H and ex vivo2 H magnetic resonance spectroscopy before and during hyperinsulinemiceuglycemic clamps with isotope dilution. Mice underwent identical clamp procedures and hepatic transcriptome analyses.RESULTS. PO administration decreased whole-body, hepatic, and adipose tissue insulin sensitivity by 25%, 15%, and 34%, respectively. Hepatic triglyceride and ATP content rose by 35% and 16%, respectively. Hepatic gluconeogenesis increased by 70%, and net glycogenolysis declined by 20%. Mouse transcriptomics revealed that PO differentially regulates predicted upstream regulators and pathways, including LPS, members of the TLR and PPAR families, NF-κB, and TNF-related weak inducer of apoptosis (TWEAK). CONCLUSION.Saturated fat ingestion rapidly increases hepatic lipid storage, energy metabolism, and insulin resistance. This is accompanied by regulation of hepatic gene expression and signaling that may contribute to development of NAFLD. PO results in increased circulating TG, glucagon, and incretins. After PO administration, TG in plasma rose by 59% (area under the time curve [AUC], P < 0.001) and by 156% in chylomicrons (AUC, P = 0.009) (Figure 2A). The AUC for plasma free fatty acids (FFA) ( Figure 2B) and insulin concentrations ( Figure 2C) was unchanged, while the AUC for plasma C-peptide was 28% higher after PO ingestion versus VCL (P < 0.005, Figure 2D). Of note, FFA were increased at 300, 420, and 480 minutes. Blood glucose levels were not different between PO-and VCL-treated groups ( Figure 2E). Plasma glucagon rose by 41% (AUC, P < 0.0001) only after PO ingestion ( Figure 2F). Also, glucagon-like peptide 1 (GLP-1) and gastric inhibitory polypeptide (GIP) levels were markedly increased and remained elevated after PO ingestion (both P < 0.005) (Supplemental Figure 2; supplemental material available online with this article; https://doi.org/10.1172/ JCI89444DS1). Circulating levels of TNF-α, IL-6, fetuin-A, chemerin, omentin, and cortisol were not different between PO and VCL groups (P > 0.5 for all) (Supplemental Table 1). REGISTRATION. The Journal of Clinical Investigation C L I N I C A L M E D I C I N EPO induces insulin resistance at whole-body, liver, and adipose tissue levels. Insulin sensitivity was measured using hyperinsulinemic-euglycemic clamp tests in healthy humans. Steady state was reached (Supplemental Figure 1), and pertinent parameters were analyzed during this time. PO ingestion reduced WBIS by 25% compared with VCL treatment (P = 0.0005, Figure 3A). Furthermore, after PO, volunteers also showed a decrease of 22% (P = 0.002) in the rate of glucose disappearance (Rd), mostly due to a 33% (P = 0.01) reduction in glucose oxidation (GOX), while the rate of nonoxidative glucose disposal remained unchanged

Fasting hyperglycemia is due to excessive glucose production in people with either IFG/NGT or IFG/IGT. Both insulin action and postprandial glucose concentrations are normal in IFG/NGT but abnormal in IFG/IGT. This finding suggests that hepatic and extrahepatic insulin resistance causes or exacerbates postprandial glucose intolerance in IFG/IGT. Elevated gluconeogenesis in the fasting state in IFG/NGT and impaired insulin-induced suppression of both gluconeogenesis and glycogenolysis in IFG/IGT suggest that alteration in the regulation of these pathways occurs early in the evolution of type 2 diabetes.

The contributions of hepatic glycogenolysis to fasting glucose production and direct pathway to hepatic glycogen synthesis were quantified in eight type 1 diabetic patients and nine healthy control subjects by ingestion of 2 H 2 O and acetaminophen before breakfast followed by analysis of urinary water and acetaminophen glucuronide. After overnight fasting, enrichment of glucuronide position 5 relative to body water (G5/body water) was significantly higher in type 1 diabetic patients compared with control subjects, indicating a reduced contribution of glycogenolysis to glucose production (38 ؎ 3 vs. 46 ؎ 2%). Following breakfast, G5/body water was significantly higher in type 1 diabetic patients, indicating a smaller direct pathway contribution to glycogen synthesis (47 ؎ 2 vs. 59 ؎ 2%). Glucuronide hydrogen 2 enrichment (G2) was equivalent to body water during fasting (G2/body water 0.94 ؎ 0.03 and 1.02 ؎ 0.06 for control and type 1 diabetic subjects, respectively) but was significantly lower after breakfast (G2/body water 0.78 ؎ 0.03 and 0.82 ؎ 0.05 for control and type 1 diabetic subjects, respectively). The reduced postprandial G2 levels reflect incomplete glucose-6-phosphate-fructose-6-phosphate exchange or glycogen synthesis from dietary galactose. Unlike current measurements of human hepatic glycogen metabolism, the 2 H 2 O/acetaminophen assay does not require specialized on-site clinical equipment or personnel. Diabetes

