Mitochondrial transplantation enhances murine lung viability and recovery after ischemia-reperfusion injury.

The most common cause of acute lung injury is ischemia-reperfusion injury (IRI), during which mitochondrial damage occurs. We have previously demonstrated that mitochondrial transplantation is an efficacious therapy to replace or augment mitochondria damaged by IRI, allowing for enhanced muscle viability and function in cardiac tissue. Here, we investigate the efficacy of mitochondrial transplantation in a murine lung IRI model using male C57BL/6J mice. Transient ischemia was induced by applying a microvascular clamp on the left hilum for 2 h. Upon reperfusion mice received either vehicle or vehicle-containing mitochondria either by vascular delivery (Mito V) through the pulmonary artery or by aerosol delivery (Mito Neb) via the trachea (nebulization). Sham control mice underwent thoracotomy without hilar clamping and were ventilated for 2 h before returning to the cage. After 24 h recovery, lung mechanics were assessed and lungs were collected for analysis. Our results demonstrated that at 24 h of reperfusion, dynamic compliance and inspiratory capacity were significantly increased and resistance, tissue damping, elastance, and peak inspiratory pressure (Mito V only) were significantly decreased (P < 0.05) in Mito groups as compared with their respective vehicle groups. Neutrophil infiltration, interstitial edema, and apoptosis were significantly decreased (P < 0.05) in Mito groups as compared with vehicles. No significant differences in cytokines and chemokines between groups were shown. All lung mechanics results in Mito groups except peak inspiratory pressure in Mito Neb showed no significant differences (P > 0.05) as compared with Sham. These results conclude that mitochondrial transplantation by vascular delivery or nebulization improves lung mechanics and decreases lung tissue injury.

Mitochondrial transplantation ameliorates acute limb ischemia.

OBJECTIVE: Acute limb ischemia (ALI), the most challenging form of ischemia-reperfusion injury (IRI) in skeletal muscle tissue, leads to decreased skeletal muscle viability and limb function. Mitochondrial injury has been shown to play a key role in skeletal muscle IRI. In previous studies, we have demonstrated that mitochondrial transplantation (MT) is an efficacious therapeutic strategy to replace or to augment mitochondria damaged by IRI, allowing enhanced muscle viability and function in cardiac tissue. In this study, we investigated the efficacy of MT in a murine ALI model. METHODS: C57BL/6J mice (male, 10-12 weeks) were used in a model of ALI. Ischemia was induced by applying a tourniquet on the left hindlimb. After 2 hours of ischemia, the tourniquet was released, and reperfusion of the hindlimb was re-established; either vehicle alone (n = 15) or vehicle containing mitochondria (n = 33) was injected directly into all the muscles of the hindlimb. Mitochondria were delivered at concentrations of 1 × 106 to 1 × 109 per gram wet weight to each muscle, and the animals were allowed to recover. Sham mice received no ischemia or injections but were anesthetized for 2 hours and allowed to recover. After 24 hours of recovery, limb function was assessed by DigiGait (Mouse Specifics Inc, Boston, Mass), and animals were euthanized; the gastrocnemius, soleus, and vastus medialis muscles were collected for analysis. RESULTS: After 24 hours of hindlimb reperfusion, infarct size (percentage of total mass) and apoptosis were significantly decreased (P < .001, each) in the gastrocnemius, soleus, and vastus medialis muscles in MT mice compared with vehicle mice for all mitochondrial concentrations (1 × 106 to 1 × 109 per gram wet weight). DigiGait analysis at 24 hours of reperfusion showed that percentage shared stance time was significantly increased (P < .001) and stance factor was significantly decreased (P = .001) in vehicle compared with MT and sham mice. No significant differences in percentage shared stance time (P = .160) or stance factor (P = .545) were observed between MT and sham mice. CONCLUSIONS: MT ameliorates skeletal muscle injury and enhances hindlimb function in the murine model of ALI.

Preischemic autologous mitochondrial transplantation by intracoronary injection for myocardial protection.

