Ultra-stable insulin-glucagon fusion protein exploits an endogenous hepatic switch to mitigate hypoglycemic risk.

The risk of hypoglycemia and its serious medical sequelae restrict insulin replacement therapy for diabetes mellitus. Such adverse clinical impact has motivated development of diverse glucose-responsive technologies, including algorithm-controlled insulin pumps linked to continuous glucose monitors ("closed-loop systems") and glucose-sensing ("smart") insulins. These technologies seek to optimize glycemic control while minimizing hypoglycemic risk. Here, we describe an alternative approach that exploits an endogenous glucose-dependent switch in hepatic physiology: preferential insulin signaling (under hyperglycemic conditions) versus preferential counter-regulatory glucagon signaling (during hypoglycemia). Motivated by prior reports of glucagon-insulin co-infusion, we designed and tested an ultra-stable glucagon-insulin fusion protein whose relative hormonal activities were calibrated by respective modifications; physical stability was concurrently augmented to facilitate formulation, enhance shelf life and expand access. An N-terminal glucagon moiety was stabilized by an α-helix-compatible Lys 13 -Glu 17 lactam bridge; A C-terminal insulin moiety was stabilized as a single chain with foreshortened C domain. Studies in vitro demonstrated (a) resistance to fibrillation on prolonged agitation at 37 °C and (b) dual hormonal signaling activities with appropriate balance. Glucodynamic responses were monitored in rats relative to control fusion proteins lacking one or the other hormonal activity, and continuous intravenous infusion emulated basal subcutaneous therapy. Whereas efficacy in mitigating hyperglycemia was unaffected by the glucagon moiety, the fusion protein enhanced endogenous glucose production under hypoglycemic conditions. Together, these findings provide proof of principle toward a basal glucose-responsive insulin biotechnology of striking simplicity. The fusion protein's augmented stability promises to circumvent the costly cold chain presently constraining global insulin access. Significance Statement: The therapeutic goal of insulin replacement therapy in diabetes is normalization of blood-glucose concentration, which prevents or delays long-term complications. A critical barrier is posed by recurrent hypoglycemic events that results in short- and long-term morbidities. An innovative approach envisions co-injection of glucagon (a counter-regulatory hormone) to exploit a glycemia-dependent hepatic switch in relative hormone responsiveness. To provide an enabling technology, we describe an ultra-stable fusion protein containing insulin- and glucagon moieties. Proof of principle was obtained in rats. A single-chain insulin moiety provides glycemic control whereas a lactam-stabilized glucagon extension mitigates hypoglycemia. This dual-hormone fusion protein promises to provide a basal formulation with reduced risk of hypoglycemia. Resistance to fibrillation may circumvent the cold chain required for global access.

Stimulation of the hepatoportal nerve plexus with focused ultrasound restores glucose homoeostasis in diabetic mice, rats and swine.

Peripheral neurons that sense glucose relay signals of glucose availability to integrative clusters of neurons in the brain. However, the roles of such signalling pathways in the maintenance of glucose homoeostasis and their contribution to disease are unknown. Here we show that the selective activation of the nerve plexus of the hepatic portal system via peripheral focused ultrasound stimulation (pFUS) improves glucose homoeostasis in mice and rats with insulin-resistant diabetes and in swine subject to hyperinsulinemic-euglycaemic clamps. pFUS modulated the activity of sensory projections to the hypothalamus, altered the concentrations of metabolism-regulating neurotransmitters, and enhanced glucose tolerance and utilization in the three species, whereas physical transection or chemical blocking of the liver-brain nerve pathway abolished the effect of pFUS on glucose tolerance. Longitudinal multi-omic profiling of metabolic tissues from the treated animals confirmed pFUS-induced modifications of key metabolic functions in liver, pancreas, muscle, adipose, kidney and intestinal tissues. Non-invasive ultrasound activation of afferent autonomic nerves may represent a non-pharmacologic therapy for the restoration of glucose homoeostasis in type-2 diabetes and other metabolic diseases.

