Calcium-Sensing Receptors Control CYP27B1-Luciferase Expression: Transcriptional and Posttranscriptional Mechanisms.

25-hydroxyvitamin D 1α-hydroxylase (encoded by CYP27B1), which catalyzes the synthesis of 1,25-dihydroxyvitamin D3, is subject to negative or positive modulation by extracellular Ca2+ (Ca2+ o) depending on the tissue. However, the Ca2+ sensors and underlying mechanisms are unidentified. We tested whether calcium-sensing receptors (CaSRs) mediate Ca2+ o-dependent control of 1α-hydroxylase using HEK-293 cells stably expressing the CaSR (HEK-CaSR cells). In HEK-CaSR cells, but not control HEK-293 cells, cotransfected with reporter genes for CYP27B1-Photinus pyralis (firefly) luciferase and control Renilla luciferase, an increase in Ca2+ o from 0.5mM to 3.0mM induced a 2- to 3-fold increase in firefly luciferase activity as well as mRNA and protein levels. Surprisingly, firefly luciferase was specifically suppressed at Ca2+ o ≥ 5.0mM, demonstrating biphasic Ca2+ o control. Both phases were mediated by CaSRs as revealed by positive and negative modulators. However, Ca2+ o induced simple monotonic increases in firefly luciferase and endogenous CYP27B1 mRNA levels, indicating that the inhibitory effect of high Ca2+ o was posttranscriptional. Studies with inhibitors and the CaSR C-terminal mutant T888A identified roles for protein kinase C (PKC), phosphorylation of T888, and extracellular regulated protein kinase (ERK)1/2 in high Ca2+ o-dependent suppression of firefly luciferase. Blockade of both PKC and ERK1/2 abolished Ca2+ o-stimulated firefly luciferase, demonstrating that either PKC or ERK1/2 is sufficient to stimulate the CYP27B1 promoter. A key CCAAT box (-74 bp to -68 bp), which is regulated downstream of PKC and ERK1/2, was required for both basal transcription and Ca2+ o-mediated transcriptional upregulation. The CaSR mediates Ca2+ o-dependent transcriptional upregulation of 1α-hydroxylase and an additional CaSR-mediated mechanism is identified by which Ca2+ o can promote luciferase and possibly 1α-hydroxylase breakdown.

Associations between CD36 gene polymorphisms and metabolic response to a short-term endurance-training program in a young-adult population.

Recent studies have shown that CD36 gene variants are associated with an increased prevalence of chronic disease. Although a genetic component to trainability has been proven, no data are available specifically on the influence of CD36 on training response. Two single nucleotide polymorphisms (SNPs) (rs1527479 and rs1984112) were assessed for associations with whole-body substrate oxidation, response to a 75-g dextrose oral glucose tolerance test, fasting plasma lipids, and cardiovascular disease risk factors in a young healthy cohort, both using cross-sectional analysis and following a 4-week endurance-exercise training program. Genotyping was performed using real-time polymerase chain reaction. Cross-sectional data were collected in 34 individuals (age, 22.7 ± 3.5 years), with 17 completing the training program. At baseline, TT SNP carriers at rs1527479 and wild-type GG carriers at rs1984112 were associated with significantly greater whole-body rate of fat oxidation (Fatox) during submaximal exercise (P < 0.05), whilst AA carriers at the same position were associated with elevated triglyceride (TG) levels. A significant genotype × time interaction in Fatox at SNP rs1984112 was identified at rest. Significant genotype × time interactions were present at rs1527479, with TT carriers exhibiting a favourable response to training when compared with C-allele carriers for fasting TG, diastolic blood pressure (DBP), and mean arterial pressure (MAP). In conclusion, cross-sectional assessment identified associations with Fatox and TG. Training response at both SNPs identified "at-risk" genotypes responding favourably to the training stimulus in Fatox, TG, DBP, and MAP. Although these data show potential pleiotropic influence of CD36 SNPs, assessment in a larger cohort is warranted.

