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      Custodiol HTK versus Plegisol: in-vitro comparison with the use of immature (H9C2) and mature (HCM) cardiomyocytes cultures

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          Abstract

          Background

          Although cardioplegia is used since the ‘70s of the last century, debate on cardioprotection during cardio-surgical procedures is still actual. The selection of a particular method depends mainly on the preferences and experience of a specific center or even surgeon. Crystalloid cardioplegia is an aqueous ion solution similar to intracellular (Custodiol HTK) or extracellular (Plegisol) fluid. The potensional clinical advantages of relatively new idea of cardioplegia solution based on intracellular composition (Custodiol HTK) justifies futher research, but only a few used cultured cells in laboratory conditions.

          Methods

          In this study, the authors sought to compare Custodiol HTK with Plegisol cardioplegia solutions using an in-vitro model simulating cardioplegic arrest. The efficacy of myocardial protection during ischemia was investigated with susceptible indicators like the appearance of the deleterious effect of reactive oxygen species and oxidative stress markers. Immersed human cardiomyocytes and rat cardiomyoblasts H9C2 in cardioplegia for 4 h were examined for expression of oxidative stress markers (MnSOD, iNOS, HSP27), cardioplegic solutions cytotoxicity, and peroxidation damage of the cell’s lipids and proteins. All tests were performed after 0.5 h, 1 h, 2 h, and 4 h of incubation in identical physical and biological conditions, which is difficult to achieve in clinical trials.

          Results

          The lower cytotoxicity index performed on matured cells of human cardiomyocytes and highest dehydrogenase level showed after incubation with Custodiol HTK. This did not apply to tests on immature cells H9C2. Custodiol HTK induced significantly stronger iNOS expression. The decrease of HSP27 concentration has been instantaneous and maintained troughout the study only in both cultures incubated with Custodiol HTK. The other tests: lipid peroxidation, carbonyl groups concentration and MnSOD expression show no clear superiority evidence of used cardioplegic solutions.

          Conclusions

          Considering proceeded examinations on cultured cardiomyocytes, Custodiol HTK appears to be safer than Plegisol.

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          Most cited references37

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          Oxidative stress and myocardial injury in the diabetic heart.

          Reactive oxygen or nitrogen species play an integral role in both myocardial injury and repair. This dichotomy is differentiated at the level of species type, amount and duration of free radical generated. Homeostatic mechanisms designed to prevent free radical generation in the first instance, scavenge, or enzymatically convert them to less toxic forms and water, playing crucial roles in the maintenance of cellular structure and function. The outcome between functional recovery and dysfunction is dependent upon the inherent ability of these homeostatic antioxidant defences to withstand acute free radical generation, in the order of seconds to minutes. Alternatively, pre-existent antioxidant capacity (from intracellular and extracellular sources) may regulate the degree of free radical generation. This converts reactive oxygen and nitrogen species to the role of second messenger involved in cell signalling. The adaptive capacity of the cell is altered by the balance between death or survival signal converging at the level of the mitochondria, with distinct pathophysiological consequences that extends the period of injury from hours to days and weeks. Hyperglycaemia, hyperlipidaemia and insulin resistance enhance oxidative stress in the diabetic myocardium that cannot adapt to ischaemia-reperfusion. Altered glucose flux, mitochondrial derangements and nitric oxide synthase uncoupling in the presence of decreased antioxidant defence and impaired prosurvival cell signalling may render the diabetic myocardium more vulnerable to injury, remodelling and heart failure. Copyright © 2012 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
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            The Dual Role of Inducible Nitric Oxide Synthase in Myocardial Ischemia/Reperfusion Injury: Friend or Foe?

            Nitric oxide synthases (NOSs) are a family of enzymes that are responsible for the synthesis of nitric oxide (NO) from the amino acid L-arginine in the body. Among the three key NOSs, the expression of inducible NOS (iNOS) can only be induced by inflammatory stimuli and contribute to the large amount of NO production. iNOS-derived NO plays an important role in various physiological and pathophysiological conditions, including the ischemic heart disease. Nowadays, the development of specific iNOS inhibitors and the availability of iNOS knockout mice have provided substantial evidence to support the role of iNOS/NO signaling in the myocardium. Nevertheless, the role of iNOS/NO signaling in the myocardial ischemic reperfusion injury is very complex and highly perplexing; both detrimental and beneficial effects of iNOS have been described. Thus, this review will aim at providing basic insights into the current progress of the role of iNOS in myocardial ischemia reperfusion injury. A better understanding of the dual role of iNOS in details may help facilitate the development of more effective therapies for the management of ischemic heart diseases.
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              Bradykinin induces mitochondrial ROS generation via NO, cGMP, PKG, and mitoKATP channel opening and leads to cardioprotection.

