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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. Recently, the EMU framework is expanded to simulate tandem mass spectrometry data. Tandem mass spectrometry has emerged as a new experimental approach to provide information on the positional isotope labeling of metabolites and therefore greatly improves the precision of MFA. In this review, we will discuss the new EMU framework that can accommodate the tandem mass isotopomer distributions (TMIDs) data. We will also analyze the improvement on the MFA precision by using TMID. Our analysis shows that combining the MIDs of the parent and daughter ions and the TMID for the MFA is more powerful than using TMID alone.

Nonalcoholic fatty liver disease (NAFLD), the most common liver disease, has become a silent worldwide pandemic. The incidence of NAFLD correlates with the rise in obesity, type 2 diabetes, and metabolic syndrome. A hallmark featureof NAFLD is excessive hepatic fat accumulation or steatosis, due to dysregulated hepatic fat metabolism, which can progress to nonalcoholic steatohepatitis (NASH), fibrosis, and cirrhosis. Currently, there are no approved pharmacotherapies to treat this disease. Here, we have found that activation of the kisspeptin 1 receptor (KISS1R) signaling pathway has therapeutic effects in NAFLD. Using high-fat diet-fed mice, we demonstrated that a deletion of hepatic Kiss1r exacerbated hepatic steatosis. In contrast, enhanced stimulation of KISS1R protected against steatosis in wild-type C57BL/6J mice and decreased fibrosis using a diet-induced mouse model of NASH. Mechanistically, we found that hepatic KISS1R signaling activates the master energy regulator, AMPK, to thereby decrease lipogenesis and progression to NASH. In patients with NAFLD and in high-fat diet-fed mice, hepatic KISS1/KISS1R expression and plasma kisspeptin levels were elevated, suggesting a compensatory mechanism to reduce triglyceride synthesis. These findings establish KISS1R as a therapeutic target to treat NASH.

Despite significant scientific and medical discoveries, the genetics of novel infectious diseases like COVID-19 remains far from understanding. SARS-CoV-2 is a single-stranded RNA respiratory virus that causes COVID-19 by binding to the ACE2 receptor in the lung and other organs. Understanding its clinical presentation and metabolomic and genetic profile will lead to the discovery of diagnostic, prognostic and predictive biomarkers, which may lead to more effective medical therapy. It is important to investigate correlations and overlap between reported diagnoses of a patient with COVID-19 in clinical data with identified germline and somatic mutations, and highly expressed genes from genomics data analysis. Timely model clinical, genomics and metabolomics data to find statistical patterns across millions of features to identify underlying biological pathways, modifiable risk factors and actionable information that supports early detection and prevention of COVID-19, and development of new therapies for better patient care. Next, ensuring security reconcile noise, need to build and train machine learning prognostic models to find actionable information that supports early detection and prevention of COVID-19. Based on the myriad data, applying appropriate machine learning algorithms to stratify patients, understand scenarios, optimise decision-making, identify high-risk rare variants (including ACE2, TMPRSS2) and making medically relevant predictions. Innovative and intelligent solutions are required to improve the traditional symptom-driven practice, and allow earlier interventions using predictive diagnostics and tailor better personalised treatments, when confronted with the challenges of pandemic situations.

Background: 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 Thra and Thrb. 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. Methods: We generated knock-in mouse models expressing endogenously and identically 2X hemagglutenin epitope (HA)-tagged THRs (THRA1/2, THRB1, and THRB2), which could then be detected by commercially available anti-HA antibodies. Using nuclear enrichment, immunoprecipitation, and Western blotting, we determined relative THR protein expression in 16 mouse organs. Results: In all peripheral organs tested except the liver, the predominant THR isoform was THRA1. Surprisingly, in metabolically active organs such as fat and muscle, THRB1 protein levels were up to 10 times lower than that of THRA1, while their mRNA levels appeared similar. In contrast to peripheral organs, the central nervous system (CNS) had a unique pattern with relatively low levels of both THRB1 and THRA1, and high levels of THRA2 expression. As expected, THRB2 was highly expressed in the pituitary, but a previously unknown sex-specific difference in THRB2 expression was found (female mice having higher pituitary expression than male mice). Higher THRB2 expression appears to make the central axis more sensitive to TH as both serum thyrotropin and Tshb mRNA levels were lower in female mice. Conclusions: Direct comparison of THR protein abundance in different organs using endogenously tagged HA-THR mouse lines shows that expression of THR isoforms is regulated at transcriptional and posttranscriptional levels, and in organ-specific manner. The prevalence of THRA1 and low abundance of THRB1 in majority of peripheral tissues suggest that peripheral actions of these isoforms should be revisited. A unique pattern of high THRA2 in CNS warrants further exploration of this non-TH binding isoform in brain development. Finally, THRB2, in addition to cell-specific control, is also regulated in a sex-specific manner, which may change the hypothalamus–pituitary–thyroid axis set point and perhaps metabolism in males and females.

