Muscle Mitochondrial ATP Synthesis and Glucose Transport/Phosphorylation in Type 2 Diabetes

BACKGROUND: Muscular insulin resistance is frequently characterized by blunted increases in glucose-6-phosphate (G-6-P) reflecting impaired glucose transport/phosphorylation. These abnormalities likely relate to excessive intramyocellular lipids and mitochondrial dysfunction. We hypothesized that alterations in insulin action and mitochondrial function should be present even in nonobese patients with well-controlled type 2 diabetes mellitus (T2DM). METHODS AND FINDINGS: We measured G-6-P, ATP synthetic flux (i.e., synthesis) and lipid contents of skeletal muscle with (31)P/(1)H magnetic resonance spectroscopy in ten patients with T2DM and in two control groups: ten sex-, age-, and body mass-matched elderly people; and 11 younger healthy individuals. Although insulin sensitivity was lower in patients with T2DM, muscle lipid contents were comparable and hyperinsulinemia increased G-6-P by 50% (95% confidence interval [CI] 39%–99%) in all groups. Patients with diabetes had 27% lower fasting ATP synthetic flux compared to younger controls (p = 0.031). Insulin stimulation increased ATP synthetic flux only in controls (younger: 26%, 95% CI 13%–42%; older: 11%, 95% CI 2%–25%), but failed to increase even during hyperglycemic hyperinsulinemia in patients with T2DM. Fasting free fatty acids and waist-to-hip ratios explained 44% of basal ATP synthetic flux. Insulin sensitivity explained 30% of insulin-stimulated ATP synthetic flux. CONCLUSIONS: Patients with well-controlled T2DM feature slightly lower flux through muscle ATP synthesis, which occurs independently of glucose transport /phosphorylation and lipid deposition but is determined by lipid availability and insulin sensitivity. Furthermore, the reduction in insulin-stimulated glucose disposal despite normal glucose transport/phosphorylation suggests further abnormalities mainly in glycogen synthesis in these patients

Plasma Metabolomic Profiles Reflective of Glucose Homeostasis in Non-Diabetic and Type 2 Diabetic Obese African-American Women

Insulin resistance progressing to type 2 diabetes mellitus (T2DM) is marked by a broad perturbation of macronutrient intermediary metabolism. Understanding the biochemical networks that underlie metabolic homeostasis and how they associate with insulin action will help unravel diabetes etiology and should foster discovery of new biomarkers of disease risk and severity. We examined differences in plasma concentrations of >350 metabolites in fasted obese T2DM vs. obese non-diabetic African-American women, and utilized principal components analysis to identify 158 metabolite components that strongly correlated with fasting HbA1c over a broad range of the latter (r = −0.631; p<0.0001). In addition to many unidentified small molecules, specific metabolites that were increased significantly in T2DM subjects included certain amino acids and their derivatives (i.e., leucine, 2-ketoisocaproate, valine, cystine, histidine), 2-hydroxybutanoate, long-chain fatty acids, and carbohydrate derivatives. Leucine and valine concentrations rose with increasing HbA1c, and significantly correlated with plasma acetylcarnitine concentrations. It is hypothesized that this reflects a close link between abnormalities in glucose homeostasis, amino acid catabolism, and efficiency of fuel combustion in the tricarboxylic acid (TCA) cycle. It is speculated that a mechanism for potential TCA cycle inefficiency concurrent with insulin resistance is “anaplerotic stress” emanating from reduced amino acid-derived carbon flux to TCA cycle intermediates, which if coupled to perturbation in cataplerosis would lead to net reduction in TCA cycle capacity relative to fuel delivery

Genotyping of adiponectin receptor polymorphisms and association with the presence of Coronary Artery Disease and insulin resistance