Glutamate-Serine-Glycine Index: A Novel Potential Biomarker in Pediatric Non-Alcoholic Fatty Liver Disease.

Preliminary evidence suggests that the glutamate-serine-glycine (GSG) index, which combines three amino acids involved in glutathione synthesis, may be used as a potential biomarker of non-alcoholic fatty liver disease (NAFLD). We investigated whether the GSG index is associated with NAFLD in youth, independent of other risk factors. Intrahepatic fat content (HFF%) and abdominal fat distribution were measured by magnetic resonance imaging (MRI) in a multiethnic cohort of obese adolescents, including Caucasians, African Americans, and Hispanics. NAFLD was defined as HFF% ≥ 5.5%. Plasma amino acids were measured by mass spectrometry. The GSG index was calculated as glutamate/(serine + glycine). The GSG index was higher in NAFLD patients (p = 0.03) and positively correlated with HFF% (r = 0.26, p = 0.02), alanine aminotransferase (r = 0.39, p = 0.0006), and aspartate aminotransferase (r = 0.26, p = 0.03). Adolescents with a high GSG index had a twofold higher prevalence of NAFLD than those with a low GSG index, despite similar adiposity, abdominal fat distribution, and liver insulin resistance. NAFLD prevalence remained significantly different between groups after adjustment for age, sex, race/ethnicity, and body mass index (OR 3.07, 95% confidence interval 1.09-8.61, p = 0.03). This study demonstrates the ability of the GSG index to detect NAFLD in at-risk pediatric populations with different genetically determined susceptibilities to intrahepatic fat accumulation, independent of traditional risk factors.

O-GlcNAcase targets pyruvate kinase M2 to regulate tumor growth.

Cancer cells are known to adopt aerobic glycolysis in order to fuel tumor growth, but the molecular basis of this metabolic shift remains largely undefined. O-GlcNAcase (OGA) is an enzyme harboring O-linked ß-N-acetylglucosamine (O-GlcNAc) hydrolase and cryptic lysine acetyltransferase activities. Here, we report that OGA is upregulated in a wide range of human cancers and drives aerobic glycolysis and tumor growth by inhibiting pyruvate kinase M2 (PKM2). PKM2 is dynamically O-GlcNAcylated in response to changes in glucose availability. Under high glucose conditions, PKM2 is a target of OGA-associated acetyltransferase activity, which facilitates O-GlcNAcylation of PKM2 by O-GlcNAc transferase (OGT). O-GlcNAcylation inhibits PKM2 catalytic activity and thereby promotes aerobic glycolysis and tumor growth. These studies define a causative role for OGA in tumor progression and reveal PKM2 O-GlcNAcylation as a metabolic rheostat that mediates exquisite control of aerobic glycolysis.

Mitochondrial MsrB2 serves as a switch and transducer for mitophagy.

Mitophagy can selectively remove damaged toxic mitochondria, protecting a cell from apoptosis. The molecular spatial-temporal mechanisms governing autophagosomal selection of reactive oxygen species (ROS)-damaged mitochondria, particularly in a platelet (no genomic DNA for transcriptional regulation), remain unclear. We now report that the mitochondrial matrix protein MsrB2 plays an important role in switching on mitophagy by reducing Parkin methionine oxidation (MetO), and transducing mitophagy through ubiquitination by Parkin and interacting with LC3. This biochemical signaling only occurs at damaged mitochondria where MsrB2 is released from the mitochondrial matrix. MsrB2 platelet-specific knockout and in vivo peptide inhibition of the MsrB2/LC3 interaction lead to reduced mitophagy and increased platelet apoptosis. Pathophysiological importance is highlighted in human subjects, where increased MsrB2 expression in diabetes mellitus leads to increased platelet mitophagy, and in platelets from Parkinson's disease patients, where reduced MsrB2 expression is associated with reduced mitophagy. Moreover, Parkin mutations at Met192 are associated with Parkinson's disease, highlighting the structural sensitivity at the Met192 position. Release of the enzyme MsrB2 from damaged mitochondria, initiating autophagosome formation, represents a novel regulatory mechanism for oxidative stress-induced mitophagy.

