Rosiglitazone Metabolism in Human Liver Microsomes Using a Substrate Depletion Method

Background: Elimination of rosiglitazone in humans is via hepatic metabolism. The existing studies suggest that CYP2C8 is the major enzyme responsible, with a minor contribution from CYP2C9; however, other studies suggest the involvement of additional cytochrome P450 enzymes and metabolic pathways. Thus a full picture of rosiglitazone metabolism is unclear. Objective: This study aimed to improve the current understanding of potential drug–drug interactions and implications for therapy by evaluating the kinetics of rosiglitazone metabolism and examining the impact of specific inhibitors on its metabolism using the substrate depletion method. Methods: In vitro oxidative metabolism of rosiglitazone in human liver microsomes obtained from five donors was determined over a 0.5–500 µM substrate range including the contribution of CYP2C8, CYP2C9, CYP3A4, CYP2E1, and CYP2D6. Results: The maximum reaction velocity was 1.64 ± 0.98 nmol·mg−1·min−1. The CYP2C8 (69 ± 20%), CYP2C9 (42 ± 10%), CYP3A4 (52 ± 23%), and CEP2E1 (41 ± 13%) inhibitors all significantly inhibited rosiglitazone metabolism. Conclusion: The results suggest that other cytochrome P450 enzymes, including CYP2C9, CYP3A4, and CEP2E1, in addition to CYP28, also play an important role in the metabolism of rosiglitazone. This example demonstrates that understanding the complete metabolism of a drug is important when evaluating the potential for drug–drug interactions and will assist to improve the current therapeutic strategies.</p

Expression of cholesterol packaging and transport genes in human and rat placenta: impact of obesity and a high-fat diet

Evidence suggests that sub-optimal maternal nutrition has implications for the developing offspring. We have previously shown that exposure to a low-protein diet during gestation was associated with upregulation of genes associated with cholesterol transport and packaging within the placenta. This study aimed to elucidate the effect of altering maternal dietary linoleic acid (LA; omega-6) to alpha-linolenic acid (ALA; omega-6) ratios as well as total fat content on placental expression of genes associated with cholesterol transport. The potential for maternal body mass index (BMI) to be associated with expression of these genes in human placental samples was also evaluated. Placentas were collected from 24 Wistar rats at 20-day gestation (term = 21–22-day gestation) that had been fed one of four diets containing varying fatty acid compositions during pregnancy, and from 62 women at the time of delivery. Expression of 14 placental genes associated with cholesterol packaging and transfer was assessed in rodent and human samples by quantitative real time polymerase chain reaction. In rats, placental mRNA expression of ApoA2, ApoC2, Cubn, Fgg, Mttp and Ttr was significantly elevated (3–30 fold) in animals fed a high LA (36% fat) diet, suggesting increased cholesterol transport across the placenta in this group. In women, maternal BMI was associated with fewer inconsistent alterations in gene expression. In summary, sub-optimal maternal nutrition is associated with alterations in the expression of genes associated with cholesterol transport in a rat model. This may contribute to altered fetal development and potentially programme disease risk in later life. Further investigation of human placenta in response to specific dietary interventions is required

The effect of maternal dietary fat content and omega-6 to omega-3 ratio on offspring growth and hepatic gene expression in the rat

© The Authors 2020. Omega-6 fatty acids have been shown to exert pro-adipogenic effects whereas omega-3 fatty acids work in opposition. Increasing intakes of LA (linoleic acid; omega-6) vs ALA (alpha-linolenic acid; omega-3) in Western diets has led to the hypothesis that consumption of this diet during pregnancy may be contributing to adverse offspring health. This study investigated the effects of feeding a maternal dietary LA:ALA ratio similar to that of the Western diet (9:1) compared to a proposed 'ideal' ratio (∼1:1.5), at two total fat levels (18% vs 36% fat w/w), on growth and lipogenic gene expression in the offspring. Female Wistar rats were assigned to one of the four experimental groups throughout gestation and lactation. Offspring were culled at 1 and 2 weeks of age for sample collection. Offspring of dams consuming a -36% fat diet were ∼20% lighter than those exposed to a 18% fat diet (

Temperature and time-dependent effects of delayed blood processing on oxylipin concentrations in human plasma.

