In vivo glycerol metabolism in patients with glycerol kinase deficiency.

Glycerol kinase deficiency (GKD) is an X-linked recessive disorder due to () gene mutations resulting in hyperglycerolermia, hyperglyceroluria, and "pseudohypertriglyceridemia." In vivo glycerol metabolism has not been assessed in GKD. A 62-year-old man with suspected GKD and his extended family underwent whole exome sequencing and fasting blood work with two modes of lipid measurements: (1) standard lipase-based methodology and (2) nuclear magnetic resonance (NMR).

Regulation of hypothalamic reactive oxygen species and feeding behavior by phosphorylation of the beta 2 thyroid hormone receptor isoform.

Unlike other thyroid hormone receptors (THRs), the beta 2 isoform (THRB2) has a restricted expression pattern and is uniquely and abundantly phosphorylated at a conserved serine residue S101 (S102 in humans). Using tagged and or phosphorylation-defective (S101A) THRB2 mutant mice, we show that THRB2 is present in a large subset of POMC neurons and mitigates ROS accumulation during ROS-triggering events, such as fasting/refeeding or high fat diet (HFD). Excessive ROS accumulation in mutant POMC neurons was accompanied by a skewed production of orexigenic/anorexigenic hormones, resulting in elevated food intake.

Leukemia inhibitory factor suppresses hepatic de novo lipogenesis and induces cachexia in mice.

Cancer cachexia is a systemic metabolic syndrome characterized by involuntary weight loss, and muscle and adipose tissue wasting. Mechanisms underlying cachexia remain poorly understood. Leukemia inhibitory factor (LIF), a multi-functional cytokine, has been suggested as a cachexia-inducing factor.

Protocol for measuring mitochondrial size in mouse and human liver tissues.

Mitochondrial dynamic process is important for cell viability, metabolic activity, and mitochondria health. Here, we present a protocol for measuring mitochondrial size through immunofluorescence staining, confocal imaging, and analysis in ImageJ. We describe the steps for tissue processing, antigen retrieval, mitochondrial staining using an integrating immunofluorescence assay, and computerized image analysis to measure each mitochondrial size in mouse and human liver tissues.

Blocking AMPKαS496 phosphorylation improves mitochondrial dynamics and hyperglycemia in aging and obesity.

Impaired mitochondrial dynamics causes aging-related or metabolic diseases. Yet, the molecular mechanism responsible for the impairment of mitochondrial dynamics is still not well understood. Here, we report that elevated blood insulin and/or glucagon levels downregulate mitochondrial fission through directly phosphorylating AMPKα at S496 by AKT or PKA, resulting in the impairment of AMPK-MFF-DRP1 signaling and mitochondrial dynamics and activity.

Gluconeogenesis Flux in Metabolic Disease.

Gluconeogenesis is a critical biosynthetic process that helps maintain whole-body glucose homeostasis and becomes altered in certain medical diseases. We review gluconeogenic flux in various medical diseases, including common metabolic disorders, hormonal imbalances, specific inborn genetic errors, and cancer. We discuss how the altered gluconeogenic activity contributes to disease pathogenesis using data from experiments using isotopic tracer and spectroscopy methodologies.

Glucagon changes substrate preference in gluconeogenesis.

Fasting hyperglycemia in diabetes mellitus is caused by unregulated glucagon secretion that activates gluconeogenesis (GNG) and increases the use of pyruvate, lactate, amino acids, and glycerol. Studies of GNG in hepatocytes, however, tend to test a limited number of substrates at nonphysiologic concentrations. Therefore, we treated cultured primary hepatocytes with three identical substrate mixtures of pyruvate/lactate, glutamine, and glycerol at serum fasting concentrations, where a different U-C- or 2-C-labeled substrate was substituted in each mix.

Differential Metabolism of Glycerol Based on Oral versus Intravenous Administration in Humans.

Glycerol can be metabolized to glucose via gluconeogenesis or lactate via glycolysis. It is unknown if glycerol is metabolized similarly in the portal and systemic circulations in humans. Eight metabolically healthy overnight-fasted individuals received equimolar amounts of C-glycerol orally and intravenously on two separate occasions with serial blood draws over four hours.

Activation of the canonical ER stress IRE1-XBP1 pathway by insulin regulates glucose and lipid metabolism.

Knockout of the transcription factor X-box binding protein (XBP1) is known to decrease liver glucose production and lipogenesis. However, whether insulin can regulate gluconeogenesis and lipogenesis through XBP1 and how insulin activates the inositol-requiring enzyme-XBP1 ER stress pathway remains unexplored. Here, we report that in the fed state, insulin-activated kinase AKT directly phosphorylates inositol-requiring enzyme 1 at S724, which in turn mediates the splicing of XBP1u mRNA, thus favoring the generation of the spliced form, XBP1s, in the liver of mice.

Glycerol's contribution to lactate production outside of a glucose intermediate in fasting humans.

Background: Glycerol is a well-recognized substrate for new glucose production via gluconeogenesis in the liver. However, its carbon contribution to the glycolytic intermediate lactate is not known in humans.

Methods: Here we infused stable isotope tracers C-glycerol and 6,6-D-glucose into six metabolically healthy individuals after an overnight fast to study glycerol metabolism and measure glucose rate of appearance.

Optimized 3D Culture of Hepatic Cells for Liver Organoid Metabolic Assays.

