Brigitte I. Frohnert

The Fatty Acid Transport Protein (FATP1) Is a Very Long Chain Acyl-CoA Synthetase

The primary sequence of the murine fatty acid transport protein (FATP1) is very similar to the multigene family of very long chain (C20-C26) acyl-CoA synthetases. To determine if FATP1 is a long chain acyl coenzyme A synthetase, FATP1-Myc/His fusion protein was expressed in COS1 cells, and its enzymatic activity was analyzed. In addition, mutations were generated in two domains conserved in acyl-CoA synthetases: a 6- amino acid substitution into the putative active site (amino acids 249–254) generating mutant M1 and a 59-amino acid deletion into a conserved C-terminal domain (amino acids 464–523) generating mutant M2. Immunolocalization revealed that the FATP1-Myc/His forms were distributed between the COS1 cell plasma membrane and intracellular membranes. COS1 cells expressing wild type FATP1-Myc/His exhibited a 3-fold increase in the ratio of lignoceroyl-CoA synthetase activity (C24:0) to palmitoyl-CoA synthetase activity (C16:0), characteristic of very long chain acyl-CoA synthetases, whereas both mutant M1 and M2 were catalytically inactive. Detergent-solubilized FATP1-Myc/His was partially purified using nickel-based affinity chromatography and demonstrated a 10-fold increase in very long chain acyl-CoA specific activity (C24:0/C16:0). These results indicate that FATP1 is a very long chain acyl-CoA synthetase and suggest that a potential mechanism for facilitating mammalian fatty acid uptake is via esterification coupled influx.

Increased adipose protein carbonylation in human obesity

Insulin resistance is associated with obesity but mechanisms controlling this relationship in humans are not fully understood. Studies in animal models suggest a linkage between adipose reactive oxygen species (ROS) and insulin resistance. ROS oxidize cellular lipids to produce a variety of lipid hydroperoxides that in turn generate reactive lipid aldehydes that covalently modify cellular proteins in a process termed carbonylation. Mammalian cells defend against reactive lipid aldehydes and protein carbonylation by glutathionylation using glutathione‐S‐transferase A4 (GSTA4) or carbonyl reduction/oxidation via reductases and/or dehydrogenases. Insulin resistance in mice is linked to ROS production and increased level of protein carbonylation, mitochondrial dysfunction, decreased insulin‐stimulated glucose transport, and altered adipokine secretion. To assess protein carbonylation and insulin resistance in humans, eight healthy participants underwent subcutaneous fat biopsy from the periumbilical region for protein analysis and frequently sampled intravenous glucose tolerance testing to measure insulin sensitivity. Soluble proteins from adipose tissue were analyzed using two‐dimensional gel electrophoresis and the major carbonylated proteins identified as the adipocyte and epithelial fatty acid‐binding proteins. The level of protein carbonylation was directly correlated with adiposity and serum free fatty acids (FFAs). These results suggest that in human obesity oxidative stress is linked to protein carbonylation and such events may contribute to the development of insulin resistance.

Oral glucose tolerance testing in children with cystic fibrosis

Ode KL, Frohnert B, Laguna T, Phillips J, Holme B, Regelmann W, Thomas W, Moran A. Oral glucose tolerance testing in children with cystic fibrosis.

Identification and treatment of metabolic complications in pediatric obesity

Metabolic consequences of obesity including insulin resistance, type 2 diabetes mellitus, hyperlipidemia, hypertension, polycystic ovarian syndrome, and non-alcoholic fatty liver infiltration are rapidly emerging in the pediatric population. Identifying effective strategies for identifying and treating these obesity related co-morbidities in children are crucial to the prevention of future cardiovascular disease and poor health outcomes. This review discusses the pathophysiologic connections between obesity, metabolic disease and cardiovascular risk. Current evidence and recommendations for screening and treatment for the metabolic consequences of pediatric obesity are reviewed.

Protein Carbonylation, Mitochondrial Dysfunction, and Insulin Resistance

Oxidative stress has been identified as a common mechanism for cellular damage and dysfunction in a wide variety of disease states. Current understanding of the metabolic changes associated with obesity and the development of insulin resistance has focused on the role of oxidative stress and its interaction with inflammatory processes at both the tissue and organismal level. Obesity-related oxidative stress is an important contributing factor in the development of insulin resistance in the adipocyte as well as the myocyte. Moreover, oxidative stress has been linked to mitochondrial dysfunction, and this is thought to play a role in the metabolic defects associated with oxidative stress. Of the various effects of oxidative stress, protein carbonylation has been identified as a potential mechanism underlying mitochondrial dysfunction. As such, this review focuses on the relationship between protein carbonylation and mitochondrial biology and addresses those features that point to either the causal or casual relationship of lipid peroxidation-induced protein carbonylation as a determining factor in mitochondrial dysfunction.

