Whole-body Glucose Utilization Research Articles

Aiding Cancer's "Sweet Tooth": Role of Hexokinases in Metabolic Reprogramming.

Hexokinases (HKs) convert hexose sugars to hexose-6-phosphate, thus trapping them inside cells to meet the synthetic and energetic demands. HKs participate in various standard and altered physiological processes, including cancer, primarily through the reprogramming of cellular metabolism. Four canonical HKs have been identified with different expression patterns across tissues. HKs 1-3 play a role in glucose utilization, whereas HK 4 (glucokinase, GCK) also acts as a glucose sensor. Recently, a novel fifth HK, hexokinase domain containing 1 (HKDC1), has been identified, which plays a role in whole-body glucose utilization and insulin sensitivity. Beyond the metabolic functions, HKDC1 is differentially expressed in many forms of human cancer. This review focuses on the role of HKs, particularly HKDC1, in metabolic reprogramming and cancer progression.

The Role of Hexokinase Domain Containing Protein-1 in Glucose Regulation During Pregnancy.

Gestational diabetes mellitus (GDM) is a common pregnancy complication conferring an increased risk to the individual of developing type 2 diabetes. As such, a thorough understanding of the pathophysiology of GDM is warranted. Hexokinase domain containing protein-1 (HKDC1) is a recently discovered protein containing hexokinase activity which has been shown to be associated with glucose metabolism during pregnancy. Here, we discuss recent evidence suggesting roles for the novel HKDC1 in gestational glucose homeostasis and the development of GDM and overt diabetes. Genome-wide association studies identified variants of the HKDC1 gene associated with maternal glucose metabolism. Studies modulating HKDC1 protein expression in pregnant mice demonstrate that HKDC1 has roles in whole-body glucose utilization and nutrient balance, with liver-specific HKDC1 influencing insulin sensitivity, glucose tolerance, gluconeogenesis, and ketone production. HKDC1 has important roles in maintaining maternal glucose homeostasis extending beyond traditional hexokinase functions and may serve as a potential therapeutic target.

Reduced Insulin Sensitivity in Patients with Myeloid Malignancies and Clonal Hematopoiesis Mutations

Cancer survivors are at increased risk for insulin resistance, which can lead to diabetes mellitus, dyslipidemia, and cardiovascular disease. Whether susceptibility to insulin resistance is due to shared onco-metabolic risk factors, like obesity and older age, or if it originates from malignancy and its treatment is unknown. We investigated metabolic and malignancy-associated risk factors and their effect on insulin sensitivity in patients with treated hematological malignancies. Due to previous glucocorticoid exposure, we hypothesized that patients with lymphoid malignancies would be less insulin sensitive. Nineteen adult patients with treated hematological malignancies but without diabetes mellitus were evaluated with a 2-hour, 75-gram, oral glucose tolerance test (OGTT) and a hyperinsulinemic-euglycemic clamp. Hyperinsulinemic-euglycemic clamps were performed to measure steady-state glucose infusion rate (M-value) as an indicator of whole-body glucose utilization during high-dose insulin stimulation. Decreasing M-values reflect increasing insulin resistance. Patient characteristics were analyzed and compared with OGTT and clamp results. The majority of individuals had myeloid malignancies [MDS/AML (n=7) and CML/MPN (n = 4)]. The remainder were diagnosed with NHL (n=7) and CML lymphoid blast crisis (n=1). All patients were treated before metabolic testing. Sixteen individuals (84%) received cytotoxic chemotherapy and 3 (16%) were administered only tyrosine kinase inhibitors. Median lines of systemic treatment were 2 (range, 1-6), and median time from cancer diagnosis to metabolic testing was 10 months (range, 3-132). Immediately before OGTTs and clamps, 10 (53%) patients were in a complete response, and 9 (47%) had persistent/stable disease. During OGTTs, impaired fasting glucose (plasma glucose 100-125 mg/dL) and impaired glucose tolerance (2-hour plasma glucose 140-199 mg/dL) were respectively observed in 5 (26%) and 6 (32%) patients. M-values were decreased in patients with impaired glucose tolerance. Overweight and obese cancer patients also had lower M-values than individuals with normal body mass indices (BMI) (Table 1). There were otherwise no differences in insulin resistance per age, sex, performance status, ethnicity, or diabetes family history. Differences in insulin resistance were not observed based on exposure to cytotoxic chemotherapy, non-cranial irradiation, numbers of lines of treatment, response, or malignancy duration. Previous glucocorticoid exposure did not impact insulin sensitivity. Surprisingly, patients with myeloid malignancies demonstrated lower M-values (6.92 + 0.54 vs. 12.11 + 1.89; p=0.026) than did those with lymphoid cancers. Acquired mutations involved with myeloid malignancies and clonal hematopoiesis (CH) can increase the risk for atherosclerosis and cardiovascular disease, which are sequelae of insulin resistance. Accordingly, insulin resistance was increased in patients with both a myeloid malignancy and mutations involving either DNMT3A, TET2, ASXL1, or JAK2 compared to myeloid malignancies with unknown/absent CH mutations or lymphoid cancers (6.71 + 0.77 vs. 7.28 + 0.72 vs. 12.11 + 1.89; p=0.031). After adjustment for interaction effect in linear regression modeling, both myeloid malignancy and elevated BMI continued to predict lower insulin sensitivity (Table 2). Pre-diabetes was diagnosed by fasting glucose or OGTT in 26-32% of patients with treated hematological cancers, more than double the prevalence of impaired fasting glucose or impaired glucose tolerance reported by the National Health and Nutrition Examination Survey III. Contrary to our hypothesis, lymphoid cancers and short-term steroids did not contribute to insulin resistance. Instead, myeloid malignancies and CH mutations were associated with reduced insulin sensitivity. Interventions targeting energy balance and obesity will be needed to prevent the harmful sequelae of insulin resistance in long-term cancer survivors. Alternatively, elimination of CH mutations could prove to be a novel treatment of insulin resistance. Disclosures Dholaria: bms: Research Funding; Poseida: Research Funding; Angiocrine: Research Funding; Takeda: Research Funding; J&J: Research Funding.