Oxidized Derivatives of Linoleic Acid in Pediatric Metabolic Syndrome: Is Their Pathogenic Role Modulated by the Genetic Background and the Gut Microbiota?

We tested whether oxidized linoleic acid metabolites (OXLAM) are associated with pediatric metabolic syndrome (MetS) and a proatherogenic lipoprotein profile in 122 obese adolescents. Furthermore, we examined whether genetic and metagenomic factors can modulate plasma OXLAM concentrations by genotyping the fatty acid desaturase 1/2 (FADS) gene and by characterizing the gut microbiota. Subjects with MetS (n = 50) showed higher concentrations of 9- and 13-oxo-octadecadienoic acid (9- and 13-oxo-ODE) than subjects without MetS (n = 72). Both metabolites were associated with an adverse lipoprotein profile that was characterized by elevated very small-dense low-density lipoprotein (p < 0.005) and large very low-density lipoprotein particles (p = 0.01). Plasma 9- and 13-oxo-ODE were higher in subjects carrying the haplotype AA of the FADS gene cluster (p = 0.030 and p = 0.048, respectively). Furthermore, the reduced gut bacterial load was associated with higher 9-oxo-ODE concentrations (p = 0.035). This is the first study showing that high plasma OXLAM concentrations are associated with MetS and suggesting that the leading factors for high plasma concentrations of OXLAM might be the genetic background and the composition of the gut microbiota. In conclusion, high concentrations of 9- and 13-oxo-ODE, which may be the result of a genetic predisposition and a reduced gut bacterial load, are associated with MetS and with a proatherogenic lipoprotein profile in obese adolescents.

Diabetes Exacerbates Myocardial Ischemia/Reperfusion Injury by Down-Regulation of MicroRNA and Up-Regulation of O-GlcNAcylation.

Management for patients with diabetes experiencing myocardial infarction remains a challenge. Here the authors show that hyperglycemia- and hyperinsulinemia-induced microRNA-24 (miR-24) reduction and O-GlcNAcylation in the diabetic heart contribute to poor survival and increased infarct size in diabetic myocardial ischemia/reperfusion (I/R). In a mouse model of myocardial I/R, pharmacological or genetic overexpression of miR-24 in hearts significantly reduced myocardial infarct size. Experimental validation revealed that miR-24 targets multiple key proteins, including O-GlcNac transferase, ATG4A, and BIM, to coordinately protect the myocardium from I/R injury. These results establish miR-24 as a promising therapeutic candidate for diabetic I/R injury.

Selective proton-observed, carbon-edited (selPOCE) MRS method for measurement of glutamate and glutamine 13 C-labeling in the human frontal cortex.

PURPOSE: 13 C magnetic resonance spectroscopy (MRS) in combination with infusion of 13 C-labeled substrates has led to unique insights into human brain metabolism and neurotransmitter cycling. However, the low sensitivity of direct 13 C MRS and high radiofrequency power requirements has limited 13 C MRS studies to predominantly data acquisition in large volumes of the occipital cortex. The purpose of this study is to develop an MRS technique for localized detection of 13 C-labeling of glutamate and glutamine in the human frontal lobe. METHODS: We used an indirect (1 H-[13 C]), proton-observed, carbon-edited MRS sequence (selPOCE) for detection of 13 C-labeled metabolites in relatively small volumes located in the frontal lobe at 4 T. The SelPOCE method allows for selective and separate detection of glutamate and glutamine resonances, which significantly overlap at magnetic field strengths used for clinical MRI. RESULTS: Phantom data illustrate how selPOCE can be tuned to selectively detect 13 C labeling in different metabolites. Three-dimensional specific absorption rate simulations of radiofrequency power deposition show that the selPOCE method operates comfortably within the global and local Food and Drug Administration specific absorption rate guidelines. In vivo selPOCE data are presented, which were acquired from a 45-mL volume in the frontal lobe of healthy subjects. The in vivo data show the time-dependent 13 C-labeling of glutamate and glutamine during intravenous infusion of [1-13 C]-glucose. Metrics describing spectral fitting quality of the glutamate and glutamine resonances are reported. CONCLUSIONS: The SelPOCE sequence allows the detection of 13 C-labeling in glutamate and glutamine from a relatively small volume in the human frontal lobe at low radiofrequency power requirements. Magn Reson Med 80:11-20, 2018. © 2017 International Society for Magnetic Resonance in Medicine.