Adiponectin is a hormone secreted by adipose tissue. To this date 2 adiponectin receptors are been discovered, named ADIPOR1 and ADIPOR2. AIM The aim of this thesis is the investigation of a possible association between eight single nucleotide polymorphisms (SNPs) of ADIPOR2 and one single nucleotide polymorphism of ADIPOR1 with coronary artery disease and insulin-resistance. METHODS We studied 70 individuals with age 45-70 years. We divided our sample in two groups. Group A had individuals with diagnosed coronary artery disease while in group B with no coronary artery disease. Also we made subgroups according to the status of insulin tolerance. All subjects underwent coronary angiography and exercise treadmill test. Carotid Intima-Media Thickness (IMT) as index of aterosclerosis and Flow Mediated Dilatation (FMD) of the brachial artery as index of endothelial disfunction were calculated. All subjects underwent oral glucose tolerance test. HOMA and Matsuda index were calculated. The genetic analysis of the polymorphisms was performed using two techniques: real time PCR with hybridization probes and Allele Specific Primer Extension (ASPE) with microspheres. For the statistical analysis of the data we used SPSS for Windows. RESULTS-DISCUSSION There was a statistically significant difference in the distribution of alleles of rs767870 G/A from ADIPOR2 gene between group A and group B (p=0.013). AG heterozygotes of the same polymorphism had significantly lower FMD compared with AA homozygotes (p=0.046). Polymorphism rs767870 G/A of ADIPOR2 can be associated with coronary artery disease and insulin resistance as we saw in the different allelic distribution and the association of low FMD with heterozygotes. It is possible to use this polymorphism as a genetic marker for early diagnosis of cardiovascular disease.Η λιπονεκτίνη ή αλλιώς αντιπονεκτίνη είναι μια ορμόνη που εκκρίνεται απο τον λιπώδη ιστό. Μέχρι σήμερα έχουν εντοπιστεί δύο υποδοχείς για την λιπονεκτίνη. Τα δύο γονίδια που κωδικοποιούν την έκφραση των υποδοχέων ονομάστηκαν ADIPOR1 και ADIPOR2. ΣΤΟΧΟΣ Στόχος της παρούσας διατριβής είναι η διερεύνηση συσχέτισης οκτώ πολυμορφισμών του γονιδίου που κωδικοποιεί την έκφραση του υποδοχέα της λιπονεκτίνης ADIPOR2 και ενός πολυμορφισμού του γονιδίου που κωδικοποιεί την έκφραση του υποδοχέα της λιπονεκτίνης ADIPOR1 με την παρουσία Στεφανιαίας Νόσου και ινσουλινοαντοχής. ΜΕΘΟΔΟΛΟΓΙΑ Η μελέτη έγινε σε 70 εθελοντές, άνδρες και γυναίκες ηλικίας 45-70 ετών. Η πρώτη ομάδα (ομάδα Α) περιλάμβανε 42 ασθενείς με βεβαιωμένη Στεφανιαία Νόσο, ενώ η δεύτερη ομάδα (ομάδα Β) περιλάμβανε 28 άτομα χωρίς Στεφανιαία Νόσο. Xωρίσαμε τα άτομα των δύο ομάδων σε υποομάδες ανάλογα με την κατάσταση ανοχής τους στη γλυκόζη. Οι εθελοντές υποβλήθηκαν σε στεφανιογραφία, σε δοκιμασία κόπωσης και σε από του στόματος δοκιμασία ανοχής στη γλυκόζη. Ως δείκτης αθηρωμάτωσης υπολογίστηκε το ΙΜΤ και ως δείκτης ενδοθηλιακής δυσλειτουργίας το FMD. Υπολογίστηκαν ο δείκτης ινσουλινοευαισθησίας Matsuda καθώς και ο δείκτης ινσουλινοαντίστασης HOMA. Για τη γονοτυπική ανάλυση των πολυμορφισμών χρησιμοποιήσαμε δύο τεχνικές, την τεχνική της αντίδρασης πολυμεράσης (PCR) με χρήση φθοριζόντων ιχνηλατών και την τεχνική πολλαπλής ανίχνευσης αλληλίων με επέκταση εκκινητών (ASPE) ειδικών για τα αλλήλια. Η ανάλυση των δεδομένων έγινε με τη χρήση του στατιστικού πακέτου SPSS for Windows. ΑΠΟΤΕΛΕΣΜΑΤΑ-ΣΥΜΠΕΡΑΣΜΑΤΑ Παρατηρήσαμε ότι ο πολυμορφισμός rs767870 G/A του γονιδίου ADIPOR2 παρουσίασε στατιστικά σημαντική διαφορά στη κατανομή των αλληλίων μεταξύ των δύο ομάδων (p=0.013). Επιπλέον, για όλα τα άτομα της μελέτης, οι ετεροζυγώτες ΑG έχουν στατιστικά σημαντικά μικρότερη τιμή του FMD απο τους ομοζυγώτες ΑΑ (p=0.046). Επειδή κατά τα ευρήματα της μελέτης ο πολυμορφισμός rs767870 G/A συνδέεται με την παρουσία Στεφανιαίας Νόσου, πιθανό να μπορεί να χρησιμοποιηθεί ως γενετικός δείκτης για έγκαιρη διάγνωση ατόμων με προδιάθεση να αναπτύξουν στεφανιαία νόσ

