Myocardial rescue with autologous mitochondrial transplantation in a porcine model of ischemia/reperfusion Aditya K. Kaza, MD, Isaac Wamala, MD, Ingeborg.

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Myocardial rescue with autologous mitochondrial transplantation in a porcine model of ischemia/reperfusion Aditya K. Kaza, MD, Isaac Wamala, MD, Ingeborg Friehs, MD, Joseph D. Kuebler, MD, Rahul H. Rathod, MD, Ignacio Berra, MD, Maria Ericsson, BS, Rouan Yao, BS, Jerusha K. Thedsanamoorthy, BS, David Zurakowski, PhD, Sidney Levitsky, MD, Pedro J. del Nido, MD, Douglas B. Cowan, PhD, James D. McCully, PhD The Journal of Thoracic and Cardiovascular Surgery Volume 153, Issue 4, Pages 934-943 (April 2017) DOI: 10.1016/j.jtcvs.2016.10.077 Copyright © 2016 The American Association for Thoracic Surgery Terms and Conditions

Figure 1 A, Experimental protocol. Injection sites in the area at risk are indicated. B, Representative electron micrographs of isolated pig skeletal muscle mitochondria. The isolated mitochondria were free from cellular contamination and were electron dense and had preserved morphology and shape. (Scale bars: 1 μm.) RI, Regional ischemia; ECG, echocardiography. The Journal of Thoracic and Cardiovascular Surgery 2017 153, 934-943DOI: (10.1016/j.jtcvs.2016.10.077) Copyright © 2016 The American Association for Thoracic Surgery Terms and Conditions

Figure 2 Body weight (A) and heart weight (B). There was no significant difference in body weight between groups either at the time of initial or terminal surgery. The mean change in body weight over 4 weeks (δ) is shown in (A). There was no significant difference in left ventricular weight at the time of initial or terminal surgery (B). LV, Left ventricular. The Journal of Thoracic and Cardiovascular Surgery 2017 153, 934-943DOI: (10.1016/j.jtcvs.2016.10.077) Copyright © 2016 The American Association for Thoracic Surgery Terms and Conditions

Figure 3 Creatine kinase-MB isoenzyme (CK-MB) (A) and cardiac troponin I (cTnI) (B) at day 1 and day 3 after surgery in the vehicle-only and mitochondria groups. Both CK-MB and cTnI were significantly decreased (P < .001) in mitochondria hearts compared with vehicle hearts at day 3. There was no significant difference between the 2 groups at day 1. Results are shown for n = 6 in each group. Significant differences compared with vehicle are shown. ∗Significant differences compared with vehicle at P = .006. The Journal of Thoracic and Cardiovascular Surgery 2017 153, 934-943DOI: (10.1016/j.jtcvs.2016.10.077) Copyright © 2016 The American Association for Thoracic Surgery Terms and Conditions

Figure 4 A and B, Area at risk (AAR) (A) and infarct size (%AAR) (B) for vehicle and mitochondria. Results are shown as the mean ± standard deviation for n = 6 in each group. Significant differences compared with vehicle are shown. C and D, Histochemical and electron microscopy analysis of myocardial tissue at 4 weeks of recovery. Representative Masson's trichrome-stained (C) and transmission electron (D) micrographs from vehicle and mitochondria hearts are shown. Longitudinal and transverse interfibrillar separation was observed in vehicle hearts but not in mitochondria hearts (C), but there was no difference in collagen between the 2 groups (C). Electron microscopy analysis (D) shows mitochondrial damage and contraction bands in vehicle hearts that are not present in mitochondria hearts. (Scale bars: 500 μm.) LV, Left ventricular. The Journal of Thoracic and Cardiovascular Surgery 2017 153, 934-943DOI: (10.1016/j.jtcvs.2016.10.077) Copyright © 2016 The American Association for Thoracic Surgery Terms and Conditions

Figure 5 A, Electrophysiological data. Representative electrocardiograms at baseline (before surgery), ischemia (following 10 minutes of regional ischemia), and reperfusion (following 120 minutes of reperfusion). B, Echocardiography data. A, Recommended 17-segment model for the left ventricle.14 B, Global and regional circumferential strain analysis from vehicle and mitochondria hearts. Strain was measured in the midventricular short-axis plane (segments 7-12). More negative strain values indicate a more favorable strain. Values are shown as mean ± standard deviation. The Journal of Thoracic and Cardiovascular Surgery 2017 153, 934-943DOI: (10.1016/j.jtcvs.2016.10.077) Copyright © 2016 The American Association for Thoracic Surgery Terms and Conditions

Figure 6 Mitochondrial uptake at 4 weeks after injection. A and B, T2*-weighted magnetic resonance imaging of mitochondrial uptake in pig heart at 4 weeks after injection. Iron oxide–labeled mitochondria appear as dark regions in the heart (arrows). Coronal (left) and transverse slices (right) are depicted from 2 different regions of the same heart. C, Histological sections from the dark regions were stained for iron (left) and human mitochondria (right). Our results show that mitochondria are present for at least 4 weeks after injection in the pig heart. The Journal of Thoracic and Cardiovascular Surgery 2017 153, 934-943DOI: (10.1016/j.jtcvs.2016.10.077) Copyright © 2016 The American Association for Thoracic Surgery Terms and Conditions

Autologous mitochondrial transplantation decreases infarct size and preserves structure. The Journal of Thoracic and Cardiovascular Surgery 2017 153, 934-943DOI: (10.1016/j.jtcvs.2016.10.077) Copyright © 2016 The American Association for Thoracic Surgery Terms and Conditions

The Journal of Thoracic and Cardiovascular Surgery 2017 153, 934-943DOI: (10.1016/j.jtcvs.2016.10.077) Copyright © 2016 The American Association for Thoracic Surgery Terms and Conditions