This paper reports the first application of high-resolution 1 H NMR spectroscopy to the plasma of five juveniles with glycogen storage disease type 1a (GSD1a), permitting the characterisation of the plasma metabolic profile and the identification of alterations relative to a set of control samples. The relaxation-weighted spectra allowed changes in low molecular weight compounds to be detected more clearly, whereas diffusion-edited spectra were used to characterise the plasma lipoprotein profile. Low molecular weight metabolites with altered levels in most patients were lactate, ketone bodies, acetate, creatine/creatinine and glucose. One of the patients showed distinctively lower glucose levels and higher lactate and ketone body contents, suggesting poorer metabolic control of the disease compared with other patients. In addition, a metabolite tentatively identified as a-hydroxyisobutyrate was only detected in the spectra of GSD1a plasmas, representing, therefore, a possible novel GSD1a biomarker. Total lipoprotein contents were higher in the plasma from GSD1a patients. Furthermore, lower HDL and higher VLDL þ LDL levels also characterised the plasma of these patients. Preliminary results on principal component analysis of 1 H NMR spectra allowed a clear separation between GSD1a and control plasmas. The specificity of the changes observed to GSD1a is discussed, together with the recognised potential of NMR and pattern recognition methods for aiding the diagnosis of GSD1a.

Plasma glucose, insulin and glucose tolerance were quantified in diabetic Goto-Kakizaki (GK) rats (342+/-45 g, n = 5) and compared with weight-matched non-diabetic Wistars (307+/-30 g, n = 8). Compared to Wistars, GK rats had higher fasting plasma insulin (219+/-50 versus 44+/-14 pmol/l, P<0.002) and glucose (9.2+/-2.3 versus 5.5+/-0.5 mmol/l, P<0.025). GK rats showed impaired glucose tolerance (IPGTT 2 h plasma glucose=14+/-1.5 versus 6.4+/-0.1 mmol/l, P<0.001). Endogenous glucose production (EGP) from glycogenolysis, phosphoenolpyruvate (PEP) and glycerol after 6 hours of fasting was quantified by a primed infusion of [U-(13)C]glucose and (2)H(2)O tracers and (2)H/(13)C NMR analysis of plasma glucose. EGP was higher in GK compared to Wistar rats (191+/-16 versus 104+/-27 mumol/kg per min, P<0.005). This was sustained by increased gluconeogenesis from PEP (85+/-12 versus 35+/-4 mumol/kg per min, P<0.02). Gluconeogenesis from glycerol was not different (20+/-3 in Wistar versus 30+/-6 mumol/kg per min for GK), and glycogenolysis fluxes were also not significantly different (76+/-23 mumol/kg per min for GK versus 52+/-19 mumol/kg per min for Wistar). The Cori cycle accounted for most of PEP gluconeogenesis in both Wistar and GK rats (85+/-15% and 77+/-10%, respectively). Therefore, increased gluconeogenesis in GK rats is largely sustained by increased Cori cycling while the maintenance of glycogenolysis indicates a failure in hepatic autoregulation of EGP.

Quantification of 2 H and 13 C enrichment distributions in human urinary glucuronide following ingestion of 2 H 2 O and 13 C gluconeogenic tracers was achieved by NMR spectroscopy of the 1,2-O-isopropylidene-a-D-glucofuranurono-6,3-lactone and 5-O-acetyl-1,2-O-isopropylidene-a-D-glucofuranurono-6,3-lactone derivatives. The derivatization process is simple and can be applied to any glucuronide species. The derivatives are highly soluble in acetonitrile and generate well-resolved and narrow 2 H and 13 C NMR signals.The 1,2-O-isopropylidene-a-D-glucofuranurono-6,3-lactone derivative provided resolution of the six glucuronide 13 C signals and numerous 13 C isotopomer populations through one-and two-bond 13 C-13 C-coupling, while the 5-O-acetyl-1,2-Oisopropylidene-a-D-glucofuranurono-6,3-lactone derivative provided complete resolution of the 2 H NMR signals for the five glucuronide hydrogens. The isopropylidene methyl signals were also resolved and provided an internal 2 H enrichment standard following the acetonation of glucuronolactone with deuterated acetone.