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 adduc...

Rationale: 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. Here, we identify Mgll as an aging-induced factor that impairs adult neurogenesis and spatial memory in AD, and show that metformin, an FDA-approved anti-diabetic drug, can reduce the expression of Mgll to reverse impaired adult neurogenesis, prevent spatial memory decline and reduce β-amyloid accumulation. Methods: Mgll expression was assessed in both human AD patient post-mortem hippocampal tissues and 3xTg-AD mouse model. In addition, we used both the 3xTg-AD animal model and the CbpS436A genetic knock-in mouse model to identify that elevated Mgll expression is caused by the attenuation of the aPKC-CBP pathway, involving atypical protein kinase C (aPKC)-stimulated Ser436 phosphorylation of histone acetyltransferase CBP through biochemical methods. Furthermore, we performed in vivo adult neurogenesis assay with BrdU/EdU labelling and Morris water maze task in both animal models following pharmacological treatments to show the key role of Mgll in metformin-corrected neurogenesis and spatial memory deficits of AD through reactivating the aPKC-CBP pathway. Finally, we performed in vitro adult neurosphere assays using both animal models to study the role of the aPKC-CBP mediated Mgll repression in determining adult neural stem/progenitor cell (NPC) fate. Results: Here, we demonstrate that aging-dependent induction of Mgll is observed in the 3xTg-AD model and human AD patient post-mortem hippocampal tissues. Importantly, we discover that elevated Mgll expression is caused by the attenuation of the aPKC-CBP pathway. The accumulation of Mgll in the 3xTg-AD mice reduces the genesis of newborn neurons and perturbs spatial memory. However, we find that metformin-stimulated aPKC-CBP pathway decreases Mgll expression to recover these deficits in 3xTg-AD. In addition, we reveal that elevated Mgll levels in cultured adult NPCs from both 3xTg-AD and CbpS436A animal models are responsible for their NPC neuronal differentiation deficits. Conclusion: Our findings set the stage for development of a clinical protocol where Mgll would serve as a biomarker in early stages of AD to identify potential metformin-responsive AD patients to restore their neurogenesis and spatial memory.

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. Methods We used non-perturbative infusions of 13C3 lactate, 13C3 glycerol, and 13C6 glucose combined with liquid chromatography mass spectrometry and metabolic flux analysis to study the contribution of substrates in gluconeogenesis (GNG). Results During infusion studies, both lactate and glycerol significantly label about 60% and 30–50% glucose carbon, respectively, but glucose labels much more lactate (∼90%) than glycerol carbon (∼10%). Our analyses indicate that lactate, but not glycerol is largely recycled during all fasting periods such that lactate is the largest direct contributor to GNG via the Cori cycle but a minor source of new glucose carbon (overall contribution). In contrast, glycerol is not only a significant direct contributor to GNG but also the largest overall contributor to GNG regardless of fasting length. Prolonged fasting decreases both the whole body turnover rate of glucose and lactate but increases that of glycerol, indicating that the usage of glycerol in GNG become more significant with longer fasting. Conclusion Collectively, these findings suggest that glycerol is the dominant overall contributor of net glucose carbon in GNG during both short and prolonged fasting., Highlights • Prolonged fasting significantly decreases the turnover rate of glucose and lactate but increases the glycerol turnover rate in mice. • In both short and prolonged fasting, lactate is the largest direct contributor to gluconeogenesis but a minor source of new carbon entry. • Glycerol is the second largest direct contributor to gluconeogenesis and the dominant overall carbon contributor during both short and prolonged fasting.

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.Here we infused stable isotope tracersGlycerol and glucose rates of appearance were 2.21 ± 1.42 μmol/kg/min and 7.81 ± 1.15 μmol/kg/min, respectively. Under steady-state conditions, theGiven that lactate itself is a carbon source for gluconeogenesis and tricycarboxylic cycle intermediates, glycerol's ability to donate carbons to lactate may make it quantitatively more important to intermediary metabolism than currently appreciated.

