The orphan G-protein-coupled receptor GPR109B is the result of a recent gene duplication of the nicotinic acid and ketone body receptor GPR109A being found in humans but not in rodents. Like GPR109A, GPR109B is predominantly expressed in adipocytes and is supposed to mediate antilipolytic effects. Here we show that GPR109B serves as a receptor for the -oxidation intermediate 3-OH-octanoic acid, which has antilipolytic activity on human but not on murine adipocytes. GPR109B is coupled to G i -type G-proteins and is activated by 2-and 3-OH-octanoic acid with EC 50 values of about 4 and 8 M, respectively. Interestingly, 3-OH-octanoic acid plasma concentrations reach micromolar concentrations under conditions of increased -oxidation rates, like in diabetic ketoacidosis or under a ketogenic diet. These data suggest that the ligand receptor pair 3-OH-octanoic acid/GPR109B mediates in humans a negative feedback regulation of adipocyte lipolysis to counteract prolipolytic influences under conditions of physiological or pathological increases in -oxidation rates.
Postnatal liver development is characterized by hepatocyte growth, proliferation, and functional maturation. Notably, canonical Wnt signaling in hepatocytes has been identified as an important regulator of final adult liver size and metabolic liver zonation. The cellular origin of Wnt ligands responsible for homeostatic liver/body weight ratio (LW/BW) remained unclear, which was also attributable to a lack of suitable endothelial Cre driver mice. To comprehensively analyze the effects of hepatic angiocrine Wnt signaling on liver development and metabolic functions, we used endothelial subtype‐specific Stab2‐Cre driver mice to delete Wls from hepatic endothelial cells (HECs). The resultant Stab2‐Cretg/wt;Wlsfl/fl (Wls‐HECKO) mice were viable, but showed a significantly reduced LW/BW. Specifically, ablation of angiocrine Wnt signaling impaired metabolic zonation in the liver, as shown by loss of pericentral, β‐catenin‐dependent target genes such as glutamine synthase (Glul), RhBg, Axin2, and cytochrome P450 2E1, as well as by extended expression of periportal genes such as arginase 1. Furthermore, endothelial subtype‐specific expression of a c‐terminally YFP‐tagged Wls fusion protein in Wls‐HECKO mice (Stab2‐Cretg/wt;Wlsfl/fl;Rosa26:Wls‐YFPfl/wt [Wls‐rescue]) restored metabolic liver zonation. Interestingly, lipid metabolism was altered in Wls‐HECKO mice exhibiting significantly reduced plasma cholesterol levels, while maintaining normal plasma triglyceride and blood glucose concentrations. On the contrary, zonal expression of Endomucin, LYVE1, and other markers of HEC heterogeneity were not altered in Wls‐HECKO livers. Conclusion: Angiocrine Wnt signaling controls liver growth as well as development of metabolic liver zonation in mice, whereas intrahepatic HEC zonation is not affected. (Hepatology 2017).
Levels of (D)-2-hydroxyglutarate [D2HG, (R)-2-hydroxyglutarate] are increased in some metabolic diseases and in neoplasms with mutations in the isocitrate dehydrogenase 1 (IDH1) and isocitrate dehydrogenase 2 (IDH2) genes. Determination of D2HG is of relevance to diagnosis and monitoring of disease. Standard detection methods of D2HG levels are liquid-chromatography-mass spectrometry or gas-chromatography-mass spectrometry. Here we present a rapid, inexpensive and sensitive enzymatic assay for the detection of D2HG levels. The assay is based on the conversion of D2HG to α-ketoglutarate (αKG) in the presence of the enzyme (D)-2-hydroxyglutarate dehydrogenase (HGDH) and nicotinamide adenine dinucleotide (NAD(+)). Determination of D2HG concentration is based on the detection of stoichiometrically generated NADH. The quantification limit of the enzymatic assay for D2HG in tumor tissue is 0.44 μM and in serum 2.77 μM. These limits enable detection of basal D2HG levels in human tumor tissues and serum without IDH mutations. Levels of D2HG in frozen and paraffin-embedded tumor tissues containing IDH mutations or in serum from acute myeloid leukemia patients with IDH mutations are significantly higher and can be easily identified with this assay. In conclusion, the assay presented is useful for differentiating basal from elevated D2HG levels in tumor tissue, serum, urine, cultured cells and culture supernatants.
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