BACKGROUND.While saturated fat intake leads to insulin resistance and nonalcoholic fatty liver, Mediterranean-like diets enriched in monounsaturated fatty acids (MUFA) may have beneficial effects. This study examined effects of MUFA on tissue-specific insulin sensitivity and energy metabolism. METHODS.A randomized placebo-controlled cross-over study enrolled 16 glucose-tolerant volunteers to receive either oil (OIL, ~1.18 g/kg), rich in MUFA, or vehicle (VCL, water) on 2 occasions. Insulin sensitivity was assessed during preclamp and hyperinsulinemic-euglycemic clamp conditions. Ingestion of 2 H 2 O/acetaminophen was combined with [6,6-2 H 2 ]glucose infusion and in vivo 13 C/ 31 P/ 1 H/ ex vivo 2 H-magnet resonance spectroscopy to quantify hepatic glucose and energy fluxes. RESULTS.OIL increased plasma triglycerides and oleic acid concentrations by 44% and 66% compared with VCL. Upon OIL intervention, preclamp hepatic and whole-body insulin sensitivity markedly decreased by 28% and 27%, respectively, along with 61% higher rates of hepatic gluconeogenesis and 32% lower rates of net glycogenolysis, while hepatic triglyceride and ATP concentrations did not differ from VCL. During insulin stimulation hepatic and whole-body insulin sensitivity were reduced by 21% and 25%, respectively, after OIL ingestion compared with that in controls. CONCLUSION.A single MUFA-load suffices to induce insulin resistance but affects neither hepatic triglycerides nor energy-rich phosphates. These data indicate that amount of ingested fat, rather than its composition, primarily determines the development of acute insulin resistance. TRIAL REGISTRATION. ClinicalTrials.gov NCT01736202.

OBJECTIVEIntravenous insulin infusion partly improves liver glucose fluxes in type 1 diabetes (T1D). This study tests the hypothesis that continuous subcutaneous insulin infusion (CSII) normalizes hepatic glycogen metabolism.RESEARCH DESIGN AND METHODST1D with poor glycemic control (T1Dp; HbA1c: 8.5 ± 0.4%), T1D with improved glycemic control on CSII (T1Di; 7.0 ± 0.3%), and healthy humans (control subjects [CON]; 5.2 ± 0.4%) were studied. Net hepatic glycogen synthesis and glycogenolysis were measured with in vivo 13C magnetic resonance spectroscopy. Endogenous glucose production (EGP) and gluconeogenesis (GNG) were assessed with [6,6-2H2]glucose, glycogen phosphorylase (GP) flux, and gluconeogenic fluxes with 2H2O/paracetamol.RESULTSWhen compared with CON, net glycogen synthesis was 70% lower in T1Dp (P = 0.038) but not different in T1Di. During fasting, T1Dp had 25 and 42% higher EGP than T1Di (P = 0.004) and CON (P < 0.001; T1Di vs. CON: P = NS). GNG was 74 and 67% higher in T1Dp than in T1Di (P = 0.002) and CON (P = 0.001). In T1Dp, GP flux (7.0 ± 1.6 μmol ⋅ kg−1 ⋅ min−1) was twofold higher than net glycogenolysis, but comparable in T1Di and CON (3.7 ± 0.8 and 4.9 ± 1.0 μmol ⋅ kg−1 ⋅ min−1). Thus T1Dp exhibited glycogen cycling (3.5 ± 2.0 μmol ⋅ kg−1 ⋅ min−1), which accounted for 47% of GP flux.CONCLUSIONSPoorly controlled T1D not only exhibits augmented fasting gluconeogenesis but also increased glycogen cycling. Intensified subcutaneous insulin treatment restores these abnormalities, indicating that hepatic glucose metabolism is not irreversibly altered even in long-standing T1D.