High-Resolution Glucose Fate-Mapping Reveals LDHB-Dependent Lactate Production by Human Pancreatic Islets

Abstract

Using ¹³C₆ glucose labeling coupled to GC-MS and 2D ¹H-¹³C HSQC NMR spectroscopy, we have obtained a comparative high-resolution map of glucose fate underpinning steady state insulin release and β cell function. In both mouse and human islets, the contribution of glucose to the TCA cycle is similar. Pyruvate-fueling of the TCA cycle is found to be mediated primarily by the activity of pyruvate dehydrogenase (PDH), with only a limited contribution from pyruvate carboxylase (PC). While conversion of pyruvate to lactate by lactate dehydrogenase (LDH) can be detected in both species, lactate accumulation via this route is unexpectedly six-fold higher in human islets. Immunohistochemical and transcriptomic analyses reveal that human islets express abundant LDH, with β cell-specific expression of the LDHB isoenzyme. Thus, glycolytically-derived acetyl CoA preferentially feeds the TCA cycle in both mouse and human β cells. However, human β cells possess the machinery needed to generate significant intracellular lactate, reflecting a key mechanism to balance the reducing activity of NADH-producing pathways.

Ethics Approval Statement: Animal studies were regulated by the Animals (Scientific Procedures) Act 1986 of the U.K. (Personal Project Licences P2ABC3A83 and PP1778740). Approval was granted by the University of Birmingham’s Animal Welfare and Ethical Review Body (AWERB).

Human islets (Lille): Human pancreatic tissues were harvested from brain-dead adult donors in accordance with the Lille clinical islet transplantation program's traceability requirements (clinicaltrials.gov, NCT01123187, NCT00446264, NCT01148680), and were approved in agreement with French regulations and the Ethical Committees of the University of Lille and the Centre Hospitalier Régional Universitaire de Lille.

Human islets (Milan): The use of human islets for research was approved by the Ethics Committee of San Raffaele Hospital in Milan (IPF002-2014).

Pancreas sections (Oxford): Post-mortem pancreas and liver samples were obtained from patients, with appropriate permissions registered under CUREC R83564/RE001.

Studies with human islets and pancreases were approved by the University of Birmingham Ethics Committee, the University of Oxford Ethics Committee, as well as the National Research Ethics Committee (REC 16/NE/0107, Newcastle and North Tyneside, UK).

Liver sections (Oxford): Authorisation for research use of steatotic livers unsuitable for transplantation was obtained by a specialist nurse in organ donation in accordance with NHSBT guidelines, with appropriate permissions registered under REC 14/LO/0182. The study was approved by the North East – Tyne and Weir South research ethics committee (16/NE/0248) and by the NHSBT Research, Innovation and Novel Technologies Advisory Group.

Note:
Funding Information: D.J.H. was supported by MRC (MR/N00275X/1 and MR/S025618/1) and Diabetes UK (17/0005681) Project Grants, as well as a UKRI ERC Frontier Research Guarantee Grant (EP/X026833/1). This project has received funding from the European Research Council (ERC) under the European Union’s Horizon 2020 research and innovation programme (Starting Grant 715884 to D.J.H.). G.G.L. was supported by a Wellcome Trust Senior Fellowship (104612/Z/14/Z). L.H. was supported by a British Heart Foundation Senior Baic Science Research Fellowship (FS/15/56/31645). C.D.L.C. was supported by a clinical research training fellowship from the MRC. G.A.R was supported by a Wellcome Trust Investigator Award (212625/Z/18/Z), MRC Programme grant (MR/R022259/1), Diabetes UK Project grant (BDA16/0005485), CRCHUM start-up funds, an Innovation Canada John R Evans Leader Award, JDRF, CIHR and NIH-NIDDK project grants. D.T. was supported by a Cancer Research UK Programme Grant (C42109/A24747). The research was funded by the National Institute for Health Research (NIHR) Oxford Biomedical Research Centre (BRC). The views expressed are those of the author(s) and not necessarily those of the NHS, the NIHR or the Department of Health. The project involves an element of animal work not funded by the NIHR but by another funder, as well as an element focussed on patients and people appropriately funded by the NIHR.

Declaration of Interests: G.A.R. has received grant funding from, and is a consultant for, Sun Pharmaceuticals Industries Ltd. D.J.H. receives licensing revenue from Celtarys Research.

Keywords: Glucose tracing, islet, metabolism, pyruvate, lactate, LDH, Pyruvate dehydrogenase, GC-MS, NMR