Structure-functional changes in eNAMPT at high concentrations mediate mouse and human beta cell dysfunction in type 2 diabetes

Abstract

Aims/hypothesis Progressive decline in functional beta cell mass is central to the development of type 2 diabetes. Elevated serum levels of extracellular nicotinamide phosphoribosyltransferase (eNAMPT) are associated with beta cell failure in type 2 diabetes and eNAMPT immuno-neutralisation improves glucose tolerance in mouse models of diabetes. Despite this, the effects of eNAMPT on functional beta cell mass are poorly elucidated, with some studies having separately reported beta cell-protective effects of eNAMPT. eNAMPT exists in structurally and functionally distinct monomeric and dimeric forms. Dimerisation is essential for the NAD-biosynthetic capacity of NAMPT. Monomeric eNAMPT does not possess NAD-biosynthetic capacity and may exert distinct NAD-independent effects. This study aimed to fully characterise the structure-functional effects of eNAMPT on pancreatic beta cell functional mass and to relate these to beta cell failure in type 2 diabetes. Methods CD-1 mice and serum from obese humans who were without diabetes, with impaired fasting glucose (IFG) or with type 2 diabetes (from the Body Fat, Surgery and Hormone [BodyFatS&H] study) or with or at risk of developing type 2 diabetes (from the VaSera trial) were used in this study. We generated recombinant wild-type and monomeric eNAMPT to explore the effects of eNAMPT on functional beta cell mass in isolated mouse and human islets. Beta cell function was determined by static and dynamic insulin secretion and intracellular calcium microfluorimetry. NAD-biosynthetic capacity of eNAMPT was assessed by colorimetric and fluorescent assays and by native mass spectrometry. Islet cell number was determined by immunohistochemical staining for insulin, glucagon and somatostatin, with islet apoptosis determined by caspase 3/7 activity. Markers of inflammation and beta cell identity were determined by quantitative reverse transcription PCR. Total, monomeric and dimeric eNAMPT and nicotinamide mononucleotide (NMN) were evaluated by ELISA, western blot and fluorometric assay using serum from non-diabetic, glucose intolerant and type 2 diabetic individuals. Results eNAMPT exerts bimodal and concentration- and structure-functional-dependent effects on beta cell functional mass. At low physiological concentrations (~1 ng/ml), as seen in serum from humans without diabetes, eNAMPT enhances beta cell function through NAD-dependent mechanisms, consistent with eNAMPT being present as a dimer. However, as eNAMPT concentrations rise to ~5 ng/ml, as in type 2 diabetes, eNAMPT begins to adopt a monomeric form and mediates beta cell dysfunction, reduced beta cell identity and number, increased alpha cell number and increased apoptosis, through NAD-independent proinflammatory mechanisms. Conclusions/interpretation We have characterised a novel mechanism of beta cell dysfunction in type 2 diabetes. At low physiological levels, eNAMPT exists in dimer form and maintains beta cell function and identity through NAD-dependent mechanisms. However, as eNAMPT levels rise, as in type 2 diabetes, structure-functional changes occur resulting in marked elevation of monomeric eNAMPT, which induces a diabetic phenotype in pancreatic islets. Strategies to selectively target monomeric eNAMPT could represent promising therapeutic strategies for the treatment of type 2 diabetes.

The Cytokine Nicotinamide Phosphoribosyltransferase (eNAMPT; PBEF; Visfatin) Acts as a Natural Antagonist of C-C Chemokine Receptor Type 5 (CCR5)

“…All this data suggests that eNAMPT selectively antagonizes Rantes-dependent calcium signaling. Very recently, Sayers et al, demonstrated a modulation of calcium release mediated by eNAMPT, which, as a dimer, increased glucose-stimulated intracellular calcium levels [23]. In this respect, in our model we were unable to appreciate any effect of eNAMPT alone at the concentrations tested (500-1000 ng/mL = 9-18 nM).…”

“…To add to the complexity, it has been recently demonstrated that the effects on pancreatic beta cell functional mass of eNAMPT are bimodal and concentration and structure functionally dependent [23]. In our study, we used concentrations between 250 and 1000 ng/mL (corresponding to 4.5-18 nM), with the only exception of internalization and cell labelling experiments, in which higher concentrations were used to counteract the higher concentrations used of RANTES (in the second instance, RANTES induces rapid internalization only at high concentrations, while in the first instance RANTES-PE binding could be appreciated only at higher concentrations).…”

Exercise-Induced Benefits on Glucose Handling in a Model of Diet-Induced Obesity Are Reduced by Concurrent Nicotinamide Mononucleotide

“…Earlier work by Revollo and colleagues using Nampt-deficient mice [45] established the importance of NAD + in the regulation of β -cell insulin secretion. Subsequent studies have demonstrated the protective effects of NAD + modulation in isolated islets, through pre-incubation with NAD + precursors and enzymes, before functional assessments such as apoptosis and GSIS measures [46,47]. Here, oral NMN administration in obese mice did not induce any benefits on islet insulin secretion capacity.…”

Nicotinamide Mononucleotide: A Promising Molecule for Therapy of Diverse Diseases by Targeting NAD+ Metabolism

“…The role of eNAMPT has been indicated to be inflammatory promotion, inflammatory suppression (Revollo et al, 2007;Li et al, 2008;Pillai et al, 2013;Zhao et al, 2013;Jing et al, 2014), enhancement of food intake (Brunetti et al, 2012), and regulation of insulin resistance as well as plasma free fatty acid concentration (Stromsdorfer et al, 2016;Nielsen et al, 2018). The function of eNAMPT remains elusive in recent research, demonstrating that low concentration of dimeric eNAMPT benefited for beta cell function through NAD+, however, a higher monomeric eNAMPT level presented hostile effect on beta cell function (Sayers et al, 2020). Obesity has also been associated with the dampened NAD+/SIRT pathway in adipose tissue (Jukarainen et al, 2016).…”