8e97e8466d56930c822d80354e7b18c3 302825.pdf 3e7c06edb1e068b01ead1c3eaca19c9ca46cc375 302825.pdf 78488db54b0c8a6959f7d5b40dca3597f80aa70c0a790599fec37c79eb744014 302825.pdf Title: Increased Density of Endogenous Adenosine A2A Receptors in Atrial Fibrillation: From Cellular and Porcine Models to Human Patients Subject: Adenosine, an endogenous nucleoside, plays a critical role in maintaining homeostasis during stressful situations, such as energy deprivation or cellular damage. Therefore, extracellular adenosine is generated locally in tissues under conditions such as hypoxia, ischemia, or inflammation. In fact, plasma levels of adenosine in patients with atrial fibrillation (AF) are elevated, which also correlates with an increased density of adenosine A2A receptors (A2ARs) both in the right atrium and in peripheral blood mononuclear cells (PBMCs). The complexity of adenosine-mediated effects in health and disease requires simple and reproducible experimental models of AF. Here, we generate two AF models, namely the cardiomyocyte cell line HL-1 submitted to Anemonia toxin II (ATX-II) and a large animal model of AF, the right atrium tachypaced pig (A-TP). We evaluated the density of endogenous A2AR in those AF models. Treatment of HL-1 cells with ATX-II reduced cell viability, while the density of A2AR increased significantly, as previously observed in cardiomyocytes with AF. Next, we generated the animal model of AF based on tachypacing pigs. In particular, the density of the key calcium regulatory protein calsequestrin-2 was reduced in A-TP animals, which is consistent with the atrial remodelling shown in humans suffering from AF. Likewise, the density of A2AR in the atrium of the AF pig model increased significantly, as also shown in the biopsies of the right atrium of subjects with AF. Overall, our findings revealed that these two experimental models of AF mimicked the alterations in A2AR density observed in patients with AF, making them attractive models for studying the adenosinergic system in AF. Keywords: adenosine A2AR receptor; atrial fibrillation; HL-1 cells; tachypaced pig; A2AR density; AF patients Author: Héctor Godoy-Marín, Verónica Jiménez-Sábado, Carmen Tarifa, Antonino Ginel, Joana Larupa Dos Santos, Bo Hjorth Bentzen, Leif Hove-Madsen and Francisco Ciruela Creator: LaTeX with hyperref Producer: pdfTeX-1.40.21 CreationDate: Tue Feb 21 10:04:23 2023 CET ModDate: Wed Feb 22 03:17:08 2023 CET Custom Metadata: no Metadata Stream: no Tagged: no UserProperties: no Suspects: no Form: none JavaScript: no Pages: 11 Encrypted: no Page size: 595.276 x 841.89 pts (A4) Page rot: 0 File size: 1779505 bytes Optimized: no PDF version: 1.7 name type encoding emb sub uni object ID ------------------------------------ ----------------- ---------------- --- --- --- --------- JYPLJJ+VnURWPalladioL-Bold Type 1 Custom yes yes no 10 0 OPZDHV+VnURWPalladioL Type 1 Custom yes yes no 15 0 ZFQLCG+URWPalladioL-Bold Type 1 Custom yes yes no 19 0 RCQYDM+URWPalladioL-Roma Type 1 Custom yes yes no 24 0 WJPFQU+URWPalladioL-Ital Type 1 Custom yes yes no 28 0 IRWHZX+TimesNewRomanPSMT Type 1C WinAnsi yes yes no 46 0 CKUPDE+TimesNewRomanPS-BoldItalicMT Type 1C WinAnsi yes yes no 49 0 IHGURL+EURM7 Type 1 Builtin yes yes no 71 0 QMDDXF+CMSY10 Type 1 Builtin yes yes no 78 0 NCNJDD+PalatinoLinotype,Bold TrueType WinAnsi yes yes no 91 0 NCNJDF+PalatinoLinotype TrueType WinAnsi yes yes no 94 0 NCNJEH+PalatinoLinotype,Italic TrueType WinAnsi yes yes no 97 0 ZXDKIP+EURM10 Type 1 Builtin yes yes no 104 0 NCNJDD+PalatinoLinotype,Bold TrueType WinAnsi yes yes no 115 0 NCNJDE+PalatinoLinotype CID TrueType Identity-H yes yes yes 118 0 NCNJDF+PalatinoLinotype TrueType WinAnsi yes yes no 124 0 NCNJEH+PalatinoLinotype,Italic TrueType WinAnsi yes yes no 127 0 NCNJDD+PalatinoLinotype,Bold TrueType WinAnsi yes yes no 141 0 NCNJDE+PalatinoLinotype CID TrueType Identity-H yes yes yes 144 0 NCNJDF+PalatinoLinotype TrueType WinAnsi yes yes no 150 0 NCNJEH+PalatinoLinotype,Italic TrueType WinAnsi yes yes no 153 0 NCNJDD+PalatinoLinotype,Bold TrueType WinAnsi yes yes no 165 0 NCNJDE+PalatinoLinotype CID TrueType Identity-H yes yes yes 168 0 NCNJDF+PalatinoLinotype TrueType WinAnsi yes yes no 174 0 NCNJEH+PalatinoLinotype,Italic TrueType WinAnsi yes yes no 177 0 NCNJDD+PalatinoLinotype,Bold TrueType WinAnsi yes yes no 192 0 NCNJDE+PalatinoLinotype CID TrueType Identity-H yes yes yes 195 0 NCNJDF+PalatinoLinotype TrueType WinAnsi yes yes no 201 0 NCNJEH+PalatinoLinotype,Italic TrueType WinAnsi yes yes no 204 0 Jhove (Rel. 