The liver is among the principal organs for glucose homeostasis and metabolism. Studies of liver metabolism are limited by the inability to expand primary hepatocytes in vitro while maintaining their metabolic functions. Human hepatic three-dimensional (3D) organoids have been established using defined factors, yet hepatic organoids from adult donors showed impaired expansion.

Thyroid hormone receptor phosphorylation regulates acute fasting-induced suppression of the hypothalamic-pituitary-thyroid axis.

Fasting induces profound changes in the hypothalamic-pituitary-thyroid (HPT) axis. After binding thyroid hormone (TH), the TH receptor beta 2 isoform (THRB2) represses and subunit genes and is the principle negative regulator of the HPT axis. Using mass spectrometry, we identified a major phosphorylation site in the AF-1 domain of THRB2 (serine 101, S101), which is conserved among many members of the nuclear hormone receptor superfamily.

Metabolic Flux Analysis-Linking Isotope Labeling and Metabolic Fluxes.

Metabolic flux analysis (MFA) is an increasingly important tool to study metabolism quantitatively. Unlike the concentrations of metabolites, the fluxes, which are the rates at which intracellular metabolites interconvert, are not directly measurable. MFA uses stable isotope labeled tracers to reveal information related to the fluxes.

Utilizing tandem mass spectrometry for metabolic flux analysis.

Metabolic flux analysis (MFA) aims at revealing the metabolic reaction rates in a complex biochemical network. To do so, MFA uses the input of stable isotope labeling patterns of the intracellular metabolites. Elementary metabolic unit (EMU) is the computational framework to simulate the metabolite labeling patterns in a network, which was originally designed for simulating mass isotopomer distributions (MIDs) at the MS1 level.

Tracking the carbons supplying gluconeogenesis.

As the burden of type 2 diabetes mellitus (T2DM) grows in the 21st century, the need to understand glucose metabolism heightens. Increased gluconeogenesis is a major contributor to the hyperglycemia seen in T2DM. Isotope tracer experiments in humans and animals over several decades have offered insights into gluconeogenesis under euglycemic and diabetic conditions.

Dysregulated expression of monoacylglycerol lipase is a marker for anti-diabetic drug metformin-targeted therapy to correct impaired neurogenesis and spatial memory in Alzheimer's disease.

Monoacylglycerol lipase (Mgll), a hydrolase that breaks down the endocannabinoid 2-arachidonoyl glycerol (2-AG) to produce arachidonic acid (ARA), is a potential target for neurodegenerative diseases, such as Alzheimer's disease (AD). Increasing evidence shows that impairment of adult neurogenesis by perturbed lipid metabolism predisposes patients to AD. However, it remains unknown what causes aberrant expression of Mgll in AD and how Mgll-regulated lipid metabolism impacts adult neurogenesis, thus predisposing to AD during aging.

In-Source CID Ramping and Covariant Ion Analysis of Hydrophilic Interaction Chromatography Metabolomics.

A large proportion of the complexity and redundancy of LC-MS metabolomics data comes from adduct formation. To reduce such redundancy, many tools have been developed to recognize and annotate adduct ions. These tools rely on predefined adduct lists that are generated empirically from reversed-phase LC-MS studies.

A Direct Comparison of Thyroid Hormone Receptor Protein Levels in Mice Provides Unexpected Insights into Thyroid Hormone Action.

Thyroid hormone (TH) action is mediated by three major thyroid hormone receptor (THR) isoforms α1, β1, and β2 (THRA1, THRB1, and THRB2). These THRs and a fourth major but non-TH binding isoform, THRA2, are encoded by two genes and . Reliable antibodies against all THR isoforms are not available, and THR isoform protein levels in mammalian tissues are often inferred from messenger RNA (mRNA) levels.

G protein-coupled kisspeptin receptor induces metabolic reprograming and tumorigenesis in estrogen receptor-negative breast cancer.

Triple-negative breast cancer (TNBC) is a highly metastatic and deadly disease. TNBC tumors lack estrogen receptor (ERα), progesterone receptor (PR), and HER2 (ErbB2) and exhibit increased glutamine metabolism, a requirement for tumor growth. The G protein-coupled kisspeptin receptor (KISS1R) is highly expressed in patient TNBC tumors and promotes malignant transformation of breast epithelial cells.

KISS1/KISS1R and Breast Cancer: Metastasis Promoter.

Kisspeptins (KPs), peptide products of the kisspeptin-1 () gene, are the endogenous ligands for the KISS1 receptor, KISS1R, which is a G protein-coupled receptor. In many human tumors, functions as a metastasis-suppressor gene and KISS1/KISS1R signaling has antimetastatic and tumor-suppressor roles. On the contrary, emerging evidence indicates that the KP/KISS1R pathway plays detrimental roles in triple negative breast cancer (TNBC), the most difficult type of breast cancer to treat.

Glycerol not lactate is the major net carbon source for gluconeogenesis in mice during both short and prolonged fasting.

Objective: Fasting results in major metabolic changes including a switch from glycogenolysis to gluconeogenesis to maintain glucose homeostasis. However, the relationship between the length of fasting and the relative contribution of gluconeogenic substrates remains unclear. We investigated the relative contribution of glycogen, lactate, and glycerol in glucose production of male C57BL/6 J-albino mice after 6, 12, and 18 h of fasting.