Quantification of Cardiotonic Steroids Potentially Regulated by Paraoxonase 3 in a Rat Model of Chronic Kidney Disease Using UHPLC-Orbitrap-MS

Endogenous cardiotonic steroids (CTSs), such as telocinobufagin (TCB) and marinobufagin (MBG) contain a lactone moiety critical to their binding and signaling through the Na<sup>+</sup>/K<sup>+</sup>-ATPase. Their concentrations elevate in response to sodium intake and under...

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Main Authors: Sabitri Lamichhane, Chrysan J. Mohammed, Steven T. Haller, David J. Kennedy, Dragan Isailovic
Format: Article
Language:English
Published: MDPI AG 2022-11-01
Series:International Journal of Molecular Sciences
Subjects:
Online Access:https://www.mdpi.com/1422-0067/23/21/13565
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author Sabitri Lamichhane
Chrysan J. Mohammed
Steven T. Haller
David J. Kennedy
Dragan Isailovic
author_facet Sabitri Lamichhane
Chrysan J. Mohammed
Steven T. Haller
David J. Kennedy
Dragan Isailovic
author_sort Sabitri Lamichhane
collection DOAJ
description Endogenous cardiotonic steroids (CTSs), such as telocinobufagin (TCB) and marinobufagin (MBG) contain a lactone moiety critical to their binding and signaling through the Na<sup>+</sup>/K<sup>+</sup>-ATPase. Their concentrations elevate in response to sodium intake and under volume-expanded conditions. Paraoxonase 3 (PON3) is an enzyme that can hydrolyze lactone substrates. Here, we examine the role of PON3 in regulating CTS levels in a rat model of chronic kidney diseases (CKD). TCB and MBG were extracted from rat urine samples, and the analyses were carried out using ultra-high pressure liquid chromatography–Orbitrap-mass spectrometry (UHPLC-Orbitrap-MS). Ten-week-old Dahl salt-sensitive wild type (SS-WT) and Dahl salt-sensitive PON3 knockout (SS-PON3 KO) rats were maintained on a high-salt diet (8% NaCl) for 8 weeks to initiate salt-sensitive hypertensive renal disease characteristic of this model. CTS extraction recovery from urine >80% was achieved. For animals maintained on a normal chow diet, the baseline amount of TCB excreted in 24 h urine of SS-PON3 KO rats (6.08 ± 1.47 ng/24 h; or 15.09 ± 3.25 pmol) was significantly higher than for SS-WT rats (1.48 ± 0.69 ng/24 h; or 3.67 ± 1.54 pmol, <i>p</i> < 0.05). Similarly, for the same animals, the amount of excreted MBG was higher in the urine of SS-PON3 KO rats (4.74 ± 1.30 ng/24 h versus 1.03 ± 0.25 ng/24 h in SS-WT; or 11.83 ± 2.91 pmol versus 2.57 ± 0.56 pmol in SS-WT, <i>p</i> < 0.05). For animals on a high-salt diet, the SS-PON3 KO rats had significantly increased levels of TCB (714.52 ± 79.46 ng/24 h; or 1774.85 ± 175.55 pmol) compared to SS-WT control (343.84 ± 157.54 ng/24 h; or 854.09 ± 350.02 pmol, <i>p</i> < 0.05), and comparatively higher levels of MBG were measured for SS-PON3 KO (225.55 ± 82.61 ng/24 h; or 563.19 ± 184.5 pmol) versus SS-WT (157.56 ± 85.53 ng/24 h; or 393.43 ± 191.01 pmol, <i>p</i> > 0.05) rats. These findings suggest that the presence and absence of PON3 dramatically affect the level of endogenous CTSs, indicating its potential role in CTS regulation.