1.28.0, 2023-05-18) Date: 2024-11-19 02:45:30 CET RepresentationInformation: 302825.pdf ReportingModule: PDF-hul, Rel. 1.12.4 (2023-03-16) LastModified: 2024-11-18 10:01:32 CET Size: 1779505 Format: PDF Version: 1.7 Status: Well-Formed and valid SignatureMatches: PDF-hul MIMEtype: application/pdf PDFMetadata: Objects: 346 FreeObjects: 1 IncrementalUpdates: 0 DocumentCatalog: PageLayout: SinglePage PageMode: UseNone Outlines: Item: Title: Introduction Destination: section.1 Item: Title: Results Destination: section.2 Children: Item: Title: Endogenous A2AR Density in the HL-1 Cell Line Destination: subsection.2.1 Item: Title: Expression of A2AR in the Atrium of a Porcine Model of AF Destination: subsection.2.2 Item: Title: Expression of A2AR in the Atrium of Patients with AF Destination: subsection.2.3 Item: Title: Discussion Destination: section.3 Item: Title: Materials and Methods Destination: section.4 Children: Item: Title: Reagents Destination: subsection.4.1 Item: Title: Cell Culture and Viability Assays Destination: subsection.4.2 Item: Title: Porcine Model of AF Destination: subsection.4.3 Item: Title: Human Samples and Subject Demographics Destination: subsection.4.4 Item: Title: Preparation of Membrane Extracts Destination: subsection.4.5 Item: Title: Gel Electrophoresis and Immunoblotting Destination: subsection.4.6 Item: Title: Data and Statistical Analysis Destination: subsection.4.7 Item: Title: References Destination: subsection.4.7 Info: Title: Increased Density of Endogenous Adenosine A2A Receptors in Atrial Fibrillation: From Cellular and Porcine Models to Human Patients Author: Héctor Godoy-Marín, Verónica Jiménez-Sábado, Carmen Tarifa, Antonino Ginel, Joana Larupa Dos Santos, Bo Hjorth Bentzen, Leif Hove-Madsen and Francisco Ciruela Subject: Adenosine, an endogenous nucleoside, plays a critical role in maintaining homeostasis during stressful situations, such as energy deprivation or cellular damage. Therefore, extracellular adenosine is generated locally in tissues under conditions such as hypoxia, ischemia, or inflammation. In fact, plasma levels of adenosine in patients with atrial fibrillation (AF) are elevated, which also correlates with an increased density of adenosine A2A receptors (A2ARs) both in the right atrium and in peripheral blood mononuclear cells (PBMCs). The complexity of adenosine-mediated effects in health and disease requires simple and reproducible experimental models of AF. Here, we generate two AF models, namely the cardiomyocyte cell line HL-1 submitted to Anemonia toxin II (ATX-II) and a large animal model of AF, the right atrium tachypaced pig (A-TP). We evaluated the density of endogenous A2AR in those AF models. Treatment of HL-1 cells with ATX-II reduced cell viability, while the density of A2AR increased significantly, as previously observed in cardiomyocytes with AF. Next, we generated the animal model of AF based on tachypacing pigs. In particular, the density of the key calcium regulatory protein calsequestrin-2 was reduced in A-TP animals, which is consistent with the atrial remodelling shown in humans suffering from AF. Likewise, the density of A2AR in the atrium of the AF pig model increased significantly, as also shown in the biopsies of the right atrium of subjects with AF. Overall, our findings revealed that these two experimental models of AF mimicked the alterations in A2AR density observed in patients with AF, making them attractive models for studying the adenosinergic system in AF. Keywords: adenosine A2AR receptor; atrial fibrillation; HL-1 cells; tachypaced pig; A2AR density; AF patients Creator: LaTeX with hyperref Producer: pdfTeX-1.40.21 CreationDate: Tue Feb 21 10:04:23 CET 2023 ModDate: Wed Feb 22 03:17:08 CET 2023 ID: 0xd06f92539f222a0383657c53a0250ce2, 0xd06f92539f222a0383657c53a0250ce2 Filters: FilterPipeline: FlateDecode Fonts: Type1: Font: BaseFont: ZFQLCG+URWPalladioL-Bold FontSubset: true FirstChar: 3 LastChar: 122 FontDescriptor: FontName: ZFQLCG+URWPalladioL-Bold Flags: Symbolic FontBBox: -152, -301, 1000, 935 FontFile: true EncodingDictionary: Differences: true Font: BaseFont: IHGURL+EURM7 FontSubset: true FirstChar: 11 LastChar: 11 FontDescriptor: FontName: IHGURL+EURM7 Flags: Symbolic FontBBox: -5, -243, 1229, 720 FontFile: true Font: BaseFont: RCQYDM+URWPalladioL-Roma FontSubset: true FirstChar: 2 LastChar: 252 FontDescriptor: FontName: RCQYDM+URWPalladioL-Roma Flags: Symbolic FontBBox: -166, -283, 1021, 943 FontFile: true EncodingDictionary: 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