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 non-physiologic 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

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. Subsequently, XBP1s stimulate the expression of lipogenic genes and upregulates liver lipogenesis as previously reported. Intriguingly, we find that fasting leads to an increase in XBP1u along with a drastic decrease in XBP1s in the liver of mice, and XBP1u, not XBP1s, significantly increases PKA-stimulated CRE reporter activity in cultured hepatocytes. Furthermore, we demonstrate that overexpression of XBP1u significantly increases cAMP-stimulated expression of rate-limiting gluconeogenic genes, G6pc and Pck1, and glucose production in primary hepatocytes. Reexpression of XBP1u in the liver of mice with XBP1 depletion significantly increases fasting blood glucose levels and gluconeogenic gene expression. These data support an important role of XBP1u in upregulating gluconeogenesis in the fasted state. Taken together, we reveal that insulin signaling via AKT controls the expression of XBP1 isoforms and that XBP1u and XBP1s function in different nutritional states to regulate liver gluconeogenesis and lipogenesis, respectively.

Fasting induces profound changes in the hypothalamic–pituitary–thyroid (HPT) axis. After binding thyroid hormone (TH), the TH receptor beta 2 isoform (THRB2) represses Trh and Tsh 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. More than 50% of THRB2 is phosphorylated at S101 in cultured thyrotrophs (TαT1.1) and in the mouse pituitary. All other THR isoforms lack this site and exhibit limited overall levels of phosphorylation. To determine the importance of THRB2 S101 phosphorylation, we used the TαT1.1 cell line and S101A mutant knock-in mice (Thrb2(S101A)). We found that TH promoted S101 THRB2 phosphorylation and was essential for repression of the axis at physiologic TH concentrations. In mice, THRB2 phosphorylation was also increased by fasting and mimicked Trh and Tshb repression by TH. In vitro studies demonstrated that a master metabolic sensor, AMP-activated kinase (AMPK) induced phosphorylation at the same site and caused Tshb repression independent of TH. Furthermore, we identified cyclin-dependent kinase 2 (CDK2) as a direct kinase phosphorylating THRB2 S101 and propose that AMPK or TH increase S101 phosphorylation through the activity of CDK2. This study provides a physiologically relevant function for THR phosphorylation, which permits nutritional deprivation and TH to use a common mechanism for acute suppression of the HPT axis.

Gluconeogenesis (GNG) is de novo production of glucose from endogenous carbon sources. Although it is a commonly studied pathway, particularly in disease, there is a lack of consensus about substrate preference. Moreover, primary hepatocytes are the current gold standard for in vitro liver studies, but no direct comparison of substrate preference at physiological fasting concentrations has been performed. We show that mouse primary hepatocytes prefer glycerol to pyruvate/lactate in glucose production assays and 13C isotope tracing studies at the high concentrations commonly used in the literature, as well as at more relevant fasting, physiological concentrations. In addition, when glycerol, pyruvate/lactate, and glutamine are all present, glycerol is responsible for over 75% of all glucose carbons labeled. We also found that glycerol can induce a rate-limiting enzyme of GNG, glucose-6-phosphatase. Lastly, we suggest that glycerol is a better substrate than pyruvate to test in vivo production of glucose in fasting mice. In conclusion, glycerol is the major carbon source for GNG in vitro and in vivo and should be compared with other substrates when studying GNG in the context of metabolic disease states.

Summary: Impaired mitochondrial respiratory activity contributes to the development of insulin resistance in type 2 diabetes. Metformin, a first-line antidiabetic drug, functions mainly by improving patients’ hyperglycemia and insulin resistance. However, its mechanism of action is still not well understood. We show here that pharmacological metformin concentration increases mitochondrial respiration, membrane potential, and ATP levels in hepatocytes and a clinically relevant metformin dose increases liver mitochondrial density and complex 1 activity along with improved hyperglycemia in high-fat- diet (HFD)-fed mice. Metformin, functioning through 5′ AMP-activated protein kinase (AMPK), promotes mitochondrial fission to improve mitochondrial respiration and restore the mitochondrial life cycle. Furthermore, HFD-fed-mice with liver-specific knockout of AMPKα1/2 subunits exhibit higher blood glucose levels when treated with metformin. Our results demonstrate that activation of AMPK by metformin improves mitochondrial respiration and hyperglycemia in obesity. We also found that supra-pharmacological metformin concentrations reduce adenine nucleotides, resulting in the halt of mitochondrial respiration. These findings suggest a mechanism for metformin’s anti-tumor effects. : The mechanism of metformin action still remains controversial, in particular on mitochondrial activity and the involvement of AMPK. Wang et al. show that pharmacological metformin concentration or dose improves mitochondrial respiration by increasing mitochondrial fission through AMPK-Mff signaling; in contrast, supra-pharmacological metformin concentrations reduce mitochondrial respiration through decreasing adenine nucleotide levels. Keywords: metformin, diabetes, insulin resistance, mitochondrial respiration/fission, membrane potential, adenine nucleotides, AMPK, Drp1

