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Circulating metabolite levels may reflect the state of the human organism in health and disease, however, the genetic architecture of metabolites is not fully understood. We have performed a whole-genome sequencing association analysis of both common and rare variants in up to 11,840 multi-ethnic participants from five studies with up to 1666 circulating metabolites. We have discovered 1985 novel variant-metabolite associations, and validated 761 locus-metabolite associations reported previously. Seventy-nine novel variant-metabolite associations have been replicated, including three genetic loci located on the X chromosome that have demonstrated its involvement in metabolic regulation. Gene-based analysis have provided further support for seven metabolite-replicated loci pairs and their biologically plausible genes. Among those novel replicated variant-metabolite pairs, follow-up analyses have revealed that 26 metabolites have colocalized with 21 tissues, seven metabolite-disease outcome associations have been putatively causal, and 7 metabolites might be regulated by plasma protein levels. Our results have depicted the genetic contribution to circulating metabolite levels, providing additional insights into understanding human disease.

Gut microbiome studies have documented depletion of butyrate-producing taxa in type 2 diabetes. We analyzed associations between butyrate-producing taxa and detailed measures of insulin homeostasis, whose dysfunction underlies diabetes in 224 non-Hispanic Whites and 129 African Americans, all of whom completed an oral glucose tolerance test. Stool microbiome was assessed by whole-metagenome shotgun sequencing with taxonomic profiling. We examined associations among 36 butyrate-producing taxa (n = 7 genera and 29 species) and insulin sensitivity, insulin secretion, disposition index, insulin clearance, and prevalence of dysglycemia (prediabetes plus diabetes, 46% of cohort), adjusting for age, sex, BMI, and race. The genus Coprococcus was associated with higher insulin sensitivity (β = 0.14; P = 0.002) and disposition index (β = 0.12; P = 0.012) and a lower rate of dysglycemia (odds ratio [OR] 0.91; 95% CI 0.85-0.97; P = 0.0025). In contrast, Flavonifractor was associated with lower insulin sensitivity (β = -0.13; P = 0.004) and disposition index (β = -0.11; P = 0.04) and higher prevalence of dysglycemia (OR 1.22; 95% CI 1.08-1.38; P = 0.0013). Species-level analyses found 10 bacteria associated with beneficial directions of effects and two bacteria with adverse associations on insulin homeostasis and dysglycemia. Although most butyrate producers analyzed appear to be metabolically beneficial, this is not the case for all such bacteria, suggesting that microbiome-directed therapeutic measures to prevent or treat diabetes should be targeted to specific butyrate-producing taxa rather than all butyrate producers.

Short-chain fatty acids (SCFAs) have been extensively studied for potential beneficial roles in glucose homeostasis and risk of diabetes; however, most of this research has focused on butyrate, acetate, and propionate. The effect on metabolism of branched SCFAs (BSCFAs; isobutyrate, isovalerate, and methylbutyrate) is largely unknown. In a cohort of 219 non-Hispanic White participants and 126 African American participants, we examined the association of BSCFA with dysglycemia (prediabetes and diabetes) and oral glucose tolerance test–based measures of glucose and insulin homeostasis, as well as with demographic, anthropometric, lifestyle, and lipid traits, and other SCFAs. We observed a bimodal distribution of BSCFAs, with 25 individuals having high levels (H-BSCFA group) and 320 individuals having lower levels (L-BSCFA group). The prevalence of dysglycemia was lower in the H-BSCFA group compared with the L-BSCFA group (16% vs. 49%; P = 0.0014). This association remained significant after adjustment for age, sex, race, BMI, and levels of other SCFAs. Consistent with the lower rate of dysglycemia, fasting and postprandial glucose levels were lower and the disposition index was higher in the H-BSCFA group. Additional findings in H-BSCFA versus L-BSCFA included lower fasting and postprandial C-peptide levels and lower insulin clearance without differences in insulin levels, insulin sensitivity, insulin secretion, or other variables examined, including diet and physical activity. As one of the first human studies associating higher BSCFA levels with lower odds of dysglycemia and improved glucose homeostasis, this study sets the stage for further investigation of BSCFA as a novel target for prevention or treatment of diabetes. Article Highlights: This study was undertaken to explore the relationship between short-chain fatty acids (SCFAs) and dysglycemia and to explore detailed measures of glucose and insulin homeostasis. We sought to determine whether less frequently studied SCFAs have a role in glucose homeostasis. Individuals with higher levels of branched SCFAs had a lower prevalence of dysglycemia and had improved glucose tolerance. The study findings indicate further investigation is warranted of branched SCFA as novel targets for prevention or treatment of diabetes. [ABSTRACT FROM AUTHOR]