Interactions Between Fatty Acid Transport Proteins, Genes That Encode for Them, and Exercise: A Systematic Review.

Long-chain fatty acid (LCFA) movement into skeletal muscle involves a highly mediated process in which lipid rafts are utilized in the cellular membrane, involving numerous putative plasma membrane-associated LCFA transport proteins. The process of LCFA uptake and oxidation is of particular metabolic significance both at rest and during light to moderate exercise. A comprehensive systematic search of electronic databases was conducted to investigate whether exercise alters protein and/or gene expression of putative LCFA transport proteins. There were 31 studies meeting all eligibility criteria, of these 13 utilized an acute exercise protocol and 18 examined chronic exercise adaptations. Seventeen involved a study design incorporating an exercise stimulus, while the remaining 14 incorporated a combined exercise and diet stimulus. Divergent data relating to acute exercise, as well as prolonged exercise training (≥3 weeks), on protein content (PC) response was identified for proteins CD36, FABPpm and CAV1. Messenger ribonucleic acid (mRNA) data did not always correspond to functional PC, supporting previous suggestions of a disconnect due to potentially limiting factors post gene expression. The large array of study designs, cohorts, and primary dependent variables within the studies included in the present review elucidate the complexity of the interaction between exercise and LCFA transport proteins. Summary of the results in the present review validate the need for further targeted investigation within this topic, and provide an important information base for such research. J. Cell. Physiol. 231: 1671-1687, 2016. © 2015 Wiley Periodicals, Inc.

The associations between polymorphisms in the CD36 gene, fat oxidation and cardiovascular disease risk factors in a young adult Australian population: a pilot study.

Our pilot study in a young adult Australian cohort aimed to investigate potential associations between CD36 polymorphisms (rs1527479 and rs1984112), fat oxidation and cardiovascular disease risk. CD36 genotype was associated with fat oxidation during sub-maximal exercise, resting heart rate and blood pressure, indicating increased chronic disease risk in this otherwise healthy cohort.

Effects of dietary protein to carbohydrate balance on energy intake, fat storage, and heat production in mice.

OBJECTIVE: Protein leverage plays a role in driving increased energy intakes that may promote weight gain. The influence of the protein to carbohydrate ratio (P:C) in diets of C57BL/6J mice on total energy intake, fat storage, and thermogenesis was investigated. DESIGN AND METHODS: Male mice (9 weeks old) were provided ad libitum access to one of five isocaloric diets that differed in P:C. Food intake was recorded for 12 weeks. After 16 weeks, white adipose tissue (WAT) and brown adipose tissue (BAT) deposits were dissected, weighed, and the expression levels of key metabolic regulators were determined in BAT. In a separate cohort, body surface temperature was measured in response to 25 diets differing in protein, fat, and carbohydrate content. RESULTS: Mice on low P:C diets (9:72 and 17:64) had greater total energy intake and increased WAT and BAT stores. Body surface temperature increased with total energy intake and with protein, fat, and carbohydrate, making similar contributions per kJ ingested. Expression of three key regulators of thermogenesis were downregulated in BAT in mice on the lowest P:C diet. CONCLUSIONS: Low-protein diets induced sustained hyperphagia and a generalized expansion of fat stores. Increased body surface temperature on low P:C diets was consistent with diet-induced thermogenesis (DIT) as a means to dissipate excess ingested energy on such diets, although this was not sufficient to prevent development of increased adiposity. Whether BAT was involved in DIT is not clear. Increased BAT mass on low P:C diets might suggest so, but patterns of thermogenic gene expression do not support a role for BAT in DIT, although they might reflect failure of thermogenic function with prolonged exposure to a low P:C diet.

Dietary fiber intake increases the risk of zinc deficiency in healthy and diabetic women.