Nanopatterned Bulk Metallic Glass Biosensors.

Nanopatterning as a surface area enhancement method has the potential to increase signal and sensitivity of biosensors. Platinum-based bulk metallic glass (Pt-BMG) is a biocompatible material with electrical properties conducive for biosensor electrode applications, which can be processed in air at comparably low temperatures to produce nonrandom topography at the nanoscale. Work presented here employs nanopatterned Pt-BMG electrodes functionalized with glucose oxidase enzyme to explore the impact of nonrandom and highly reproducible nanoscale surface area enhancement on glucose biosensor performance. Electrochemical measurements including cyclic voltammetry (CV) and amperometric voltammetry (AV) were completed to compare the performance of 200 nm Pt-BMG electrodes vs Flat Pt-BMG control electrodes. Glucose dosing response was studied in a range of 2 mM to 10 mM. Effective current density dynamic range for the 200 nm Pt-BMG was 10-12 times greater than that of the Flat BMG control. Nanopatterned electrode sensitivity was measured to be 3.28 µA/cm2/mM, which was also an order of magnitude greater than the flat electrode. These results suggest that nonrandom nanotopography is a scalable and customizable engineering tool which can be integrated with Pt-BMGs to produce biocompatible biosensors with enhanced signal and sensitivity.

Extracellular Mitochondrial DNA Is Generated by Fibroblasts and Predicts Death in Idiopathic Pulmonary Fibrosis.

RATIONALE: Idiopathic pulmonary fibrosis (IPF) involves the accumulation of α-smooth muscle actin-expressing myofibroblasts arising from interactions with soluble mediators such as transforming growth factor-ß1 (TGF-ß1) and mechanical influences such as local tissue stiffness. Whereas IPF fibroblasts are enriched for aerobic glycolysis and innate immune receptor activation, innate immune ligands related to mitochondrial injury, such as extracellular mitochondrial DNA (mtDNA), have not been identified in IPF. OBJECTIVES: We aimed to define an association between mtDNA and fibroblast responses in IPF. METHODS: We evaluated the response of normal human lung fibroblasts (NHLFs) to stimulation with mtDNA and determined whether the glycolytic reprogramming that occurs in response to TGF-ß1 stimulation and direct contact with stiff substrates, and spontaneously in IPF fibroblasts, is associated with excessive levels of mtDNA. We measured mtDNA concentrations in bronchoalveolar lavage (BAL) from subjects with and without IPF, as well as in plasma samples from two longitudinal IPF cohorts and demographically matched control subjects. MEASUREMENTS AND MAIN RESULTS: Exposure to mtDNA augments α-smooth muscle actin expression in NHLFs. The metabolic changes in NHLFs that are induced by interactions with TGF-ß1 or stiff hydrogels are accompanied by the accumulation of extracellular mtDNA. These findings replicate the spontaneous phenotype of IPF fibroblasts. mtDNA concentrations are increased in IPF BAL and plasma, and in the latter compartment, they display robust associations with disease progression and reduced event-free survival. CONCLUSIONS: These findings demonstrate a previously unrecognized and highly novel connection between metabolic reprogramming, mtDNA, fibroblast activation, and clinical outcomes that provides new insight into IPF.

A Branched-Chain Amino Acid-Related Metabolic Signature Characterizes Obese Adolescents with Non-Alcoholic Fatty Liver Disease.

