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Studying the Innate Immune Response to Myocardial Infarction in a Highly Efficient Experimental Animal Model Cover

Studying the Innate Immune Response to Myocardial Infarction in a Highly Efficient Experimental Animal Model

Romanian Journal of Cardiology
Volume 31 (2021): Issue 3 (September 2021)
By: Razvan Gheorghita Mares,  Doina Manu,  Istvan Adorjan Szabo,  Mihaela Elena Tomut,  Gabriela Pintican,  Bogdan Cordos,  Gabriel Jakobsson,  Minodora Dobreanu,  Ovidiu Simion Cotoi and  Alexandru Schiopu  
Open Access
|May 2022

Figures & Tables

Figure 1

Permanent coronary artery occlusion without mechanical ventilation in mice.Images show the schematic presentation of the surgical technique. A, Preparing the operating table. B, Anesthetizing the mouse in the induction chamber. C, Placing the unconscious mouse on a warm pad and removing the fur from the chest. D, Making a small skin cut in the middle position of precordial chest. E, Dissecting and retracting the pectoral major and minor muscles. F, Exposing the 4th intercostal space. G, Placing the curved mosquito clamp at the 4th intercostal space level with the help of a forceps and forcing the mosquito to open the pleural membrane and pericardium. H–J, Smoothly and gently lifting out the heart through the window, locating and ligating the LAD with silk suture. K, Placing the heart back into the thorax promptly after ligation, with manual evacuation of air by gentle but efficient side chest pressure. L, closure of the skin with Prolene suture.
Permanent coronary artery occlusion without mechanical ventilation in mice.Images show the schematic presentation of the surgical technique. A, Preparing the operating table. B, Anesthetizing the mouse in the induction chamber. C, Placing the unconscious mouse on a warm pad and removing the fur from the chest. D, Making a small skin cut in the middle position of precordial chest. E, Dissecting and retracting the pectoral major and minor muscles. F, Exposing the 4th intercostal space. G, Placing the curved mosquito clamp at the 4th intercostal space level with the help of a forceps and forcing the mosquito to open the pleural membrane and pericardium. H–J, Smoothly and gently lifting out the heart through the window, locating and ligating the LAD with silk suture. K, Placing the heart back into the thorax promptly after ligation, with manual evacuation of air by gentle but efficient side chest pressure. L, closure of the skin with Prolene suture.

Figure 2

Validation of LAD ligation by electrocardiographyElevations of the S-T segment appear a few minutes after LAD ligation, confirming the successful induction of ischemia.
Validation of LAD ligation by electrocardiographyElevations of the S-T segment appear a few minutes after LAD ligation, confirming the successful induction of ischemia.

Figure 3

Inflammatory infiltration post-AMI determined by histology and immunohistochemistryA, H&E at day 1 after AMI. B, H&E at day 3 after AMI. C, S100A8/A9 positive cells at day 1 after AMI. D, S100A8/A9 positive cells at day 3 after AMI. E, S100A8/A9 positive cells at day 7 after AMI. F, S100A8/A9 positive cells at day 14 after AMI. All 10X magnification.
Inflammatory infiltration post-AMI determined by histology and immunohistochemistryA, H&E at day 1 after AMI. B, H&E at day 3 after AMI. C, S100A8/A9 positive cells at day 1 after AMI. D, S100A8/A9 positive cells at day 3 after AMI. E, S100A8/A9 positive cells at day 7 after AMI. F, S100A8/A9 positive cells at day 14 after AMI. All 10X magnification.

Figure 4

Fibrous scar formation, neovascularization and myofibroblast presence at 7 and 14 days after AMIA, Masson's trichrome staining. B, CD31 positive cells. C, α-SMA positive cells. All 10X magnification.
Fibrous scar formation, neovascularization and myofibroblast presence at 7 and 14 days after AMIA, Masson's trichrome staining. B, CD31 positive cells. C, α-SMA positive cells. All 10X magnification.