210-LB: Combined Use of Oral Standard Cookie Meal (OSC) and the Newly Revised Insulin Suppression Test for the Evaluation of Whole Body (WB), Muscle, and Hepatic Glucose Clearance (GC) Under Endogenous or Exogenous Insulin and Its Clinical Usefulness

Adequate insulin action (sensitivity: IS) in muscle, liver and WB is the most important metabolic factor in diabetes, especially in S-GLUT-2 user. Oral intake of hypoglycemic agents followed by the standard 75g starch including 15% maltose (OSC, Saraya Co.), which is digested and absorbed into blood glucose at a constant rate for 2 hours, minus urinary glucose loss, divided by the steady state PG at 2h equals the GC in WB under the endogenous insulin secretion. In SGLUT-2 user(HbA1c, 6.8%, BMI, 25.3), this was significantly low(4.2±0.5,SE ml/kg/min, n=6 vs. normal, 8.7,n=10). Average of 1 and 2h insulin was low,10.5 vs. normal,30μU/ml. They are recommended to keep BMI below 23, upper normal range of IS, and use GLP-1 analog or insulin to compensate insufficient IS. This approximation is further evaluated using the revised half dose of glucose and insulin infusion, 3mg, 0.39mU/kg/min, with octreotide 50μg SC for muscle, 100μg for whole body, 30min before the test. Under this exogenous insulin(19.1μU/ml, at 2-4h), 2h GC is confirmed significantly low (2.3±0.1,n=6 vs. control, 3.7±1.0, n=10) reflecting impaired muscle GC in SGLUT-2 user. Additional intake of OSC under continued subcutaneous insulin(1.5 rate over iv), gives the 2h decreased GC (5.84, vs. normal, 9.5,) of WB, and the difference corresponding to hepatic GC, which was significantly low. Better GC was obtained for non-SGLUT groups on oral agents with or without insulin secretagogues. Conclusion Combined use of OSC and simplified IS test enables to evaluate muscle, hepatic and WB glucose utilization under endogenous or exogenous insulin. The method is useful to detect and clarify the various mechanisms of insulin resistance in diabetes, metabolic syndrome, heart failure, Alzheimer disease or fatty liver, etc. Disclosure Y. Harano: None.

4164 Body mass index, not chemotherapy is the major predictor of insulin resistance in patients with hematological malignancies