Glucose and glycerol are important circulating metabolites. Due to poor ionization and/or ion suppression, the liquid chromatography-mass spectrometry (LC-MS) detection of glucose and glycerol presents challenges. Here, we propose an efficient LC-MS method of quantitative glucose and glycerol detection via enzymatic derivatization to glucose-6-phosphate and sn-glycerol-3-phosphate, respectively. This derivatization protocol can be used to measure the concentrations of glucose production in a plethora of sample types for metabolic analysis and is compatible with the general metabolomics workflow. This novel approach allows us to quantitatively study glucose and glycerol metabolism using stable isotope tracers in vivo.

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. The conceptual idea of MFA is that in tracer experiments the isotope labeling patterns of intracellular metabolites are determined by the fluxes, therefore by measuring the labeling patterns we can infer the fluxes in the network. In this review, we will discuss the basic concept of MFA using a simplified upper glycolysis network as an example. We will show how the fluxes are reflected in the isotope labeling patterns. The central idea we wish to deliver is that under metabolic and isotopic steady-state the labeling pattern of a metabolite is the flux-weighted average of the substrates’ labeling patterns. As a result, MFA can tell the relative contributions of converging metabolic pathways only when these pathways make substrates in different labeling patterns for the shared product. This is the fundamental principle guiding the design of isotope labeling experiment for MFA including tracer selection. In addition, we will also discuss the basic biochemical assumptions of MFA, and we will show the flux-solving procedure and result evaluation. Finally, we will highlight the link between isotopically stationary and nonstationary flux analysis.

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. This review focuses on the current understanding of carbon flux in gluconeogenesis, including substrate contribution of various gluconeogenic precursors to glucose production. Alterations of gluconeogenic metabolites and fluxes in T2DM are discussed. We also highlight ongoing knowledge gaps in the literature that require further investigation. A comprehensive analysis of gluconeogenesis may enable a better understanding of T2DM pathophysiology and identification of novel targets for treating hyperglycemia.

Kisspeptin is a hypothalamic protein critical for neuroendocrine control of pubertal development and fertility and is modulated by nutritional signals. Kisspeptin has been localized to specific neurons located in the arcuate and anteroventral periventricular (AVPV) nuclei of the hypothalamus and is secreted to control GnRH mediated pubertal maturation and reproduction. Kisspeptin has also been localized to peripheral tissues including the liver, fat, gonads, intestine and placenta, although its role in these tissues is unclear. The objective of current study is to define the role of hepatic kisspeptin as a metabolic sensor. A floxed Kiss1 mouse has been developed, and ablation of liver-specific Kiss1 was achieved in two to three month old Kiss1f/f male and female mice given a single tail vein injection of thyroid hormone-binding globulin (TBG) promoter-driven Cre recombinase adeno-associated virus (AAV-CRE). A control group of Kiss1f/f male and female mice received an injection of AAV-GFP, expressing green fluorescent protein. Two weeks after injection, a glucose tolerance test (GTT) was performed followed by an insulin tolerance test. To determine whether changes had occurred in the reproductive axis, estrous cyclicity was assessed by daily vaginal smears and estrous cycle phases determined by vaginal cytology. Mice were euthanized four weeks post-injection and tissues were collected for RNA extraction and gene expression analysis via qRT-PCR. As expected, qRT-PCR data showed absence of Kiss1 expression in the liver of AAV-CRE mice compared to AAV-GFP mice with no changes in kisspeptin gene expression were noted in the ovary, testes, spleen, pancreas, arcuate or AVPV. Estrous cyclicity was also not affected by viral ablation of hepatic Kiss1. Elevated fasting glucose and glucose intolerance in the GTT were found in AAV-CRE compared to AAV-GFP females (P < 0.05). No differences in AAV-CRE and AAV-GFP male mice were found, indicating the importance of Kiss1 in glucose homeostasis in females. The insulin tolerance test was not statistically different between groups or treatments. Further research is required to elucidate the mechanism by which hepatic kisspeptin alters glucose metabolism in mice in a sexually-dimorphic fashion.