Figure 5

Inflammatory cells dynamics after AMI and the flow cytometry gating strategyA, neutrophils in the infarcted myocardium at 1, 3 and 7 days after AMI. (a), (b) single cells selection using the forward scatter/side scatter plots. (c) myeloid cells (CD45+CD11b+) derived from the single cells gate. (d) Ly6G/CD11b plot to identify neutrophils (CD45+CD11b+Ly6G+). B, monocytes in blood at 1, 3 and 7 days after AMI. (a) cells selection using forward/side scatter plots, to eliminate debris. (b) myeloid cells. (c) CD115/CD11b plot identifying total monocytes (CD45+CD11b+CD115+ cells). (d) CD115/Ly6C plot to divide monocytes into inflammatory Ly6Chi monocytes (upper gate) and patrolling Ly6Clow monocytes (lower gate). C, macrophages in the infarcted myocardium at 1, 3 and 7 days after AMI. (a) cells selection using forward/side scatter plots. (b) myeloid cells. (c) F4/80/CD11b plot identifying total macrophages (CD45+CD11b+F4/80+ cells). (d) MerTK/Ly6C gate to identify the reparatory Ly6ClowMerTKhi macrophages (right lower quadrant) and inflammatory Ly6ChiMerTKlow macrophages (left upper quadrant).
Inflammatory cells dynamics after AMI and the flow cytometry gating strategyA, neutrophils in the infarcted myocardium at 1, 3 and 7 days after AMI. (a), (b) single cells selection using the forward scatter/side scatter plots. (c) myeloid cells (CD45+CD11b+) derived from the single cells gate. (d) Ly6G/CD11b plot to identify neutrophils (CD45+CD11b+Ly6G+). B, monocytes in blood at 1, 3 and 7 days after AMI. (a) cells selection using forward/side scatter plots, to eliminate debris. (b) myeloid cells. (c) CD115/CD11b plot identifying total monocytes (CD45+CD11b+CD115+ cells). (d) CD115/Ly6C plot to divide monocytes into inflammatory Ly6Chi monocytes (upper gate) and patrolling Ly6Clow monocytes (lower gate). C, macrophages in the infarcted myocardium at 1, 3 and 7 days after AMI. (a) cells selection using forward/side scatter plots. (b) myeloid cells. (c) F4/80/CD11b plot identifying total macrophages (CD45+CD11b+F4/80+ cells). (d) MerTK/Ly6C gate to identify the reparatory Ly6ClowMerTKhi macrophages (right lower quadrant) and inflammatory Ly6ChiMerTKlow macrophages (left upper quadrant).

Figure 6

Immune cell dynamics post-AMI in blood and myocardium during the first week post-AMI, determined by flow cytometryPercentages of: A, neutrophils in the infarcted myocardium out of total myeloid cells. B, circulating monocytes out of total myeloid cells in blood. C, inflammatory Ly6Chi monocytes out of total blood monocytes. D, macrophages in the infarcted myocardium out of total myeloid cells. E, reparatory Ly6CloMerTKhi macrophages in the infarcted myocardium out of total macrophages. F, Ly6ChiMerTKlo macrophages in the infarcted myocardium out of total macrophages.
Immune cell dynamics post-AMI in blood and myocardium during the first week post-AMI, determined by flow cytometryPercentages of: A, neutrophils in the infarcted myocardium out of total myeloid cells. B, circulating monocytes out of total myeloid cells in blood. C, inflammatory Ly6Chi monocytes out of total blood monocytes. D, macrophages in the infarcted myocardium out of total myeloid cells. E, reparatory Ly6CloMerTKhi macrophages in the infarcted myocardium out of total macrophages. F, Ly6ChiMerTKlo macrophages in the infarcted myocardium out of total macrophages.
DOI: https://doi.org/10.47803/rjc.2021.31.3.573 | Journal eISSN: 2734-6382 | Journal ISSN: 1220-658X
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Language: English
Page range: 573 - 585
Published on: May 5, 2022
Published by: Romanian Society of Cardiology
In partnership with: Paradigm Publishing Services
Publication frequency: 4 issues per year
Keywords:
acute myocardial infarction,
experimental model,
innate immunity,
neutrophils,
monocytes,
macrophages,
myocardial fibrosis,
revascularization
Related subjects:
Medicine,
Clinical medicine,
Internal medicine,
Cardiology,
Pediatrics and juvenile medicine,
Paediatric cardiology,
Surgery,
Cardiac surgery

© 2022 Razvan Gheorghita Mares, Doina Manu, Istvan Adorjan Szabo, Mihaela Elena Tomut, Gabriela Pintican, Bogdan Cordos, Gabriel Jakobsson, Minodora Dobreanu, Ovidiu Simion Cotoi, Alexandru Schiopu, published by Romanian Society of Cardiology
This work is licensed under the Creative Commons Attribution 4.0 License.

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