Phytic acid is a major determinant of zinc bioavailability. Little is known about phytic acid intakes or indices of zinc bioavailability in type 2 diabetes mellitus (DM), a condition that predisposes to zinc deficiency. The aim of this cross-sectional study was to measure and explore the relationships among phytic acid intake, zinc bioavailability, and molecular markers of zinc homeostasis in 20 women with DM compared to 20 healthy women. The phytate/zinc, (calcium)(phytate)/zinc, and (calcium + magnesium)(phytate)/zinc molar ratios were used to indicate zinc bioavailability. Plasma zinc concentrations and zinc transporter (ZnT1, ZnT8, and Zip1) gene expression in mononuclear cells were measured. Participants with DM consumed 1,194 ± 824 mg/day (mean ± SD) phytic acid, an amount similar to the intake of healthy women (1,316 ± 708 mg/day). Bread products and breakfast cereals contributed more than 40 % of the phytic acid intake in each group. A positive relationship was observed in all participants between phytic acid and dietary fiber (r = 0.6, P < 0.001) and between dietary fiber and the (calcium)(phytate)/zinc ratio (r = 0.5, P < 0.001). Compared to the healthy group, the messenger RNA ratio of ZnT1 (zinc export) to Zip1 (zinc import) was lower in participants with DM, which may indicate perturbed zinc homeostasis in the disorder. The plasma zinc concentration was not predicted by age, body mass index, health status, zinc bioavailability, or zinc transporter expression. Healthy and diabetic women consume phytic acid in amounts that are likely to decrease the bioavailability of dietary zinc. Recommendations to consume greater amounts of dietary fiber, much of which is associated with phytate, increase the risk of zinc deficiency.

High-glycemic index carbohydrate increases nuclear factor-kappaB activation in mononuclear cells of young, lean healthy subjects.

BACKGROUND: High-glycemic index diets have been linked to greater risk of cardiovascular disease and type 2 diabetes. Postprandial glycemia within the normal range may promote oxidative stress and inflammatory processes underlying the development of disease. OBJECTIVE: We explored acute differences in the activation of the inflammatory marker nuclear factor-kappaB after consumption of 2 carbohydrate meals matched for macronutrient and micronutrient composition but differing in glycemic index. DESIGN: After an overnight fast, 10 young, lean healthy subjects were fed in random order 3 meals providing 50 g of available carbohydrate as glucose, white bread, or pasta. Venous blood samples were collected at 0, 1, 2, and 3 h, and nuclear proteins were extracted from mononuclear cells. Changes in nuclear factor-kappaB-p65 proteins were detected by Western blotting. Acute changes in other markers of oxidative stress (nitrotyrosine and soluble intercellular adhesion molecule-1) were also assessed. RESULTS: The maximum increase in nuclear factor-kappaB activation was similar after the bread meal [mean (+/-SEM) area under the curve: 69 +/- 16% optical density x h] and the glucose challenge (75 +/- 9% optical density x h), but was 3 times higher than after the pasta meal (23 +/- 5% optical density x h) (P < 0.05). Similarly, changes in nitrotyrosine, but not soluble intercellular adhesion molecule-1, were higher after glucose and bread than after pasta (P = 0.01 at 2 h). CONCLUSIONS: The findings suggest that high-normal physiologic increases in blood glucose after meals aggravate inflammatory processes in lean, young adults. This mechanism may help to explain relations between carbohydrates, glycemic index, and the risk of chronic disease.

Insulin resistance does not influence gene expression in skeletal muscle.

Insulin resistance is commonly observed in patients prior to the development of type 2 diabetes and may predict the onset of the disease. We tested the hypothesis that impairment in insulin stimulated glucose-disposal in insulin resistant patients would be reflected in the gene expression profile of skeletal muscle. We performed gene expression profiling on skeletal muscle of insulin resistant and insulin sensitive subjects using microarrays. Microarray analysis of 19,000 genes in skeletal muscle did not display a significant difference between insulin resistant and insulin sensitive muscle. This was confirmed with real-time PCR. Our results suggest that insulin resistance is not reflected by changes in the gene expression profile in skeletal muscle.