              Bradykinin (BK) mimics ischemic preconditioning by generating reactive oxygen species (ROS). To identify intermediate steps that lead to ROS generation, rabbit cardiomyocytes were incubated in reduced MitoTracker Red stain, which becomes fluorescent after exposure to ROS. Fluorescence intensity in treated cells was expressed as a percentage of that in paired, untreated cells. BK (500 nM) caused a 51 +/- 16% increase in ROS generation (P < 0.001). Coincubation with either the BK B2-receptor blocker HOE-140 (5 microM) or the free radical scavenger N-(2-mercaptopropionyl)glycine (1 mM) prevented this increase, which confirms that the response was receptor mediated and ROS were actually being measured. Closing mitochondrial ATP-sensitive K+ (mitoKATP) channels with 5-hydroxydecanoate (5-HD, 1 mM) prevented increased ROS generation. BK-induced ROS generation was blocked by Nomega-nitro-m-arginine methyl ester (m-NAME, 200 microM), which implicates nitric oxide as an intermediate. Blockade of guanylyl cyclase with 1-H-[1,2,4]oxadiazole[4,3-a]quinoxalin-1-one (ODQ, 10 microM) aborted BK-induced ROS generation but not that from diazoxide, a direct opener of mitoKATP channels. The protein kinase G (PKG) blocker 8-bromoguanosine-3',5'-cyclic monophosphorothioate (25 microM) eliminated the effects of BK. Conversely, direct activation of PKG with 8-(4-chlorophenylthio)-guanosine-3',5'-cyclic monophosphate (100 microM) increased ROS generation (39 +/- 15%; P < 0.004) similar to BK. This increase was blocked by 5-HD. Finally, the nitric oxide donor S-nitroso-N-acetylpenicillamine (1 microM) increased ROS by 34 +/- 6%. This increase was also blocked by 5-HD. In intact rabbit hearts, BK (400 nM) decreased infarction from 30.5 +/- 3.0 of the risk zone in control hearts to 11.9 +/- 1.4% (P < 0.01). This protection was aborted by either 200 microM m-NAME or 2 microM ODQ (35.4 +/- 5.7 and 30.4 +/- 3.0% infarction, respectively; P = not significant vs. control). Hence, BK preconditions through receptor-mediated production of nitric oxide, which activates guanylyl cyclase. The resulting cGMP activates PKG, which opens mitoKATP. Subsequent release of ROS triggers cardioprotection.
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                Author and article information

                Contributors
                rafal.nowicki@umw.edu.pl
                mikolaj.berezowski@umw.edu.pl
                julita.kulbacka@umw.edu.pl
                k.biezunskakusiak@gmail.com
                marek.jasinski@umw.edu.pl
                jolanta.saczko@umw.edu.pl
                Journal
                BMC Cardiovasc Disord
                BMC Cardiovasc Disord
                BMC Cardiovascular Disorders
                BioMed Central (London )
                1471-2261
                17 March 2022
                17 March 2022
                2022
                : 22
                : 108
                Affiliations
                [1 ]GRID grid.4495.c, ISNI 0000 0001 1090 049X, Clinical Department of Cardiac Surgery, , Wroclaw Medical University, ; Wroclaw, Poland
                [2 ]GRID grid.4495.c, ISNI 0000 0001 1090 049X, Doctoral School, , Wroclaw Medical University, ; Wroclaw, Poland
                [3 ]GRID grid.4495.c, ISNI 0000 0001 1090 049X, Department of Molecular and Cellular Biology, Faculty of Pharmacy, , Wroclaw Medical University, ; Wroclaw, Poland
                [4 ]Children’s Memorial Pediatric Health Institute, Warsaw, Poland
                Article
                2536
                10.1186/s12872-022-02536-6
                8928626
                35296256
                5680743f-33d9-4501-8e2d-0531d8a9ca9e
                © The Author(s) 2022

                Open AccessThis article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. The Creative Commons Public Domain Dedication waiver ( http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data.

                History
                : 30 July 2021
                : 22 February 2022
                Funding
                Funded by: Polish Ministry of Science and Higher Education
                Award ID: No N N403 573038
                Award Recipient :
                Categories
                Research
                Custom metadata
                © The Author(s) 2022

                Cardiovascular Medicine
                cardioplegia,cardiomyocyte cultures,oxidative stress
                Cardiovascular Medicine
                cardioplegia, cardiomyocyte cultures, oxidative stress

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