Novel Lipophilic Probe for Detecting Near-Membrane Reactive Oxygen Species Responses and Its Application for Studies of Pancreatic Acinar Cells: Effects of Pyocyanin and L-Ornithine



Chvanov, Michael, Huang, Wei, Jin, Tao, Wen, Li, Armstrong, Jane, Elliot, Vicky, Alston, Ben ORCID: 0000-0001-8255-7950, Burdyga, Alex, Criddle, David, Sutton, Robert ORCID: 0000-0001-6600-562X
et al (show 1 more authors) (2015) Novel Lipophilic Probe for Detecting Near-Membrane Reactive Oxygen Species Responses and Its Application for Studies of Pancreatic Acinar Cells: Effects of Pyocyanin and L-Ornithine. Antioxidants & Redox Signaling, 22 (6). 451 - 464.

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Abstract

Aims: The aim of this study was to develop a fluorescent reactive oxygen species (ROS) probe, which is preferentially localized in cellular membranes and displays a strong change in fluorescence upon oxidation. We also aimed to test the performance of this probe for detecting pathophysiologically relevant ROS responses in isolated cells. Results: We introduced a novel lipophilic ROS probe dihydrorhodamine B octadecyl ester (H2RB-C18). We then applied the new probe to characterize the ROS changes triggered by inducers of acute pancreatitis in pancreatic acinar cells. We resolved ROS changes produced by L-ornithine, L-arginine, cholecystokinin-8, acetylcholine, taurolithocholic acid 3-sulfate, palmitoleic acid ethyl ester, and the bacterial toxin pyocyanin. Particularly prominent ROS responses were induced by pyocyanin and L-ornithine. These ROS responses were accompanied by changes in cytosolic Ca2+concentration ([Ca2+]i), mitochondrial membrane potential (ΔΨ), and NAD(P)H concentration. Innovation: The study describes a novel sensitive lipophilic ROS probe. The probe is particularly suitable for detecting ROS in near-membrane regions and therefore for reporting the ROS environment of plasma membrane channels and pumps. Conclusions: In our experimental conditions, the novel probe was more sensitive than 5-(and-6)-chloromethyl-2′,7′-dichlorodihydrofluorescein (CM-H2DCF) and dihydrorhodamine123 (H2R123) and allowed us to resolve ROS responses to secretagogues, pyocyanin, and L-ornithine. Changes in the fluorescence of the new probe were particularly prominent in the peripheral plasma membrane-associated regions. Our findings suggest that the new probe will be a useful tool in studies of the contribution of ROS to the pathophysiology of exocrine pancreas and other organs/tissues.

Item Type: Article
Depositing User: Symplectic Admin
Date Deposited: 27 Oct 2016 10:33
Last Modified: 14 May 2021 07:12
DOI: 10.1089/ars.2013.5589
Related URLs:
URI: https://livrepository.liverpool.ac.uk/id/eprint/3004001