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spelling doaj.art-b78ab6db205d43e884e68b57de4f597b2023-11-24T05:10:04ZengMDPI AGInternational Journal of Molecular Sciences1661-65961422-00672022-11-0123211356510.3390/ijms232113565Quantification of Cardiotonic Steroids Potentially Regulated by Paraoxonase 3 in a Rat Model of Chronic Kidney Disease Using UHPLC-Orbitrap-MSSabitri Lamichhane0Chrysan J. Mohammed1Steven T. Haller2David J. Kennedy3Dragan Isailovic4Department of Chemistry and Biochemistry, University of Toledo, Toledo, OH 43606, USADepartment of Medicine, University of Toledo College of Medicine and Life Sciences, Toledo, OH 43614, USADepartment of Medicine, University of Toledo College of Medicine and Life Sciences, Toledo, OH 43614, USADepartment of Medicine, University of Toledo College of Medicine and Life Sciences, Toledo, OH 43614, USADepartment of Chemistry and Biochemistry, University of Toledo, Toledo, OH 43606, USAEndogenous cardiotonic steroids (CTSs), such as telocinobufagin (TCB) and marinobufagin (MBG) contain a lactone moiety critical to their binding and signaling through the Na<sup>+</sup>/K<sup>+</sup>-ATPase. Their concentrations elevate in response to sodium intake and under volume-expanded conditions. Paraoxonase 3 (PON3) is an enzyme that can hydrolyze lactone substrates. Here, we examine the role of PON3 in regulating CTS levels in a rat model of chronic kidney diseases (CKD). TCB and MBG were extracted from rat urine samples, and the analyses were carried out using ultra-high pressure liquid chromatography–Orbitrap-mass spectrometry (UHPLC-Orbitrap-MS). Ten-week-old Dahl salt-sensitive wild type (SS-WT) and Dahl salt-sensitive PON3 knockout (SS-PON3 KO) rats were maintained on a high-salt diet (8% NaCl) for 8 weeks to initiate salt-sensitive hypertensive renal disease characteristic of this model. CTS extraction recovery from urine >80% was achieved. For animals maintained on a normal chow diet, the baseline amount of TCB excreted in 24 h urine of SS-PON3 KO rats (6.08 ± 1.47 ng/24 h; or 15.09 ± 3.25 pmol) was significantly higher than for SS-WT rats (1.48 ± 0.69 ng/24 h; or 3.67 ± 1.54 pmol, <i>p</i> < 0.05). Similarly, for the same animals, the amount of excreted MBG was higher in the urine of SS-PON3 KO rats (4.74 ± 1.30 ng/24 h versus 1.03 ± 0.25 ng/24 h in SS-WT; or 11.83 ± 2.91 pmol versus 2.57 ± 0.56 pmol in SS-WT, <i>p</i> < 0.05). For animals on a high-salt diet, the SS-PON3 KO rats had significantly increased levels of TCB (714.52 ± 79.46 ng/24 h; or 1774.85 ± 175.55 pmol) compared to SS-WT control (343.84 ± 157.54 ng/24 h; or 854.09 ± 350.02 pmol, <i>p</i> < 0.05), and comparatively higher levels of MBG were measured for SS-PON3 KO (225.55 ± 82.61 ng/24 h; or 563.19 ± 184.5 pmol) versus SS-WT (157.56 ± 85.53 ng/24 h; or 393.43 ± 191.01 pmol, <i>p</i> > 0.05) rats. These findings suggest that the presence and absence of PON3 dramatically affect the level of endogenous CTSs, indicating its potential role in CTS regulation.https://www.mdpi.com/1422-0067/23/21/13565endogenous cardiotonic steroidstelocinobufaginmarinobufaginparaoxonaseurineSPE
spellingShingle Sabitri Lamichhane
Chrysan J. Mohammed
Steven T. Haller
David J. Kennedy
Dragan Isailovic
Quantification of Cardiotonic Steroids Potentially Regulated by Paraoxonase 3 in a Rat Model of Chronic Kidney Disease Using UHPLC-Orbitrap-MS
International Journal of Molecular Sciences
endogenous cardiotonic steroids
telocinobufagin
marinobufagin
paraoxonase
urine
SPE
title Quantification of Cardiotonic Steroids Potentially Regulated by Paraoxonase 3 in a Rat Model of Chronic Kidney Disease Using UHPLC-Orbitrap-MS
title_full Quantification of Cardiotonic Steroids Potentially Regulated by Paraoxonase 3 in a Rat Model of Chronic Kidney Disease Using UHPLC-Orbitrap-MS
title_fullStr Quantification of Cardiotonic Steroids Potentially Regulated by Paraoxonase 3 in a Rat Model of Chronic Kidney Disease Using UHPLC-Orbitrap-MS
title_full_unstemmed Quantification of Cardiotonic Steroids Potentially Regulated by Paraoxonase 3 in a Rat Model of Chronic Kidney Disease Using UHPLC-Orbitrap-MS
title_short Quantification of Cardiotonic Steroids Potentially Regulated by Paraoxonase 3 in a Rat Model of Chronic Kidney Disease Using UHPLC-Orbitrap-MS
title_sort quantification of cardiotonic steroids potentially regulated by paraoxonase 3 in a rat model of chronic kidney disease using uhplc orbitrap ms
topic endogenous cardiotonic steroids
telocinobufagin
marinobufagin
paraoxonase
urine
SPE
url https://www.mdpi.com/1422-0067/23/21/13565
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