Background: In patients with system based diseases, multiple pathologies coexisting in the same organ can go undiagnosed. We present a case of a patient with long standing hypertension recently diagnosed with Primary hyperaldosteronism in the setting of known Pheochromocytoma. Case: A 43 year old male with a history of Von Hippel Lindau (VHL), Pheochromocytoma status post right subtotal adrenalectomy 18 years ago, renal cell carcinoma, prediabetes, bilateral renal cysts, and hypertension presented to the clinic for evaluation of suspected hyperaldosteronism. The patient had a history of medication refractory hypertension associated with hypokalemia requiring potassium supplementation. The patient’s VHL disease involved the pancreas, adrenal, brain and spine which required surgery to remove tumors in these areas. His Pheochromocytoma resolved status post subtotal adrenalectomy. Recent imaging and laboratory work up showed a stable left adrenal nodule (unchanged over the years), unremarkable right adrenal status post subtotal adrenalectomy with normal serum and urine metanephrines. In September 2019 he was seen by his Cardiologist for persistent Atrial Fibrillation and subsequent workup for suspected hyperaldosteronism showed a morning plasma aldosterone of 44.4ng/dL (normal range 4–21 ng/dL), plasma renin activity of

Individuals with demyelinating diseases often experience difficulties during social interactions that are not well studied in preclinical models. Here, we describe a novel juvenile focal corpus callosum demyelination murine model exhibiting a social interaction deficit. Using this preclinical murine demyelination model, we discover that application of metformin, an FDA-approved drug, in this model promotes oligodendrocyte regeneration and remyelination and improves the social interaction. This beneficial effect of metformin acts through stimulating Ser436 phosphorylation in CBP, a histone acetyltransferase. In addition, we found that metformin acts through two distinct molecular pathways to enhance oligodendrocyte precursor (OPC) proliferation and differentiation, respectively. Metformin enhances OPC proliferation through early-stage autophagy inhibition, while metformin promotes OPC differentiation into mature oligodendrocytes through activating CBP Ser436 phosphorylation. In summary, we identify that metformin is a promising remyelinating agent to improve juvenile demyelination-associated social interaction deficits by promoting oligodendrocyte regeneration and remyelination.

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 pre-defined adduct lists which are learned empirically from reverse phase LC-MS studies. Meanwhile, hydrophilic interaction chromatography (HILIC) is gaining popularity in metabolomics studies due to better performance on polar compounds. HILIC methods typically use high concentration of buffer salts for improved chromatography performance. It is therefore necessary to analyze the adduct formation in HILIC metabolomics. To this end, we developed co-variant ion analysis (COVINA) to investigate the metabolite adduct formation. Using this tool, we completely annotated 201 adduct and fragment ions of 10 metabolites. Many of the metabolite adduct ions are found to contain cluster ions of mobile phase additives. We further utilized COVINA to find the major ionization forms of metabolites. Our results show that for some metabolites the adduct ion signals can be >200-fold higher than the deprotonated form, offering better sensitivity for targeted metabolomics analysis. Finally, we developed the in-source CID ramping (InCIDR) method to analyze the intensity changes of the adduct and fragment ions of the metabolites. Our analysis demonstrates a promising method to distinguish the protonated/deprotonated ions of the metabolites from the adduct and fragment ions.

Review the normal anatomy and physiology of the hypothalamic-pituitary-gland axes; highlight differences between the anterior and posterior pituitary; emphasize mechanistic connections between the endocrine gland and embryology/anatomy.

Understand the etiologic classification of diabetes mellitus; understand the diagnostic criteria for prediabetes and diabetes mellitus; understand the pathophysiologic principles behind T1DM and T2DM.

Review pancreatic anatomy relevant to diabetes mellitus; understand how blood glucose is maintained in the normal range; review insulin and glucagon action; learn the location and regulation of various subtypes of glucose transporters (GLUTs); learn the location and kinetic differences between hexokinase and glucokinase; review factors that promote insulin secretion from the pancreatic β-cell.

Understand why most thyroid nodules are benign; know the general approach for a thyroid nodule work-up; review the causes of thyroid cancer.

Emphasize mechanistic connections between the endocrine gland and embryology/anatomy; describe defects in thyroid development; review thyroid hormone synthesis; explore negative feedback in the HPT axis; and review peripheral metabolism of thyroid hormones.

Understand the vocabulary used to describe thyroid functional diseases; review the signs, symptoms, and causes of hypothyroidism; review the signs, symptoms, and causes of thyrotoxicosis; and present the euthyroid sick syndrome.

Emphasize mechanistic connections between the endocrine gland and embryology/anatomy; understand the mechanism of action of steroid hormones; be able to identify the three classes of adrenal steroids and their main actions; understand the regulation of adrenal steroid secretion.