SIRT1-Dependent Upregulation of Antiglycative Defense in HUVECs Is Essential for Resveratrol Protection against High Glucose Stress

Uncontrolled accumulation of methylglyoxal (MG) and reactive oxygen species (ROS) occurs in hyperglycemia-induced endothelial dysfunction associated with diabetes. Resveratrol (RSV) protects the endothelium upon high glucose (HG); however, the mechanisms underlying such protective effects are still debated. Here we identified key molecular players involved in the glycative/oxidative perturbations occurring in endothelial cells exposed to HG. In addition, we determined whether RSV essentially required SIRT1 to trigger adaptive responses in HG-challenged endothelial cells. We used primary human umbilical vein endothelial cells (HUVECs) undergoing a 24-h treatment with HG, with or without RSV and EX527 (i.e., SIRT1 inhibitor). We found that HG-induced glycative stress (GS) and oxidative stress (OS), by reducing SIRT1 activity, as well as by diminishing the efficiency of MG- and ROS-targeting protection. RSV totally abolished the HG-dependent cytotoxicity, and this was associated with SIRT1 upregulation, together with increased expression of GLO1, improved ROS-scavenging efficiency, and total suppression of HG-related GS and OS. Interestingly, RSV failed to induce effective response to HG cytotoxicity when EX527 was present, thus suggesting that the upregulation of SIRT1 is essential for RSV to activate the major antiglycative and antioxidative defense and avoid MG- and ROS-dependent molecular damages in HG environment

SIRT1-dependent upregulation of antiglycative defense in HUVECs is essential for resveratrol protection against high glucose stress

Uncontrolled accumulation of methylglyoxal (MG) and reactive oxygen species (ROS) occurs in hyperglycemia-induced endothelial dysfunction associated with diabetes. Resveratrol (RSV) protects the endothelium upon high glucose (HG); however, the mechanisms underlying such protective effects are still debated. Here we identified key molecular players involved in the glycative/oxidative perturbations occurring in endothelial cells exposed to HG. In addition, we determined whether RSV essentially required SIRT1 to trigger adaptive responses in HG-challenged endothelial cells. We used primary human umbilical vein endothelial cells (HUVECs) undergoing a 24-h treatment with HG, with or without RSV and EX527 (i.e., SIRT1 inhibitor). We found that HG-induced glycative stress (GS) and oxidative stress (OS), by reducing SIRT1 activity, as well as by diminishing the efficiency of MG- and ROS-targeting protection. RSV totally abolished the HG-dependent cytotoxicity, and this was associated with SIRT1 upregulation, together with increased expression of GLO1, improved ROS-scavenging efficiency, and total suppression of HG-related GS and OS. Interestingly, RSV failed to induce effective response to HG cytotoxicity when EX527 was present, thus suggesting that the upregulation of SIRT1 is essential for RSV to activate the major antiglycative and antioxidative defense and avoid MG- and ROS-dependent molecular damages in HG environment

Artificial Intelligence and liver: Opportunities and barriers

: Artificial Intelligence (AI) has recently been shown as an excellent tool for the study of the liver; however, many obstacles still have to be overcome for the digitalization of real-world hepatology. The authors present an overview of the current state of the art on the use of innovative technologies in different areas (big data, translational hepatology, imaging, and transplant setting). In clinical practice, physicians must integrate a vast array of data modalities (medical history, clinical data, laboratory tests, imaging, and pathology slides) to achieve a diagnostic or therapeutic decision. Unfortunately, machine learning and deep learning are still far from really supporting clinicians in real life. In fact, the accuracy of any technological support has no value in medicine without the support of clinicians. To make better use of new technologies, it is essential to improve clinicians' knowledge about them. To this end, the authors propose that collaborative networks for multidisciplinary approaches will improve the rapid implementation of AI systems for developing disease-customized AI-powered clinical decision support tools. The authors also discuss ethical, educational, and legal challenges that must be overcome to build robust bridges and deploy potentially effective AI in real-world clinical settings

This folder contains the source data for Figs 2A, 2C, 4B, 5A, 5B, 6A–6D, 7D, 7E, S2A, S5B,

This folder contains the source data for Figs 2A, 2C, 4B, 5A, 5B, 6A–6D, 7D, 7E, S2A, S5B, </p

Expression of the bicistronic plasmid Sirius-M₃i3(558n)-EGFP.

(A) Schematic representation of the bicistronic plasmid bearing the i3 loop of the rat muscarinic M3 receptor (558 nucleotides from nucleotide 880 to nucleotide 1437) between the coding regions of the Sirius and EGFP fluorescent proteins. (B) EGFP expression in COS-7 cells transfected with the bicistronic plasmid Sirius-M3i3(558n)-EGFP. Source data for panel B can be found in S1 Data. (AI)</p

This files contains supplementary discussion related to some of the results.

This files contains supplementary discussion related to some of the results.</p

M₂tail(368–466) predominantly localizes to the cell mitochondria.

(A) Colocalization of M2tail(368–466)-EGFP (green) together with the mitochondrial targeted dye Mitotracker (magenta) in HEK293 cells. Separate spectral channels are displayed below. (B) Co-expression of M2tail(368–466)-EGFP (green) and wild-type M2-mRuby2 (red) in HEK293 cells. Separate spectral channels are displayed below. Left: confocal micrograph of the basal membrane of 2 cells. Right: confocal cross section of a separate cell. (C) Western blot (10% TRIS-Glycine PAA gel) of mitochondria purified from HEK293 cells transfected with the M2-myc, M2tail(368–466)-myc, M2M368A-myc, together with an untransfected control are blotted using an anti-myc antibody. Loading control was verified by immunoblotting COXIV and β-actin. (D) Cellular localization of the construct M2-mRuby2-STOP-M2-i3-tail-EGFP in HEK293 cells. Expression of the M2tail(368–466)-EGFP (green) after the STOP codon is an indicator of the endogenous IRES activity. While M2-mRuby2 localizes correctly and predominantly to the cell plasma membrane, M2tail(368–466)-EGFP localizes intracellularly. Separate spectral channels are displayed below. (E) Same conditions as in D, for a separate cell where Mitotracker (magenta) was added. (F) M2-fr.sh.-EGFP (green) expressed in conjunction with Mitotracker (magenta). The EGFP is expressed despite the creation of an in-frame stop after M368, confirming endogenous cap-independent translation of the M2tail(368–466). Arrows indicate areas of colocalization with the mitochondria. Manders Colocalization Coefficient MCC2 is 0.3 ± 0.1 (n = 6). Separate spectral channels are displayed below. Scale bars are 10 μm throughout. IRES, internal ribosome entry site.</p