OBJECTIVES/GOALS: Cancer survivors are at increased risk for type 2 diabetes mellitus.It is not clear if diabetes susceptibility is due to shared risk factors for cancer and diabetes, such as obesity, or if it is directly related to cancer and its treatment. We investigated the association between malignancy and insulin resistance, a major risk factor for diabetes. METHODS/STUDY POPULATION: 20 adult patients with treated hematological malignancies and 21 controls without cancer were included in the study. Individuals with pre-existing diabetes were excluded. All patients underwent a 2-hour 75-gram oral glucose tolerance test (OGTT). Hyperinsulinemic-euglycemic clamps were performed to measure the steady-state glucose infusion rate (M-value) as an indicator of whole-body glucose utilization during insulin stimulation. Insulin sensitivity index was calculated by dividing M-value over the steady-state plasma insulin (M/I). RESULTS/ANTICIPATED RESULTS: Fasting or postprandial plasma glucose levels during the OGTT did not differ significantly between malignancy patients and controls (Table 2). Difference in the insulin-stimulated glucose utilization (M-value) was not statistically significant among cancer patients and controls (median, 7.2 [IQR, 6.2-10.4] vs. 7.3 [IQR, 5.5-8.9] mg/kg/min; P = 0.261). M/I index was significantly higher in malignancy patients compared to controls (median, 42.4 mg/kg/min/(µU/ml) [IQR, 33.9-67.2] vs. 23.4 mg/kg/min/(µU/ml) [IQR, 12.9-29.2], P <0.001), however insulin clearance was also lower in the controls. In multivariate analysis, only BMI was significantly associated with M-value (β = −0.2 (95% CI −0.4, −0.1), P = 0.004), and M/I (β = −2 (95% CI −3.4, −0.5), P = 0.009). DISCUSSION/SIGNIFICANCE OF IMPACT: Our data suggest that the major contributor to diabetes development after diagnosis of cancer in adults is obesity-induced insulin resistance, not malignancy related factors. These findings emphasize the importance of obesity management in long-term survival of cancer patients, as diabetes is a risk factor for mortality in this patient population.

MON-642 Genetic Knockout of Intestinal Hexokinase Domain-Containing Protein 1 Affects Whole-Body Glycemic Control and Triglyceride Metabolism

Hexokinase domain-containing protein 1 (HKDC1) is a recently discovered putative fifth hexokinase that is widely expressed in a variety of human and mouse tissues. Previous work indicate that HKDC1 is important for whole-body glucose homeostasis and utilization in pregnancy and aging, and suggest roles for HKDC1 in nonalcoholic fatty liver disease development and progression of hepatocellular carcinoma. Prior work in the lab further showed that global heterozygous-deleted HKDC1 mice exhibit blunted uptake of triglycerides following an olive oil bolus compared to wild-type mice, suggesting a role for intestinal HKDC1 expression in intestinal lipid metabolism (unpublished results). To specifically study the significance of intestinal HKDC1 on whole-body glucose and lipid homeostasis, we utilized Cre-mediated recombination of HKDC1 in which Cre was expressed under the control of the villin gene promoter, creating a mouse model in which HKDC1 expression is specifically deleted in the intestinal epithelium. Quantitative RT-PCR data confirmed the knockout of HKDC1 within the mouse intestine in young and aged mice, while HKDC1 expression in other tissues was comparable to wild-type mice. Next, intestinal HKDC1 knockout mice and their wild-type littermate controls were either maintained on a normal diet or were switched to a high fat diet at 6 weeks of age to simulate the state of impaired glucose tolerance, and the effects of intestinal HKDC1 on glucose and lipid homeostasis were analyzed between 28-34 weeks of age. Mice fed a normal diet did not exhibit any differences in serum glucose or triglyceride during oral/intraperitoneal glucose tolerance tests or oral olive oil bolus, respectively, regardless of intestinal HKDC1 status. Interestingly, mice lacking intestinal HKDC1 that were on a high fat diet demonstrated improved overall glycemic control compared to wild-type mice after the administration of an oral glucose load, all while there were no changes in insulin levels, gluconeogenesis or insulin tolerance related to HKDC1 status. Additionally, introduction of an intraperitoneal glucose load to mice fed a high fat diet did not alter glucose control in the presence or absence of intestinal HKDC1. However, high fat diet-fed mice lacking intestinal HKDC1 did not have a significant increase in serum triglyceride following an oral olive oil bolus, while their stool fat and triglyceride content were comparable to wild-type. Collectively, these data indicate that intestinal HKDC1 has important roles in glucose and triglyceride metabolism within the intestinal epithelium, and further suggest a role in whole-body glucose homeostasis and in the development of insulin resistance and diabetes.