Dysregulation of several metabolite pathways, including branched-chain amino acids (BCAAs), are associated with Non-Alcoholic Fatty Liver Disease (NAFLD) and insulin resistance in adults, while studies in youth reported conflicting results. We explored whether, independently of obesity and insulin resistance, obese adolescents with NAFLD display a metabolomic signature consistent with disturbances in amino acid and lipid metabolism. A total of 180 plasma metabolites were measured by a targeted metabolomic approach in 78 obese adolescents with (n = 30) or without (n = 48) NAFLD assessed by magnetic resonance imaging (MRI). All subjects underwent an oral glucose tolerance test and subsets of patients underwent a two-step hyperinsulinemic-euglycemic clamp and/or a second MRI after a 2.2 ± 0.8-year follow-up. Adolescents with NAFLD had higher plasma levels of valine (p = 0.02), isoleucine (p = 0.03), tryptophan (p = 0.02), and lysine (p = 0.02) after adjustment for confounding factors. Circulating BCAAs were negatively correlated with peripheral and hepatic insulin sensitivity. Furthermore, higher baseline valine levels predicted an increase in hepatic fat content (HFF) at follow-up (p = 0.01). These results indicate that a dysregulation of BCAA metabolism characterizes obese adolescents with NAFLD independently of obesity and insulin resistance and predict an increase in hepatic fat content over time.

Elevated α-Hydroxybutyrate and Branched-Chain Amino Acid Levels Predict Deterioration of Glycemic Control in Adolescents.

Context: Traditional risk factors for type 2 diabetes mellitus are weak predictors of changes in glucose tolerance and insulin sensitivity in youth. Objective: To identify early metabolic features of insulin resistance (IR) in youth and whether they predict deterioration of glycemic control. Design and Setting: A cross-sectional and longitudinal study was conducted at the Yale Pediatric Obesity Clinic. Patients and Intervention: Concentrations of α-hydroxybutyrate, ß-hydroxybutyrate, lactate, and branched-chain amino acids (BCAAs) were measured by nuclear magnetic resonance spectroscopy in 78 nondiabetic adolescents during an oral glucose tolerance test (OGTT). Associations between baseline metabolic alterations and longitudinal changes in glucose control were tested in 16 subjects after a mean follow-up of 2.3 years. Main Outcome Measures: The relationship between metabolite levels, parameters of IR, and glycemic control, and their progression over time. Results: Elevated fasting α-hydroxybutyrate levels were observed in adolescents with reduced insulin sensitivity after adjusting for age, sex, ethnicity, Tanner stage, and body mass index z-score (P = 0.014). Plasma α-hydroxybutyrate and BCAAs were increased throughout the course of the OGTT in this group (P < 0.03). Notably, borderline IR was associated with a progressive α-hydroxybutyrate decrease from elevated baseline concentrations to normal levels (P = 0.02). Increased baseline α-hydroxybutyrate concentrations were further associated with progressive worsening of glucose tolerance and disposition index. Conclusion: α-Hydroxybutyrate and BCAA concentrations during an OGTT characterize insulin-resistant youth and predict worsening of glycemic control. These findings provide potential biomarkers for risk assessment of type 2 diabetes and new insights into IR pathogenesis.

ß-Hydroxybutyrate Deactivates Neutrophil NLRP3 Inflammasome to Relieve Gout Flares.

Aging and lipotoxicity are two major risk factors for gout that are linked by the activation of the NLRP3 inflammasome. Neutrophil-mediated production of interleukin-1ß (IL-1ß) drives gouty flares that cause joint destruction, intense pain, and fever. However, metabolites that impact neutrophil inflammasome remain unknown. Here, we identified that ketogenic diet (KD) increases ß-hydroxybutyrate (BHB) and alleviates urate crystal-induced gout without impairing immune defense against bacterial infection. BHB inhibited NLRP3 inflammasome in S100A9 fibril-primed and urate crystal-activated macrophages, which serve to recruit inflammatory neutrophils in joints. Consistent with reduced gouty flares in rats fed a ketogenic diet, BHB blocked IL-1ß in neutrophils in a NLRP3-dependent manner in mice and humans irrespective of age. Mechanistically, BHB inhibited the NLRP3 inflammasome in neutrophils by reducing priming and assembly steps. Collectively, our studies show that BHB, a known alternate metabolic fuel, is also an anti-inflammatory molecule that may serve as a treatment for gout.