BACKGROUND:Oxidized derivatives of polyunsaturated fatty acids, collectively known as oxylipins, are labile bioactive mediators with diverse roles in human physiology and pathology. Oxylipins are increasingly being measured in plasma collected in clinical studies to investigate biological mechanisms and as pharmacodynamic biomarkers for nutrient-based and drug-based interventions. Whole blood is generally stored either on ice or at room temperature prior to processing. However, the potential impacts of delays in processing, and of temperature prior to processing, on oxylipin concentrations are incompletely understood. OBJECTIVE:To evaluate the effects of delayed processing of blood samples in a timeframe that is typical of a clinical laboratory setting, using typical storage temperatures, on concentrations of representative unesterified oxylipins measured by liquid chromatography-tandem mass spectrometry. DESIGN:Whole blood (drawn on three separate occasions from a single person) was collected into 5 mL purple-top potassium-EDTA tubes and stored for 0, 10, 20, 30, 60 or 120 min at room temperature or on wet ice, followed by centrifugation at 4 °C for 10 min with plasma collection. Each sample was run in duplicate, therefore there were six tubes and up to six data points at each time point for each oxylipin at each condition (ice/room temperature). Representative oxylipins derived from arachidonic acid, docosahexaenoic acid, and linoleic acid were quantified by liquid chromatography tandem mass spectrometry. Longitudinal models were used to estimate differences between temperature groups 2 h after blood draw. RESULTS:We found that most oxylipins measured in human plasma in traditional potassium-EDTA tubes are reasonably stable when stored on ice for up to 2 h prior to processing, with little evidence of auto-oxidation in either condition. By contrast, in whole blood stored at room temperature, substantial time-dependent increases in the 12-lipoxygenase-derived (12-HETE, 14-HDHA) and platelet-derived (thromboxane B2) oxylipins were observed. CONCLUSION:These findings suggest that certain plasma oxylipins can be measured with reasonable accuracy despite delayed processing for up to 2 h when blood is stored on ice prior to centrifugation. 12-Lipoxygenase- and platelet-derived oxylipins may be particularly sensitive to post-collection artifact with delayed processing at room temperature. Future studies are needed to determine impacts of duration and temperature of centrifugation on oxylipin concentrations

Recent developments on the role of epigenetics in obesity and metabolic disease

The increased prevalence of obesity and related comorbidities is a major public health problem. While genetic factors undoubtedly play a role in determining individual susceptibility to weight gain and obesity, the identified genetic variants only explain part of the variation. This has led to growing interest in understanding the potential role of epigenetics as a mediator of gene-environment interactions underlying the development of obesity and its associated comorbidities. Initial evidence in support of a role of epigenetics in obesity and type 2 diabetes mellitus (T2DM) was mainly provided by animal studies, which reported epigenetic changes in key metabolically important tissues following high-fat feeding and epigenetic differences between lean and obese animals and by human studies which showed epigenetic changes in obesity and T2DM candidate genes in obese/diabetic individuals. More recently, advances in epigenetic methodologies and the reduced cost of epigenome-wide association studies (EWAS) have led to a rapid expansion of studies in human populations. These studies have also reported epigenetic differences between obese/T2DM adults and healthy controls and epigenetic changes in association with nutritional, weight loss, and exercise interventions. There is also increasing evidence from both human and animal studies that the relationship between perinatal nutritional exposures and later risk of obesity and T2DM may be mediated by epigenetic changes in the offspring. The aim of this review is to summarize the most recent developments in this rapidly moving field, with a particular focus on human EWAS and studies investigating the impact of nutritional and lifestyle factors (both pre- and postnatal) on the epigenome and their relationship to metabolic health outcomes. The difficulties in distinguishing consequence from causality in these studies and the critical role of animal models for testing causal relationships and providing insight into underlying mechanisms are also addressed. In summary, the area of epigenetics and metabolic health has seen rapid developments in a short space of time. While the outcomes to date are promising, studies are ongoing, and the next decade promises to be a time of productive research into the complex interactions between the genome, epigenome, and environment as they relate to metabolic disease.Susan J. van Dijk, Ross L. Tellam, Janna L. Morrison, Beverly S. Muhlhausler, and Peter L. Mollo

DHA supplementation during pregnancy does not reduce BMI or body fat mass in children: follow-up of the DHA to Optimize Mother Infant Outcome randomized controlled trial