Relation Between Serum Free Fatty Acids and Adiposity, Insulin Resistance, and Cardiovascular Risk Factors From Adolescence to Adulthood

The objective of this study was to describe longitudinal relations of serum total free fatty acids (FFAs) to insulin resistance (IR) and cardiovascular (CV) risk factors from adolescence into adulthood. The cohort included participants in a longitudinal study of obesity and IR with complete data, including anthropometric measures, FFAs, IR measured by euglycemic clamp, blood pressure, fasting serum lipids, and insulin at mean 15 and 22 years of age (n = 207) and their parents (n = 272). FFAs and IR were not significantly related at mean 15 years of age but were significantly related at mean age 22 years. FFA did not relate to BMI at either age. FFA at 15 years of age estimated IR at 22 years of age. In parents (mean age 51 years), FFA was significantly correlated with BMI, percent body fat, systolic blood pressure, LDL, and IR. Associations with all risk factors except IR in parents were attenuated by adjustment for BMI. Most 22 years of age correlations with parents were higher than corresponding 15 years of age correlations. This study finds that FFA is associated with IR starting in young adulthood. The relation between FFA and CV risk factors does not become significant until later adulthood. The results support a significant impact of early metabolic dysfunction on later CV risk.

Glutathionylated Lipid Aldehydes are Products of Adipocyte Oxidative Stress and Activators of Macrophage Inflammation

Obesity-induced insulin resistance has been linked to adipose tissue lipid aldehyde production and protein carbonylation. Trans-4-hydroxy-2-nonenal (4-HNE) is the most abundant lipid aldehyde in murine adipose tissue and is metabolized by glutathione S-transferase A4 (GSTA4), producing glutathionyl-HNE (GS-HNE) and its metabolite glutathionyl-1,4-dihydroxynonene (GS-DHN). The objective of this study was to evaluate adipocyte production of GS-HNE and GS-DHN and their effect on macrophage inflammation. Compared with lean controls, GS-HNE and GS-DHN were more abundant in visceral adipose tissue of ob/ob mice and diet-induced obese, insulin-resistant mice. High glucose and oxidative stress induced production of GS-HNE and GS-DHN by 3T3-L1 adipocytes in a GSTA4-dependent manner, and both glutathionylated metabolites induced secretion of tumor necrosis factor-α from RAW 264.7 and primary peritoneal macrophages. Targeted microarray analysis revealed GS-HNE and GS-DHN induced expression of inflammatory genes, including C3, C4b, c-Fos, igtb2, Nfkb1, and Nos2. Transgenic overexpression of GSTA4 in mouse adipose tissue led to increased production of GS-HNE associated with higher fasting glucose levels and moderately impaired glucose tolerance. These results indicated adipocyte oxidative stress results in GSTA4-dependent production of proinflammatory glutathione metabolites, GS-HNE and GS-DHN, which may represent a novel mechanism by which adipocyte dysfunction results in tissue inflammation and insulin resistance.

Impaired fasting glucose in cystic fibrosis.

OBJECTIVE While glucose tolerance abnormalities are common in cystic fibrosis (CF), impaired fasting glucose (IFG) has scarcely been explored. No studies have examined the relation between IFG and clinical status. RESEARCH DESIGN AND METHODS Data were retrieved from the University of Minnesota CF database on oral glucose tolerance tests (OGTTs) performed in 1996–2005. Subjects were identified as normal glucose tolerance (NGT), impaired glucose tolerance (IGT), or CF–related diabetes without fasting hyperglycemia (CFRD FH−). Patients with fasting hyperglycemia were excluded. The presence of IFG was assessed within each category. In a separate case-control cohort study, subjects with IFG were matched to CF control subjects by age, sex, and OGTT class to explore outcomes. RESULTS For the total population (n = 310), the prevalence of IFG was 22%, and by OGTT class was NGT 14%, IGT 31%, CFRD FH− 53%. Within the cohort study, mortality was significantly reduced in IFG (two vs. nine deaths, odds ratio [OR] = 0.2 [95% CI 0.04–0.9]). IFG did not confer increased risk of progression to diabetes (OR 0.66 [0.29–1.48]). Lung function was better in pediatric IFG subjects with IGT and not significantly worse in adults with IGT or adults and children with NGT and CFRD FH−. BMI was not significantly different in IFG subjects versus control subjects. CONCLUSIONS Contrary to expectations in patients with CF, IFG appeared to be associated with improved survival and was not associated with worse nutritional or pulmonary status or increased progression to fasting hyperglycemia.

Blunted response to a growth hormone stimulation test is associated with unfavorable cardiovascular risk factor profile in childhood cancer survivors

Childhood cancer survivors (CCS) are at risk for growth hormone (GH) deficiency. CCS are also at increased risk for early mortality from cardiovascular (CV) disease, but the association between GH levels and CV risk remains poorly understood. The goal of this study was to examine the cross‐sectional association between stimulated GH levels and CV risk factors in CCS younger than 18 years.

No relation between cystic fibrosis-related diabetes and type 1 diabetes autoimmunity

Diabetes is the most common comorbidity in individuals with cystic fibrosis. The etiology is poorly understood. Data on the presence of diabetes autoantibodies are conflicting, and little is known about type 1 diabetes gene associations. Our goal was to determine the prevalence of antibodies and HLA haplotypes known to be associated with type 1 diabetes in cystic fibrosis–related diabetes (CFRD). Patients with CFRD with fasting hyperglycemia were recruited from the University of Minnesota. Serum for antibodies and buffy coats for HLA were sent for analysis to the Barbara Davis Center for Childhood Diabetes (BDC). All patients gave informed consent. The Eisenbarth laboratory at BDC serves as the autoantibody/HLA reference laboratory for large national diabetes studies. Insulin, insulinoma-associated protein 2 (IA-2), GAD65, and zinc transporter 8 (ZnT8) autoantibodies were measured by radioimmunoassay …