Dietary Fat and Sugar Differentially Affect β-Adrenergic Stimulation of Cardiac ERK and AKT Pathways in C57BL/6 Male Mice Subjected to High-Calorie Feeding

High dietary fat and sugar promote cardiac hypertrophy independently from an increase in blood pressure. The respective contribution that each macronutrient exerts on cardiac growth signaling pathways remains unclear. The goal of this study was to investigate the mechanisms by which high amounts of dietary fat and sugar affect cardiac growth regulatory pathways. Male C57BL/6 mice (9 wk old; n=20/group) were fed a standard rodent diet (STD; kcal% protein-fat-carbohydrate, 29-17-54), a high-fat diet (HFD; 20-60-20), a high-fat and high-sugar Western diet (WD; 20-45-35), a high-sugar diet with mixed carbohydrates (HCD; 20-10-70), or a high-sucrose diet (HSD; 20-10-70). Body composition was assessed weekly by EchoMRI. Whole-body glucose utilization was assessed with an intraperitoneal glucose tolerance test. After 6 wk on diets, mice were treated with saline or 20 mg/kg isoproterenol (ISO), and the activity of cardiac growth regulatory pathways was analyzed by immunoblotting. Data were analyzed by ANOVA with data from the STD group included for references only. Compared with HCD and HSD, WD and HFD increased body fat mass 2.7- to 3.8-fold (P<0.001), induced glucose intolerance (P<0.001), and increased insulin concentrations >1.5-fold (P<0.05), thereby enhancing basal and ISO-stimulated AKT phosphorylation at both threonine 308 and serine 473 residues (+25-63%; P<0.05). Compared with HFD, the high-sugar diets potentiated ISO-mediated stimulation of the glucose-sensitive kinases PYK2 (>47%; P<0.05 for HCD and HSD) and ERK (>34%; P<0.05 for WD, HCD, and HSD), thereby leading to increased phosphorylation of protein synthesis regulator S6K1 at threonine 389 residue (>64%; P<0.05 for WD, HCD, and HSD). Dietary fat and sugar affect cardiac growth signaling pathways in C57BL/6 mice through distinct and additive mechanisms. The findings may provide new insights into the role of overnutrition in pathological cardiac remodeling.

Metabolic effects of skeletal muscle-specific deletion of beta-arrestin-1 and -2 in mice.

Type 2 diabetes (T2D) has become a major health problem worldwide. Skeletal muscle (SKM) is the key tissue for whole-body glucose disposal and utilization. New drugs aimed at improving insulin sensitivity of SKM would greatly expand available therapeutic options. β-arrestin-1 and -2 (Barr1 and Barr2, respectively) are two intracellular proteins best known for their ability to mediate the desensitization and internalization of G protein-coupled receptors (GPCRs). Recent studies suggest that Barr1 and Barr2 regulate several important metabolic functions including insulin release and hepatic glucose production. Since SKM expresses many GPCRs, including the metabolically important β2-adrenergic receptor, the goal of this study was to examine the potential roles of Barr1 and Barr2 in regulating SKM and whole-body glucose metabolism. Using SKM-specific knockout (KO) mouse lines, we showed that the loss of SKM Barr2, but not of SKM Barr1, resulted in mild improvements in glucose tolerance in diet-induced obese mice. SKM-specific Barr1- and Barr2-KO mice did not show any significant differences in exercise performance. However, lack of SKM Barr2 led to increased glycogen breakdown following a treadmill exercise challenge. Interestingly, mice that lacked both Barr1 and Barr2 in SKM showed no significant metabolic phenotypes. Thus, somewhat surprisingly, our data indicate that SKM β-arrestins play only rather subtle roles (SKM Barr2) in regulating whole-body glucose homeostasis and SKM insulin sensitivity.

Neuronal hypothalamic regulation of body metabolism and bone density is galanin dependent

In the brain, the ventral hypothalamus (VHT) regulates energy and bone metabolism. Whether this regulation uses the same or different neuronal circuits is unknown. Alteration of AP1 signaling in the VHT increases energy expenditure, glucose utilization, and bone density, yet the specific neurons responsible for each or all of these phenotypes are not identified. Using neuron-specific, genetically targeted AP1 alterations as a tool in adult mice, we found that agouti-related peptide-expressing (AgRP-expressing) or proopiomelanocortin-expressing (POMC-expressing) neurons, predominantly present in the arcuate nucleus (ARC) within the VHT, stimulate whole-body energy expenditure, glucose utilization, and bone formation and density, although their effects on bone resorption differed. In contrast, AP1 alterations in steroidogenic factor 1-expressing (SF1-expressing) neurons, present in the ventromedial hypothalamus (VMH), increase energy but decrease bone density, suggesting that these effects are independent. Altered AP1 signaling also increased the level of the neuromediator galanin in the hypothalamus. Global galanin deletion (VHT galanin silencing using shRNA) or pharmacological galanin receptor blockade counteracted the observed effects on energy and bone. Thus, AP1 antagonism reveals that AgRP- and POMC-expressing neurons can stimulate body metabolism and increase bone density, with galanin acting as a central downstream effector. The results obtained with SF1-expressing neurons, however, indicate that bone homeostasis is not always dictated by the global energy status, and vice versa.

