Pancreatic Acinar Cell Preparation for Oxygen Consumption and Lactate Production Analysis



Armstrong, Jane, Sutton, Robert ORCID: 0000-0001-6600-562X and Criddle, David ORCID: 0000-0003-2952-8450
(2020) Pancreatic Acinar Cell Preparation for Oxygen Consumption and Lactate Production Analysis. BIO-PROTOCOL, 10 (10). e3627-.

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Abstract

Mitochondrial dysfunction is a principal feature of acute pancreatitis (AP) although the underlying mechanisms are still unclear. AP precipitants induce Ca<sup>2+</sup>-dependent formation of the mitochondrial permeability transition pore (MPTP) in pancreatic acinar cells (PACs), leading to ATP depletion and necrosis. Evaluations of mitochondrial bioenergetics have mainly been performed in isolated PACs using confocal microscopy, with assessment of mitochondrial membrane potential, NADH/FAD<sup>+</sup> and ATP levels, coupled with patch-clamp electrophysiology. These studies are technically demanding and time-consuming. Application of Seahorse flux analysis now allows detailed investigations of bioenergetics changes to be performed in cell populations using a multi-well plate-reader format; rates of oxygen consumption (OCR) and extracellular acidification (ECAR) provide important information about cellular respiration and glycolysis, respectively. Parameters such as maximal respiration, ATP-linked capacity and proton leak can be derived from application of a respiratory function "stress" test that involves pharmacological manipulation of the electron transport chain. The use of Seahorse Flux analysis therefore provides a quick, and convenient means to measure detailed cellular bioenergetics and allows results to be coupled with other plate-reader based assays, providing a fuller understanding of the pathophysiological consequences of mitochondrial bioenergetics alterations.

Item Type: Article
Uncontrolled Keywords: Mitochondrial dysfunction, Bioenergetics, Seahorse, Respiration, Glycolysis, Pancreatic acinar cells, Acute pancreatitis
Depositing User: Symplectic Admin
Date Deposited: 23 Jun 2020 09:43
Last Modified: 18 Jan 2023 23:48
DOI: 10.21769/bioprotoc.3627
Related URLs:
URI: https://livrepository.liverpool.ac.uk/id/eprint/3091327