Pigment epithelium-derived factor restoration increases bone mass and improves bone plasticity in a model of osteogenesis imperfecta type VI via Wnt3a blockade.

Null mutations in for pigment epithelium-derived factor (PEDF), the protein product of the SERPINF1 gene, are the cause of osteogenesis imperfecta (OI) type VI. The PEDF-knockout (KO) mouse captures crucial elements of the human disease, including diminished bone mineralization and propensity to fracture. Our group and others have demonstrated that PEDF directs human mesenchymal stem cell (hMSC) commitment to the osteoblast lineage and modulates Wnt/ß-catenin signaling, a major regulator of bone development; however, the ability of PEDF to restore bone mass in a mouse model of OI type VI has not been determined. In this study, PEDF delivery increased trabecular bone volume/total volume by 52% in 6-mo-old PEDF-KO mice but not in wild-type mice. In young (19-d-old) PEDF-KO mice, PEDF restoration increased bone volume fraction by 35% and enhanced biomechanical parameters of bone plasticity. A Wnt-green fluorescent protein reporter demonstrated dynamic changes in Wnt/ß-catenin signaling characterized by early activation and marked suppression during terminal differentiation of hMSCs. Continuous Wnt3a exposure impeded mineralization of hMSCs, whereas the combination of Wnt3a and PEDF potentiated mineralization. Interrogation of the PEDF sequence identified a conserved motif found in other Wnt modulators, such as the dickkopf proteins. Mutation of a single amino acid on a 34-mer PEDF peptide increased mineralization of hMSC cultures compared with the native peptide sequence. These results indicate that PEDF counters Wnt signaling to allow for osteoblast differentiation and provides a mechanistic insight into how the PEDF null state results in OI type VI.-Belinsky, G. S., Sreekumar, B., Andrejecsk, J. W., Saltzman, W. M., Gong, J., Herzog, R. I., Lin, S., Horsley, V., Carpenter, T. O., Chung, C. Pigment epithelium-derived factor restoration increases bone mass and improves bone plasticity in a model of osteogenesis imperfecta type VI via Wnt3a blockade.

Quantification of 1H NMR Spectra from Human Plasma.

Human plasma is a biofluid that is high in information content, making it an excellent candidate for metabolomic studies. 1H NMR has been a popular technique to detect several dozen metabolites in blood plasma. In order for 1H NMR to become an automated, high-throughput method, challenges related to (1) the large signal from lipoproteins and (2) spectral overlap between different metabolites have to be addressed. Here diffusion-weighted 1H NMR is used to separate lipoprotein and metabolite signals based on their large difference in translational diffusion. The metabolite 1H NMR spectrum is then quantified through spectral fitting utilizing full prior knowledge on the metabolite spectral signatures. Extension of the scan time by 3 minutes or 15% per sample allowed the acquisition of a 1H NMR spectrum with high diffusion weighting. The metabolite 1H NMR spectra could reliably be modeled with 28 metabolites. Excellent correlation was found between results obtained with diffusion NMR and ultrafiltration. The combination of minimal sample preparation together with minimal user interaction during processing and quantification provides a metabolomics technique for automated, quantitative 1H NMR of human plasma.

Hyperglycemia repression of miR-24 coordinately upregulates endothelial cell expression and secretion of von Willebrand factor.

