost twofold higher in FF (P <0.001). Primary BA levels were increased in FF, indicating that, in addition to passive diffusion, other sources of ovarian BA might exist. The key BA synthesis enzyme cytochrome P450 A1 was absent in MGC and CGC; BA production in vitro was undetectable. Therefore, local ovarian BA production is unlikely. However, common BA importers (Na+/taurocholate cotransporting polypeptide, apical sodium-dependent bile acid transporter) and an exporter (ATP-binding cassette subfamily C member 3) were identified in GC, theca cells, and oocyte. In summary, these results suggest that passive and active transport of BAs from blood into FF constitute sources of FF BA

FXR overexpression alters adipose tissue architecture in mice and limits its storage capacity leading to metabolic derangements

The bile acid-activated nuclear receptor, FXR (NR1H4), has been implicated in the control of lipid and energy metabolism, but its role in fat tissue, where it is moderately expressed, is not understood. In view of the recent development of FXR-targeting therapeutics for treatment of human metabolic diseases, understanding the tissue-specific actions of FXR is essential. Transgenic mice expressing human FXR in adipose tissue (aP2-hFXR mice) at three to five times higher levels than endogenous Fxr, i.e., much lower than its expression in liver and intestine, have markedly enlarged adipocytes and show extensive extracellular matrix remodeling. Ageing and exposure to obesogenic conditions revealed a strongly limited capacity for adipose expansion and development of fibrosis in adipose tissues of aP2-hFXR transgenic mice. This was associated with impaired lipid storage capacity, leading to elevated plasma free fatty acids and ectopic fat deposition in liver and muscle as well as wholebody insulin resistance. These studies establish that adipose FXR is a determinant of adipose tissue architecture and contributes to whole-body lipid homeostasis

Transcriptome analysis suggests a compensatory role of the cofactors coenzyme A and NAD+ in medium-chain acyl-CoA dehydrogenase knockout mice

During fasting, mitochondrial fatty-acid β-oxidation (mFAO) is essential for the generation of glucose by the liver. Children with a loss-of-function deficiency in the mFAO enzyme medium-chain acyl-Coenzyme A dehydrogenase (MCAD) are at serious risk of life-threatening low blood glucose levels during fasting in combination with intercurrent disease. However, a subset of these children remains asymptomatic throughout life. In MCAD-deficient (MCAD-KO) mice, glucose levels are similar to those of wild-type (WT) mice, even during fasting. We investigated if metabolic adaptations in the liver may underlie the robustness of this KO mouse. WT and KO mice were given a high- or low-fat diet and subsequently fasted. We analyzed histology, mitochondrial function, targeted mitochondrial proteomics, and transcriptome in liver tissue. Loss of MCAD led to a decreased capacity to oxidize octanoyl-CoA. This was not compensated for by altered protein levels of the short- and long-chain isoenzymes SCAD and LCAD. In the transcriptome, we identified subtle adaptations in the expression of genes encoding enzymes catalyzing CoA- and NAD(P)(H)-involving reactions and of genes involved in detoxification mechanisms. We discuss how these processes may contribute to robustness in MCAD-KO mice and potentially also in asymptomatic human subjects with a complete loss of MCAD activity