Biologics address a range of unmet clinical needs, but the occurrence of biologics-induced liver injury remains a major challenge. Development of cimaglermin alfa (GGF2) was terminated due to transient elevations in serum aminotransferases and total bilirubin. Tocilizumab has been reported to induce transient aminotransferase elevations, requiring frequent monitoring. To evaluate the clinical risk of biologics-induced liver injury, a novel quantitative systems toxicology modeling platform, BIOLOGXsym™, representing relevant liver biochemistry and the mechanistic effects of biologics on liver pathophysiology, was developed in conjunction with clinically relevant data from a human biomimetic liver microphysiology system. Phenotypic and mechanistic toxicity data and metabolomics analysis from the Liver Acinus Microphysiology System showed that tocilizumab and GGF2 increased high mobility group box 1, indicating hepatic injury and stress. Tocilizumab exposure was associated with increased oxidative stress and extracellular/tissue remodeling, and GGF2 decreased bile acid secretion. BIOLOGXsym simulations, leveraging the in vivo exposure predicted by physiologically-based pharmacokinetic modeling and mechanistic toxicity data from the Liver Acinus Microphysiology System, reproduced the clinically observed liver signals of tocilizumab and GGF2, demonstrating that mechanistic toxicity data from microphysiology systems can be successfully integrated into a quantitative systems toxicology model to identify liabilities of biologics-induced liver injury and provide mechanistic insights into observed liver safety signals. [ABSTRACT FROM AUTHOR]

The renin-angiotensin system (RAS) is a major mediator of renal injury in diabetic nephropathy. Our previous studies demonstrated that 1,25-dihydroxyvitamin [D.sub.3] [1,25[(OH).sub.2][D.sub.3]] plays a renoprotective role by suppressing the RAS, with renin and angiotensinogen (AGT) as the main targets. The mechanism whereby 1,25[(OH).sub.2][D.sub.3] transcriptionally suppresses renin gene expression has been elucidated; however, how vitamin D regulates AGT remains unknown. Exposure of mesangial cells or podocytes to high glucose (HG; 30 mM) markedly stimulated AGT expression. In mesangial cells, the stimulation was inhibited by 1,25[(OH).sub.2][D.sub.3] (20 nM) or NF-[kappa]B inhibitor BAY 11-7082, suggesting the involvement of NF-[kappa]B in HG-induced AGT expression and the interaction between 1,25[(OH).sub.2][D.sub.3] and NF-[kappa]B in the regulation. Plasmid pNF-[kappa]B-Luc luciferase reporter assays showed that 1,25[(OH).sub.2][D.sub.3] blocked HG-induced NF-[kappa]B activity. EMSA and ChIP assays demonstrated increased p65/p50 binding to a NF-[kappa]B binding site at--1734 in the AGT gene promoter upon high glucose stimulation, and the binding was disrupted by 1,25[(OH).sub.2][D.sub.3] treatment. Overexpression of p65/p50 overcame 1,25[(OH).sub.2][D.sub.3] suppression, and mutation of this NF-[kappa]B binding site blunted 1,25[(OH).sub.2][D.sub.3] suppression of the promoter activity. In mice lacking the vitamin D receptor, AGT mRNA expression in the kidney was markedly increased compared with wild-type mice, and AGT induction in diabetic mice was suppressed by treatment with a vitamin D analog. These data indicate that 1,25[(OH).sub.2][D.sub.3] suppresses hyperglycemia-induced AGT expression by blocking NF-[kappa]B-mediated pathway. vitamin D receptor; renin-angiotensin system; diabetic nephropathy

Recent studies have established that vitamin D plays multiple biological roles beyond calcium metabolism; however, whether vitamin D is involved in energy metabolism is unknown. To address this question, we characterized the metabolic phenotypes of vitamin D receptor (VDR)-null mutant mice. Under a normocalcemic condition, VDR-null mice displayed less body fat mass and lower plasma triglyceride and cholesterol levels compared with wild-type (WT) mice; when placed on a high-fat diet, VDR-null mice showed a slower growth rate and accumulated less fat mass globally than WT mice, even though their food intake and intestinal lipid transport capacity were the same as WT mice. Consistent with the lower adipose mass, plasma leptin levels were lower and white adipocytes were histologically smaller in VDR-null mice than WT mice. The rate of fatty acid [beta]-oxidation in the white adipose tissue was higher, and the expression of uncoupling protein (UCP) 1, UCP2 and UCP3 was markedly upregulated in VDR-null mice, suggesting a higher energy expenditure in the mutant mice. Experiments using primary brown fat culture confirmed that 1,25-dihydroxyvitamin [D.sub.3] directly suppressed the expression of the UCPs. Consistently, the energy expenditure, oxygen consumption, and C[O.sub.2] production in VDR-null mice were markedly higher than in WT mice. These data indicate that vitamin D is involved in energy metabolism and adipocyte biology in vivo in part through regulation of [beta]-oxidation and UCP expression. adipocytes; uncoupling proteins; energy metabolism