An elevated level of von Willebrand factor (VWF) in diabetic patients is associated with increased risk of thrombotic cardiovascular events. The underlying mechanism of how VWF expression is upregulated in diabetes mellitus is poorly understood. We now report that hyperglycemia-induced repression of microRNA-24 (miR-24) increases VWF expression and secretion in diabetes mellitus. In diabetic patients and diabetic mouse models (streptozotocin/high-fat diet-induced and db/db mice), miR-24 is reduced in both tissues and plasma. Knockdown of miR-24 in mice leads to increased VWF mRNA and protein levels and enhanced platelet tethering (spontaneous thrombosis). miR-24 tightly controls VWF levels through pleiotropic effects, including direct binding to the 3' untranslated region of VWF and targeting FURIN and the histamine H1 receptor, known regulators of VWF processing and secretion in endothelial cells. We present a novel mechanism for miR-24 downregulation through hyperglycemia-induced activation of aldose reductase, reactive oxygen species, and c-Myc. These findings support a critical role for hyperglycemic repression of miR-24 in VWF-induced pathology. miR-24 represents a novel therapeutic target to prevent adverse thrombotic events in patients with diabetes mellitus.

Vaccination with single chain antigen receptors for islet-derived peptides presented on I-A(g7) delays diabetes in NOD mice by inducing anergy in self-reactiveT-cells.

To develop a vaccination approach for prevention of type 1 diabetes (T1D) that selectively attenuates self-reactive T-cells targeting specific autoantigens, we selected phage-displayed single chain antigen receptor libraries for clones binding to a complex of the NOD classII MHC I-A(g7) and epitopes derived from the islet autoantigen RegII. Libraries were generated from B-cell receptor repertoires of classII-mismatched mice immunized with RegII-pulsed NOD antigen presenting cells or from T-cell receptor repertoires in pancreatic lymph nodes of NOD mice. Both approaches yielded clones recognizing a RegII-derived epitope in the context of I-A(g7), which activated autoreactive CD4(+) T-cells. A receptor with different specificity was obtained by converting the BDC2.5 TCR into single chain form. B- but not T-cells from donors vaccinated with the clones transferred protection from diabetes to NOD-SCID recipients if the specificity of the diabetes inducer cell and the single chain receptor were matched. B-cells and antibodies from donors vaccinated with the BDC2.5 single chain receptor induced a state of profound anergy in T-cells of BDC2.5 TCR transgenic NOD recipients while B-cells from donors vaccinated with a single chain receptor specific for I-A(g7) RegII peptide complexes induced only partial non-responsiveness. Vaccination of normal NOD mice with receptors recognizing I-A(g7) RegII peptide complexes or with the BDC2.5 single chain receptor delayed onset of T1D. Thus anti-idiotypic vaccination can be successfully applied to T1D with vaccines either generated from self-reactive T-cell clones or derived from antigen receptor libraries.

Lactate preserves neuronal metabolism and function following antecedent recurrent hypoglycemia.

Hypoglycemia occurs frequently during intensive insulin therapy in patients with both type 1 and type 2 diabetes and remains the single most important obstacle in achieving tight glycemic control. Using a rodent model of hypoglycemia, we demonstrated that exposure to antecedent recurrent hypoglycemia leads to adaptations of brain metabolism so that modest increments in circulating lactate allow the brain to function normally under acute hypoglycemic conditions. We characterized 3 major factors underlying this effect. First, we measured enhanced transport of lactate both into as well as out of the brain that resulted in only a small increase of its contribution to total brain oxidative capacity, suggesting that it was not the major fuel. Second, we observed a doubling of the glucose contribution to brain metabolism under hypoglycemic conditions that restored metabolic activity to levels otherwise only observed at euglycemia. Third, we determined that elevated lactate is critical for maintaining glucose metabolism under hypoglycemia, which preserves neuronal function. These unexpected findings suggest that while lactate uptake was enhanced, it is insufficient to support metabolism as an alternate substrate to replace glucose. Lactate is, however, able to modulate metabolic and neuronal activity, serving as a "metabolic regulator" instead.

Diabetes. Can tight glycemic control in diabetes benefit cognition?

Intensive glucose management, if begun early, diminishes the long-term complications of diabetes. Whether the cognitive domain also benefits from such therapy is not clear, and has remained subject to investigation. Do the new results from the ACCORD-MIND study settle the issue?