Activation of SF1 Neurons in the Ventromedial Hypothalamus by DREADD Technology Increases Insulin Sensitivity in Peripheral Tissues.

The ventromedial hypothalamus (VMH) regulates glucose and energy metabolism in mammals. Optogenetic stimulation of VMH neurons that express steroidogenic factor 1 (SF1) induces hyperglycemia. However, leptin acting via the VMH stimulates whole-body glucose utilization and insulin sensitivity in some peripheral tissues, and this effect of leptin appears to be mediated by SF1 neurons. We examined the effects of activation of SF1 neurons with DREADD (designer receptors exclusively activated by designer drugs) technology. Activation of SF1 neurons by an intraperitoneal injection of clozapine-N-oxide (CNO), a specific hM3Dq ligand, reduced food intake and increased energy expenditure in mice expressing hM3Dq in SF1 neurons. It also increased whole-body glucose utilization and glucose uptake in red-type skeletal muscle, heart, and interscapular brown adipose tissue, as well as glucose production and glycogen phosphorylase a activity in the liver, thereby maintaining blood glucose levels. During hyperinsulinemic-euglycemic clamp, such activation of SF1 neurons increased insulin-induced glucose uptake in the same peripheral tissues and tended to enhance insulin-induced suppression of glucose production by suppressing gluconeogenic gene expression and glycogen phosphorylase a activity in the liver. DREADD technology is thus an important tool for studies of the role of the brain in the regulation of insulin sensitivity in peripheral tissues.

Systemic and tissue-specific effects of aliskiren on the RAAS and carbohydrate/lipid metabolism in obese patients with hypertension

Aliskiren penetrates adipose and skeletal muscle in hypertensive patients with abdominal obesity and reduces renin–angiotensin–aldosterone system activity. After discontinuation, blood pressure–lowering effects are observed possibly through drug–tissue binding. We performed microdialysis evaluation of adipose tissue and skeletal muscle before and during an insulin-modified frequently sampled intravenous glucose tolerance test (IM-FSIGT). Aliskiren 300 mg (n = 8) or amlodipine 5 mg (n = 8) once daily were administered during a 12-week randomized treatment period. Aliskiren elicited variable changes in median interstitial angiotensin II (Ang II) in adipose (2.60–1.30 fmol/mL) and skeletal muscle (2.23–0.68 fmol/mL); amlodipine tended to increase adipose and skeletal muscle Ang II (P = .066 for skeletal muscle treatment difference). Glucose/insulin increased median plasma Ang II 1 hour after glucose injection (1.04–2.50 fmol/mL; P = .001), which was markedly attenuated by aliskiren but not amlodipine. Aliskiren increased glucose disposition index (P = .012) and tended to increase acute insulin response to glucose (P = .067). Fasting adipose glycerol (−17%; P = .064) and fasting muscle glucose dialysate (−17%; P = .025) were decreased by aliskiren but not amlodipine. In summary, aliskiren decreased Ang II production in response to glucose/insulin stimulus and elicited metabolic effects in adipose and skeletal muscle suggestive of increased whole-body glucose utilization.