A comparison of different dna extraction methods and molecular techniques for the detection and identification of foodborne pathogens

Abatract: Foodborne infections continue to plague Europe. Food safety monitoring is in crisis as the existing techniques for detecting pathogens do not keep up with the global rising of food production and consumption. Thus, the development of innovative techniques for detecting and identifying pathogenic bacteria has become critical. The aim of the present study was firstly to develop an innovative simple and low cost method of extracting bacterial DNA from contaminated food and water samples with Salmonella enteric(a) subsp. enteric(a) serovar Typhimurium and Listeria monocytogenes and its comparison with two commercial DNA extraction kits (Qiagen, Macherey-Nagel). Finally, pathogens’ detection using two molecular techniques (PCR-electrophoresis, LAMP), in order to evaluate the best combination of DNA extraction and identification based on their sensitivity, cost, rapidity and simplicity. Considering the above criteria, among them, best was proved an in-house bacterial DNA extraction method, based on the chloroform-isoamyl alcohol protocol, with certain modifications. This technique showed statistically similar results in terms of sensitivity, compared to the commercial kits, while at the same time maintained high rapidity and much lower cost. Lastly, between the molecular techniques, LAMP was found more promising considering its simplicity, high rapidity and sensitivity. Conclusively, the in-house DNA extraction method along with the LAMP technique, was proven to be the best among the presented combinations. © 2021 the Author(s), licensee AIMS Press

Characterization of <i>Lacticaseibacillus rhamnosus</i>, <i>Levilactobacillus brevis</i> and <i>Lactiplantibacillus plantarum</i> Metabolites and Evaluation of Their Antimicrobial Activity against Food Pathogens

Lactic acid bacteria (LAB) play an important role as natural food preservatives. However, the characterization of the variety of their metabolites is limited. The objective of this study was to determine the production of specific metabolites of Lacticaseibacillus rhamnosus, Levilactobacillus brevis and Lactiplantibacillus plantarum by an optimized liquid chromatography with an ultraviolet/diode detection (HPLC-UV/DAD) method and to investigate their potential antimicrobial activity against specific food pathogens. Based on the results of this study, the main metabolites detected in Levilactobacillus brevis were 103.4 μg mL−1 DL-p-Hydroxyphenyllactic acid (OH-PLA) and 2.59 μg mL−1 vanillic acid, while 216.2 μg mL−1 OH-PLA, 19.0 μg mL−1 salicylic acid, 3.7 μg mL−1 vanillic acid, 6.9 μg mL−1 ferulic acid, 4.2 μg mL−1 benzoic acid and 1.4 μg mL−1 4-Hydrocinnamic acid were identified in the Lactiplantibacillus plantarum strain and 147.6 μg mL−1 OH-PLA and 4.9 μg mL−1 ferulic acid were identified in Lacticaseibacillus rhamnosus. This study provides alternative approaches for the molecules involved in the antimicrobial activity of food microorganism fermentation. These molecules may be used as antimicrobial ingredients in the food industry instead of conventional chemical preservatives