Mariola Olkowicz (Presenter)
Medical University of Gdansk
Bio: I'm a postdoctoral scientist within the TEAM programme of Foundation for Polish Science at the Medical University of Gdansk where perform research tasks in the project: “Nucleotides in pathology, diagnosis and therapy of heart disease”. The main area of my research includes: characterization of CVD (cardiovascular) biomarkers in body fluids/tissues, targeted quantitative analysis of proteins with the use of miniaturized HPLC and tandem mass spectrometry techniques, new methods/solutions in protein fractionation from complex biological matrices (development of the new chromatographic media), differential analysis of global proteome between physiological and pathophysiological states in the cardiovascular system.
Authorship: Mariola Olkowicz (1,2), Joanna Suraj (3,4), Agnieszka Kij (3,4), Maria Walczak (3,4), Stefan Chlopicki (3,4), Ryszard T. Smolenski (1)
(1) Medical University of Gdansk, Gdansk, Poland, (2) Poznan University of Life Sciences, Poznan, Poland, (3) JCET, Krakow, Poland, (4) Jagiellonian University Medical College, Krakow, Poland
| Short Abstract The aim of this study was to develop an analytical methodology for accurate quantification of angiotensins: Ang I, Ang II, Ang-(1-7), Ang III and Ang IV in plasma of atherosclerotic mice. A triple quadrupole mass detector equipped with chip-based nanospray source connected to nanoHPLC was used. Plasma angiotensin profile was substantially modified in ApoE/LDLR double knock-out mice with increase in concentration of Ang II from 37.6 ± 21.3 pg mL-1 in WT to 200.2 ± 47.6 pg mL-1. Concentrations of Ang I, III and IV were also elevated 3-10 fold in ApoE-/-/LDLR-/- mice while that of Ang-(1-7) was unchanged. |
Long Abstract
The renin-angiotensin system (RAS) and its effector molecules, angiotensin peptides, are known to exert a wide variety of actions in the cardiovascular and renal systems. Understanding the RAS function requires precise and comprehensive quantification of its elements. The aim of this study was to develop an analytical methodology for the rapid extraction, separation and sensitive detection of the selected angiotensin peptides in plasma of atherosclerotic mice.
A triple quadrupole (TSQ Vantage, Thermo) mass detector equipped with chip-based nanospray source (ChipMate, Advion) connected to nanoflow HPLC (Dionex RSLCnano, Thermo) was used to simultaneously determine the concentrations of angiotensin (Ang) I, Ang II, Ang-(1-7), Ang III and Ang IV in biological samples. Plasma was collected from eight ApoE-/-/LDLR-/- and five wild-type (WT) mice. The samples were deproteinated with acetonitrile, and separated by a reverse-phase C18 column using acetonitrile in water with 1% acetic acid as a mobile phase. Multiple reaction monitoring (MRM) mode was used for recording two transitions (from doubly/triply charged ions) for each peptide.
The method provided excellent analytical performance with LOQ < 5 pg mL-1. Plasma angiotensin profile was substantially modified in ApoE/LDLR double knock-out mice with increase in concentration of Ang II from 37.6 ± 21.3 pg mL-1 in WT to 200.2 ± 47.6 pg mL-1. Concentrations of Ang I, III and IV were also increased 3-10 fold in ApoE-/-/LDLR-/- mice while that of Ang-(1-7) was unchanged.
We conclude that the method developed could be effectively used for accurate, comprehensive profiling of angiotensin peptides in mouse plasma. We identified substantial changes in renin-angiotensin system in a genetic mouse model of atherosclerosis consistent with the overactivation of angiotensin converting enzyme (ACE) and the impairment of ACE2.
References & Acknowledgements:
This study was supported by the European Union from the resources of the European Regional Development Fund under the Innovative Economy Programme (a grant coordinated by JCET-UJ, No POIG.01.01.02-00-069/09) and TEAM programme of the Foundation for Polish Science (TEAM/2011-8/7).
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