Glucose requirements of an activated immune system in lactating Holstein cows

Accurately quantifying activated immune system energy requirements in vivo is difficult, but a better understanding may advance strategies to maximize animal productivity. Study objectives were to estimate whole-body glucose utilization following an i.v. endotoxin challenge. Lactating Holstein cows were jugular catheterized and assigned 1 of 3 bolus treatments: control (CON; 5 mL of saline; n = 6), lipopolysaccharide (LPS)-administered (LPS-C; 1.5 μg/kg of body weight; Escherichia coli 055:B5; n = 6), and LPS + euglycemic clamp (LPS-Eu; 1.5 μg/kg of body weight; 50% glucose solution infusion; n = 6). After LPS administration, blood glucose was determined every 10 min and glucose infusion rates were adjusted in LPS-Eu cows to maintain euglycemia for 720 min. Blood samples were obtained 180, 360, 540, and 720 min postbolus for further analysis. Cows were milked 360 and 720 min postbolus. Blood glucose was increased 84% in LPS-administered cows for up to 150 min postbolus; thereafter, circulating glucose was decreased 30% in LPS-C relative to LPS-Eu and CON cows. Mild hyperthermia (+0.5°C) occurred between 30 and 90 min postbolus in LPS-administered relative to CON cows; thereafter, rectal temperature did not differ between treatments. Milk yield and lactose percentage were decreased 80 and 11%, respectively, in LPS-administered relative to CON cows. Circulating insulin was increased 4 fold and nonesterified fatty acids, β-hydroxybutyrate, and ionized Ca were decreased ∼50% in LPS-administered compared with CON cows. Plasma l-lactate, haptoglobin, and serum amyloid A increased ∼160, 260, and 75%, respectively, in LPS-administered relative to CON cows. Overall, LPS-binding protein was increased 87% in LPS-administered relative to CON cows; however, at 720 min, it was decreased 25% in LPS-Eu compared with LPS-C cows. White blood cell count decreased ∼90% in LPS-administered cows at 180 min and progressively increased to ∼50% of CON values by 720 min. Total glucose deficit during the 720 min following LPS administration was calculated as the decrease in the amount of glucose required to synthesize milk (due to the decrease in milk yield relative to prebolus levels) plus the amount of glucose infused to maintain euglycemia (in LPS-Eu cows only) and was 461, 1,259, and 1,553 g for CON, LPS-C, and LPS-Eu cows, respectively. Our data indicate an acutely activated immune system uses >1 kg of glucose within 720 min and maintaining euglycemia did not rescue milk synthesis.

Metabolic abnormalities induced by mitochondrial dysfunction in skeletal muscle of the renal carcinoma Eker (TSC2+/-) rat model.

Tuberous sclerosis complex 2 (TSC2) is a mediator of insulin signal transduction, and a loss of function in TSC2 induces hyperactivation of mTORC1 pathway, which leads to tumorigenesis. We have previously demonstrated that Eker rat model, which is heterozygous for a TSC2 mutation, exhibits hyperglycemia and hyperketonemia. The present study was to investigate whether these changes also can affect metabolism in skeletal muscle of the Eker rat. Wild-type (TSC2+/+) and Eker (TSC2+/-) rats underwent an oral glucose tolerance test, and the latter showed decrease in whole-body glucose utilization. Additionally, reductions in the expression of glycolysis-, lipolysis-, and ketone body-related genes in skeletal muscle were observed in Eker rats. Furthermore, ATP content and mitochondrial DNA copy number were lower in skeletal muscle of Eker rats. These data demonstrate that heterozygous to mutation TSC2 not only affects the liver metabolism, but also skeletal muscle metabolism, via mitochondrial dysfunction.

Role of reduced insulin-stimulated bone blood flow in the pathogenesis of metabolic insulin resistance and diabetic bone fragility

Worldwide, 387 million adults live with type 2 diabetes (T2D) and an additional 205 million cases are projected by 2035. Because T2D has numerous complications, there is significant morbidity and mortality associated with the disease. Identification of early events in the pathogenesis of insulin resistance and T2D might lead to more effective treatments that would mitigate health and monetary costs. Here, we present our hypothesis that impaired bone blood flow is an early event in the pathogenesis of whole-body metabolic insulin resistance that ultimately leads to T2D. Two recent developments in different fields form the basis for this hypothesis. First, reduced vascular function has been identified as an early event in the development of T2D. In particular, before the onset of tissue or whole body metabolic insulin resistance, insulin-stimulated, endothelium-mediated skeletal muscle blood flow is impaired. Insulin resistance of the vascular endothelium reduces delivery of insulin and glucose to skeletal muscle, which leads to tissue and whole-body metabolic insulin resistance. Second is the paradigm-shifting discovery that the skeleton has an endocrine function that is essential for maintenance of whole-body glucose homeostasis. Specifically, in response to insulin signaling, osteoblasts secret osteocalcin, which stimulates pancreatic insulin production and enhances insulin sensitivity in skeletal muscle, adipose, and liver. Furthermore, the skeleton is not metabolically inert, but contributes to whole-body glucose utilization, consuming 20% that of skeletal muscle and 50% that of white adipose tissue. Without insulin signaling or without osteocalcin activity, experimental animals become hyperglycemic and insulin resistant. Currently, it is not known if insulin-stimulated, endothelium-mediated blood flow to bone plays a role in the development of whole body metabolic insulin resistance. We hypothesize that it is a key, early event. Microvascular dysfunction is a primary cause of diabetic nephropathy, retinopathy and neuropathy and poor bone blood flow is associated with bone loss. Therefore, we also hypothesize that dysfunction of the bone vascular endothelium contributes to the bone fragility observed in T2D. The most important consequence of our-dual hypothesis is the public health significance. Namely, identification of the proximal cause of T2D and associated bone complications allows pursuit of the appropriate therapeutic target to treat and prevent T2D. If our hypothesis that reduced bone blood flow is an early event in the pathogenesis of T2D and diabetic bone fragility is correct, then the endothelium of the bone vasculature should be a therapeutic target.