OBJECTIVE: To investigate preischemic intracoronary autologous mitochondrial transplantation (MT) as a therapeutic strategy for prophylactic myocardial protection in a porcine model of regional ischemia-reperfusion injury (IRI). METHODS: The left coronary artery was cannulated in Yorkshire pigs (n = 26). Mitochondria (1 × 109) or buffer (vehicle [Veh]) were delivered as a single bolus (MTS) or serially (10 injections over 60 minutes; MTSS). At 15 minutes after injection, the heart was subjected to temporary regional ischemia (RI) by snaring the left anterior descending artery. After 30 minutes of RI, the snare was released, and the heart was reperfused for 120 minutes. RESULTS: Coronary blood flow (CBF) and myocardial function were increased temporarily during the pre-RI period. Following 30 minutes of RI, MTS and MTSS hearts had significantly increased CBF that persisted throughout reperfusion (Veh vs MTS and MTSS; P = .04). MTS and MTSS showed a significantly enhanced ejection fraction (Veh vs MTS, P < .001; Veh vs MTSS, P = .04) and developed pressure (Veh vs MTS, P < .001; Veh vs MTSS, P = .03). Regional function, assessed through segmental shortening (Veh vs MTS, P = .03; Veh vs MTSS, P < .001), fractional shortening (Veh vs MTS, P < .001; Veh vs MTSS, P = .04), and strain analysis (Veh vs MTS, P = .002; Veh vs MTSS, P = .003), was also significantly improved. Although there was no difference in the area at risk between treatment groups, infarct size was significantly reduced in both MT groups (Veh vs MTS and MTSS, P < .001). CONCLUSIONS: Preischemic MT by single or serial intracoronary injections provides prophylactic myocardial protection from IRI, significantly decreasing infarct size and enhancing global and regional function.

Alloreactivity and allorecognition of syngeneic and allogeneic mitochondria.

Previously, we have demonstrated that the transplantation of autologous mitochondria is cardioprotective. No immune or autoimmune response was detectable following the single injection of autologous mitochondria. To expand the therapeutic potential and safety of mitochondrial transplantation, we now investigate the immune response to single and serial injections of syngeneic and allogeneic mitochondria delivered by intraperitoneal injection. Our results demonstrate that there is no direct or indirect, acute or chronic alloreactivity, allorecognition or damage-associated molecular pattern molecules (DAMPs) reaction to single or serial injections of either syngeneic or allogeneic mitochondria.

Mitochondrial transplantation prolongs cold ischemia time in murine heart transplantation.

BACKGROUND: Cold ischemia time (CIT) causes ischemia‒reperfusion injury to the mitochondria and detrimentally effects myocardial function and tissue viability. Mitochondrial transplantation replaces damaged mitochondria and enhances myocardial function and tissue viability. Herein we investigated the efficacy of mitochondrial transplantation in enhancing graft function and viability after prolonged CIT. METHODS: Heterotopic heart transplantation was performed in C57BL/6J mice. Upon heart harvesting from C57BL/6J donors, 0.5 ml of either mitochondria (1 × 108 in respiration buffer; mitochondria group) or respiration buffer (vehicle group) was delivered antegrade to the coronary arteries via injection to the coronary ostium. The hearts were excised and preserved for 29 ± 0.3 hours in cold saline (4°C). The hearts were then heterotopically transplanted. A second injection of either mitochondria (1 × 108) or respiration buffer (vehicle) was delivered antegrade to the coronary arteries 5 minutes after transplantation. Grafts were analyzed for 24 hours. Beating score, graft function, and tissue injury were measured. RESULTS: Beating score, calculated ejection fraction, and shortening fraction were significantly enhanced (p < 0.05), whereas necrosis and neutrophil infiltration were significantly decreased (p < 0.05) in the mitochondria group as compared with the vehicle group at 24 hours of reperfusion. Transmission electron microscopy showed the presence of contraction bands in vehicle but not in mitochondria grafts. CONCLUSIONS: Mitochondrial transplantation prolongs CIT to 29 hours in the murine heart transplantation model, significantly enhances graft function, and decreases graft tissue injury. Mitochondrial transplantation may provide a means to reduce graft failure and improve transplantation outcomes after prolonged CIT.

A Novel Biological Strategy for Myocardial Protection by Intracoronary Delivery of Mitochondria: Safety and Efficacy.

Mitochondrial dysfunction is the determinant insult of ischemia-reperfusion injury. Autologous mitochondrial transplantation involves supplying one's healthy mitochondria to the ischemic region harboring damaged mitochondria. The authors used in vivo swine to show that mitochondrial transplantation in the heart by intracoronary delivery is safe, with specific distribution to the heart, and results in significant increase in coronary blood flow, which requires intact mitochondrial viability, adenosine triphosphate production, and, in part, the activation of vascular KIR channels. Intracoronary mitochondrial delivery after temporary regional ischemia significantly improved myocardial function, perfusion, and infarct size. The authors concluded that intracoronary delivery of mitochondria is safe and efficacious therapy for myocardial ischemia-reperfusion injury.