First published March 30, 2016The omega-3 (n-3) long-chain polyunsaturated fatty acid (LCPUFA) docosahexaenoic acid (DHA) has proven effective at reducing fat storage in animal studies. However, a systematic review of human trials showed a lack of quality data to support or refute this hypothesis.We sought to determine whether maternal DHA supplementation during the second half of pregnancy results in a lower body mass index (BMI) and percentage of body fat in children.We conducted a follow-up at 3 and 5 y of age of children who were born to mothers enrolled in the DOMInO (DHA to Optimize Mother Infant Outcome) double-blind, randomized controlled trial, in which women with a singleton pregnancy were provided with DHA-rich fish-oil capsules (800 mg DHA/d) or vegetable-oil capsules (control group) in the second half of pregnancy. Primary outcomes were the BMIzscore and percentage of body fat at 3 and 5 y of age. Potential interactions between prenatal DHA and the peroxisome proliferator-activated receptor-γ (PPARγ) genotype as a measure of the genetic predisposition to obesity were investigated.A total of 1614 children were eligible for the follow-up. Parent or caregiver consent was obtained for 1531 children (95%), and these children were included in the analysis. BMIzscores and percentages of body fat of children in the DHA group did not differ from those of children in the control group at either 3 y of age [BMIzscore adjusted mean difference: 0.03 (95% CI: -0.07, 0.13;P= 0.61); percentage of body fat adjusted mean difference: -0.26 (95% CI: -0.99, 0.46;P= 0.47)] or 5 y of age [BMIzscore adjusted mean difference: 0.02 (95% CI: -0.08, 0.12;P= 0.66); percentage of body fat adjusted mean difference: 0.11 (95% CI: -0.60, 0.82;P= 0.75)]. No treatment effects were modified by thePPARγgenotype of the child.Independent of a genetic predisposition to obesity, maternal intake of DHA-rich fish oil during the second half of pregnancy does not affect the growth or body composition of children at 3 or 5 y of age. This trial was registered atwww.anzctr.org.auas ACTRN1260500056906 and ACTRN12611001127998.Beverly S Muhlhausler, Lisa N Yelland, Robyn McDermott, Linda Tapsell, Andrew McPhee, Robert A Gibson, and Maria Makride

Acute changes to breast milk composition following consumption of high-fat and high-sugar meals

© 2021 The Authors. Maternal & Child Nutrition published by John Wiley & Sons Ltd. Breast milk composition is influenced by habitual diet, yet little is known about the short-term effects of changes in maternal diet on breast milk macronutrient concentrations. Our aim was to determine the acute effect of increased consumption of sugar/fat on breast milk protein, lactose and lipids. Exclusively breastfeeding women (n = 9) were provided with a control, higher fat (+28 g fat) and higher sugar (+66 g sugar) diet over three separate days at least 1 week apart. Hourly breast milk samples were collected concurrently for the analysis of triglycerides, cholesterol, protein, and lactose concentrations. Breast milk triglycerides increased significantly following both the higher fat and sugar diet with a greater response to the higher sugar compared to control diet (mean differences of 3.05 g/dL ± 0.39 and 13.8 g/dL ± 0.39 in higher fat and sugar diets, respectively [P < 0.001]). Breast milk cholesterol concentrations increased most in response to the higher sugar diet (0.07 g/dL ± 0.005) compared to the control (0.04 g/dL) and the higher fat diet (0.05 g/dL) P < 0.005. Breast milk triglyceride and lactose concentrations increased (P < 0.001, P = 0.006), whereas protein decreased (p = 0.05) in response to the higher fat diet compared to the control. Independent of diet, there were significant variations in breast milk composition over the day; triglycerides and cholesterol concentrations were higher at end of day (P < 0.001), whereas protein and lactose concentrations peaked at Hour 10 (of 12) (P < 0.001). In conclusion, controlled short-term feeding to increase daily sugar/fat consumption altered breast milk triglycerides, cholesterol, protein and lactose. The variations observed in breast milk protein and lactose across the 12 h period is suggestive of a circadian rhythm

Whole animal experiments should be more like human randomized controlled trials

Beverly S. Muhlhausler, Frank H. Bloomfield, Matthew W. Gillma

Placental glucocorticoid receptor isoforms in a sheep model of maternal allergic asthma

Maternal asthma increases the risk of adverse pregnancy outcomes and may affect fetal growth and placental function by differential effects on the expression of glucocorticoid receptor (GR) isoforms, leading to altered glucocorticoid signalling. Our aim was to examine the effect of maternal asthma on placental GR profiles using a pregnant sheep model of asthma. Nine known GR isoforms were detected. There was a significant increase in the expression of placental GR isoforms that are known to have low trans-activational activity in other species including GR A, GR P and GRγ which may result in a pro-inflammatory environment in the presence of allergic asthma