Characterization of the role of sphingomyelin synthase 2 in glucose metabolism in whole-body and peripheral tissues in mice

Sphingomyelin synthase 2 (SMS2) is a proposed potential therapeutic target for obesity and insulin resistance. However, the contributions of SMS2 to glucose metabolism in tissues and its possible therapeutic mechanisms remain unclear. Thus, to determine whole-body glucose utilization and the contributions of each insulin-targeted tissue to glucose uptake, we performed a glucose kinetics study, using the radiolabeled glucose analog 18F-2-fluoro-2-deoxy-D-glucose (18F-FDG), in wild-type (WT) and SMS2 knockout (KO) mice. Insulin signaling was enhanced in the liver, white adipose tissue and skeletal muscle of SMS2 KO mice compared with those of WT mice. In addition, compared with in WT mice, blood clearance of 18F-FDG was accelerated in SMS2 KO mice when they were fed either a normal or a high fat diet. 18F-FDG uptake was also increased in insulin-targeted tissues such as skeletal muscle in the SMS2 KO mice. Whereas skeletal muscle sphingolipid content was not clearly affected, plasma levels of very long-chain fatty acid (VLCFA)-containing ceramides were markedly increased in SMS2 KO mice, compared with in WT mice. We also generated liver-conditional SMS2 KO mice and performed glucose and insulin tolerance tests on mice with a high fat diet. However, no significant effect was observed. Thus, our study provided evidence that genetic inhibition of SMS2 elevated glucose clearance through activation of glucose uptake into insulin-targeted tissues such as skeletal muscle by a mechanism independent of hepatic SMS2. Our findings further indicate that this occurs, at least in part, via indirect mechanisms such as elevation of VLCFA-containing ceramides.

Short-term glucagon stimulation test of C-peptide effect on glucose utilization in patients with type 1 diabetes mellitus.

This work aimed to evaluate the use of a four-point glucagon stimulation test of C-peptide effect on glucose utilization in type 1 diabetic patients using a new mathematical model. A group of 32 type 1 diabetic patients and a group of 10 healthy control subjects underwent a four-point glucagon stimulation test with blood sampling at 0, 6, 15 and 30min after 1mg glucagon bolus intravenous administration. Pharmacokinetic and pharmacokinetic/pharmacodynamic models of C-peptide effect on glucose utilization versus area under curve (AUC) were used. A two-sample t test and ANOVA with Bonferroni correction were used to test the significance of differences between parameters. A significant difference between control and patient groups regarding the coefficient of whole-body glucose utilization and AUC C-peptide/AUC glucose ratio (p≪0.001 and p=0.002, respectively) was observed. The high correlation (r=0.97) between modeled coefficient of whole-body glucose utilization and numerically calculated AUC C-peptide/AUC glucose ratio related to entire cohort indicated the stability of used method. The short-term four-point glucagon stimulation test allows the numerically calculated AUC C-peptide/AUC glucose ratio and/or the coefficient of whole-body glucose utilization calculated from model to be used to diagnostically identify type 1 diabetic patients.

Pathological consequences of C-peptide deficiency in insulin-dependent diabetes mellitus.

Diabetes is associated with several complications such as retinopathy, nephropathy, neuropathy and cardiovascular diseases. Currently, insulin is the main used medication for management of insulin-dependent diabetes mellitus (type-1 diabetes). In this metabolic syndrome, in addition to decrease of endogenous insulin, the plasma level of connecting peptide (C-peptide) is also reduced due to beta cell destruction. Studies in the past decade have shown that C-peptide is much more than a byproduct of insulin biosynthesis and possess different biological activities. Therefore, it may be possible that C-peptide deficiency be involved, at least in part, in the development of different complications of diabetes. It has been shown that a small level of remaining C-peptide is associated with significant metabolic benefit. The purpose of this review is to describe beneficial effects of C-peptide replacement on pathological features associated with insulin-dependent diabetes. Also, experimental and clinical findings on the effects of C-peptide on whole-body glucose utilization, adipose tissue metabolism and tissues blood flow are summarized and discussed. The hypoglycemic, antilipolytic and vasodilator effects of C-peptide suggest that it may contribute to fine-tuning of the tissues metabolism under different physiologic or pathologic conditions. Therefore, C-peptide replacement together with the classic insulin therapy may prevent, retard, or ameliorate diabetic complications in patients with type-1 diabetes.