Emerging evidence supports an inhibitory role for vitamin D in colorectal carcinogenesis; however, the mechanism remains unclear. The adenomatous polyposis coli (APC)/β-catenin pathway plays a critical role in colorectal carcinogenesis. The purpose of our study is to explore the interactions of vitamin D and APC/β-catenin pathways in intestinal tumor development. APCmin/+ mice with genetic inactivation of the vitamin D receptor (VDR) were generated through breeding. Intestinal tumorigenesis was compared between APCmin/+ and APCmin/+VDR-/- mice at different ages. No differences were seen in the number of small intestinal and colonic tumors between APCmin/+ and APCmin/+VDR-/- mice aged 3, 4, 6 and 7 months. The size of the tumors, however, was significantly increased in APCmin/+VDR-/- mice in all age groups. Immunostaining showed significant increases in β-catenin, cyclin D1, phosphorylated Stat-3 and MSH-2 levels and decreases in Stat-1 in APCmin/+VDR-/- tumors compared to APCmin/+ tumors. These observations suggest that VDR signaling inhibits tumor growth rather than tumor initiation in the intestine. Thus, the increased tumor burden in APCmin/+VDR-/- mice is likely due to the loss of the growth-inhibiting effect of VDR. This study provides strong evidence for the in vivo relevance of the interaction demonstrated in vitro between the vitamin D and β-catenin signaling pathways in intestinal tumorigenesis. [ABSTRACT FROM AUTHOR]

As the prevalence of obesity and metabolic syndrome escalates, it has become increasingly important to understand the physiology of adipocytes. 1,25-dihydroxyvitamin D (1,25(OH)[sub 2]D[sub 3]) is a steroid hormone that is traditionally known to be a regulator of mineral homeostasis. 1,25(OH)[sub 2]D[sub 3] exerts its effects through the vitamin D receptor (VDR), a nuclear receptor that regulates gene transcription. Our recent studies suggest that VDR plays an integral role in adipocyte differentiation. VDR expression is tightly regulated during adipocyte differentiation and treatment of preadipocytic cells with 1,25(OH)[sub 2]D[sub 3] suppresses differentiation. However, the in vivo role of VDR in lipid metabolism has not been explored. We, therefore, evaluated the body fat composition and lipid parameters of VDR(-/-) mice. On a regular diet, VDR (-/-) mice weighed less and contained less body fat than wild type (WT) mice. On the high calcium-high lactose diet that rescues the hypocalcemia, the difference between VDR(-/-) and WT mice appeared sex dependent. In males, VDR(-/-) mice weighed less and had a lower body fat than WT mice; however no significant difference was observed in females. On a high fat diet, VDR (-/-) mice gained weight at a much lower rate than WT mice regardless of the sex. The gene profile of adipocytes from VDR(-/-) mice was also found to be altered. Levels of leptin and adiponectin were reduced in VDR(-/-) mice compared to WT mice, whereas resistin levels were increased in VDR(-/-) mice. Plasma cholesterol and triglyceride levels were significantly lower in VDR (-/-) mice than WT mice. These observations indicate that VDR plays a role in the regulation of adipose mass and lipid metabolism in vivo, which is independent of its role in calcium metabolism. [ABSTRACT FROM AUTHOR]

We have shown that 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) down-regulates renin expression. To explore the molecular mechanism, we analyzed the mouse Ren-1c gene promoter by luciferase reporter assays. Deletion analysis revealed two DNA fragments from -2725 to -2647 (distal fragment) and from -117 to +6 (proximal fragment) that are sufficient to mediate the repression. Mutation of the cAMP response element (CRE) in the distal fragment blunted forskolin stimulation as well as 1,25(OH)2D3 inhibition of the transcriptional activity, suggesting the involvement of CRE in 1,25(OH)2D3-induced suppression. EMSA revealed that 1,25(OH)2D3 markedly inhibited nuclear protein binding to the CRE in the promoter. ChIP and GST pull-down assays demonstrated that liganded VDR blocked the binding of CREB to the CRE by directly interacting with CREB with the ligand-binding domain, and the VDR-mediated repression can be rescued by CREB, CBP, or p300 overexpression. These data indicate that 1,25(OH)2D3 suppresses renin gene expression at least in part by blocking the formation of CRE-CREB-CBP complex. [ABSTRACT FROM AUTHOR]

Circulating metabolite levels partly reflect the state of human health and diseases, and can be impacted by genetic determinants. Hundreds of loci associated with circulating metabolites have been identified; however, most findings focus on predominantly European ancestry or single study analyses. Leveraging the rich metabolomics resources generated by the NHLBI Trans-Omics for Precision Medicine (TOPMed) Program, we harmonized and accessibly cataloged 1,729 circulating metabolites among 25,058 ancestrally-diverse samples. We provided recommendations for outlier and imputation handling to process metabolite data, as well as a general analytical framework. We further performed a pooled analysis following our practical recommendations and discovered 1,778 independent loci associated with 667 metabolites. Among 108 novel locus - metabolite pairs, we detected not only novel loci within previously implicated metabolite associated genes, but also novel genes (such as GAB3 and VSIG4 located in the X chromosome) that have putative roles in metabolic regulation. In the sex-stratified analysis, we revealed 85 independent locus-metabolite pairs with evidence of sexual dimorphism, including well-known metabolic genes such as FADS2 , D2HGDH , SUGP1 , UTG2B17 , strongly supporting the importance of exploring sex difference in the human metabolome. Taken together, our study depicted the genetic contribution to circulating metabolite levels, providing additional insight into the understanding of human health.