Preclinical characterization of recombinant human tissue kallikrein-1 as a novel treatment for type 2 diabetes mellitus.

Modulation of the kallikrein-kinin system (KKS) has been shown to have beneficial effects on glucose homeostasis and several other physiological responses relevant to the progression of type 2 diabetes mellitus (T2D). The importance of bradykinin and its receptors in mediating these responses is well documented, but the role of tissue kallikrein-1, the protease that generates bradykinin in situ, is much less understood. We developed and tested DM199, recombinant human tissue kallikrein-1 protein (rhKLK-1), as a potential novel therapeutic for T2D. Hyperinsulinemic-euglycemic clamp studies suggest that DM199 increases whole body glucose disposal in non-diabetic rats. Single-dose administration of DM199 in obese db/db mice and ZDF rats, showed an acute, dose-dependent improvement in whole-body glucose utilization. Sub-acute dosing for a week in ZDF rats improved glucose utilization, with a concomitant rise in fasting insulin levels and HOMA1-%B scores. After cessation of sub-acute dosing, fasting blood glucose levels were significantly lower in ZDF rats during a drug wash-out period. Our studies show for the first time that DM199 administration results in acute anti-hyperglycemic effects in several preclinical models, and demonstrate the potential for further development of DM199 as a novel therapeutic for T2D.

Diets enriched in trans-11 vaccenic acid alleviate ectopic lipid accumulation in a rat model of NAFLD and metabolic syndrome

Trans11-18:1 (vaccenic acid, VA) is one of the most predominant naturally occurring trans fats in our food chain and has recently been shown to exert hypolipidemic effects in animal models. In this study, we reveal new mechanism(s) by which VA can alter body fat distribution, energy utilization and dysfunctional lipid metabolism in an animal model of obesity displaying features of the metabolic syndrome (MetS). Obese JCR:LA-cp rats were assigned to a control diet that included dairy-derived fat or the control diet supplemented with 1% VA. VA reduced total body fat (−6%), stimulated adipose tissue redistribution [reduced mesenteric fat (−17%) while increasing inguinal fat mass (29%)] and decreased adipocyte size (−44%) versus control rats. VA supplementation also increased metabolic rate (7%) concomitantly with an increased preference for whole-body glucose utilization for oxidation and increased insulin sensitivity [lower HOMA-IR (−59%)]. Further, VA decreased nonalcoholic fatty liver disease activity scores (−34%) and reduced hepatic (−27%) and intestinal (−39%) triglyceride secretion relative to control diet, while exerting differential transcriptional regulation of SREBP1 and FAS amongst other key genes in the liver and the intestine. Adding VA to dairy fat alleviates features of MetS potentially by remodeling adipose tissue and attenuating ectopic lipid accumulation in a rat model of obesity and MetS. Increasing VA content in the diet (naturally or by fortification) may be a useful approach to maximize the health value of dairy-derived fats.

Putting Rac1 on the Path to Glucose Uptake

In spite of numerous and key advances in our understanding of insulin signaling, the molecular basis for impaired insulin-stimulated glucose uptake underlying insulin resistance remains unclear. Because skeletal muscle accounts for the majority of glucose utilization in the postprandial insulin-regulated state, defects in insulin action in muscle can determine whole-body glucose utilization. In muscle and fat cells, overall similarities exist in the signals emanating from the insulin receptor that mobilize glucose transporter GLUT4 to the membrane. Importantly, defects in the signaling network connecting the activated insulin receptor to GLUT4 vesicles are of particular consequence to insulin resistance in muscle, where, unlike the case of fat cells, GLUT4 expression is not significantly reduced. The complex structural organization of muscle tissue and its associated cells have limited the molecular scrutiny of glucose uptake regulation. Instead, cell culture systems have been instrumental in revealing intricacies in the signals and traffic machinery mobilizing GLUT4 to the membrane. This signaling relay is initiated by insulin receptor substrate (IRS)-1 (and not IRS-2)–associated activation of class I phosphatidylinositol-3-kinase (PI3K). At this point, a signal bifurcation takes place, one arm leading to Akt2 activation, inhibition of its substrate AS160 (a GTPase-activating protein for Rabs) and consequent activation of its target Rab GTPases, which in skeletal muscle cells are Rab8A and Rab13 (1). The other arm leads to Rac1, a Rho-family GTPase (2,3) that enacts a dynamic cycle of cortical actin filament remodeling through the Arp2/3 complex and cofilin (4). Joint activation of these distinct signaling arms is required …