Original Investigation
Mitochondria-Rich Extracellular Vesicles From Autologous Stem Cell–Derived Cardiomyocytes Restore Energetics of Ischemic Myocardium

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Abstract

Background

Mitochondrial dysfunction results in an imbalance between energy supply and demand in a failing heart. An innovative therapy that targets the intracellular bioenergetics directly through mitochondria transfer may be necessary.

Objectives

The purpose of this study was to establish a preclinical proof-of-concept that extracellular vesicle (EV)-mediated transfer of autologous mitochondria and their related energy source enhance cardiac function through restoration of myocardial bioenergetics.

Methods

Human-induced pluripotent stem cell–derived cardiomyocytes (iCMs) were employed. iCM-conditioned medium was ultracentrifuged to collect mitochondria-rich EVs (M-EVs). Therapeutic effects of M-EVs were investigated using in vivo murine myocardial infarction (MI) model.

Results

Electron microscopy revealed healthy-shaped mitochondria inside M-EVs. Confocal microscopy showed that M-EV–derived mitochondria were transferred into the recipient iCMs and fused with their endogenous mitochondrial networks. Treatment with 1.0 × 108/ml M-EVs significantly restored the intracellular adenosine triphosphate production and improved contractile profiles of hypoxia-injured iCMs as early as 3 h after treatment. In contrast, isolated mitochondria that contained 300× more mitochondrial proteins than 1.0 × 108/ml M-EVs showed no effect after 24 h. M-EVs contained mitochondrial biogenesis-related messenger ribonucleic acids, including proliferator-activated receptor γ coactivator-1α, which on transfer activated mitochondrial biogenesis in the recipient iCMs at 24 h after treatment. Finally, intramyocardial injection of 1.0 × 108 M-EVs demonstrated significantly improved post-MI cardiac function through restoration of bioenergetics and mitochondrial biogenesis.

Conclusions

M-EVs facilitated immediate transfer of their mitochondrial and nonmitochondrial cargos, contributing to improved intracellular energetics in vitro. Intramyocardial injection of M-EVs enhanced post-MI cardiac function in vivo. This therapy can be developed as a novel, precision therapeutic for mitochondria-related diseases including heart failure.

Key Words

bioenergetics
heart failure
human stem cells
mitochondria
myocardial infarction

Abbreviations and Acronyms

ATP
adenosine triphosphate
CMR
cardiac magnetic resonance
COX
cytochrome c oxidase
D15s
induced pluripotent stem cell–derived cardiomyocytes cultured for shorter than 15 days
D50s
induced pluripotent stem cell–derived cardiomyocytes cultured for longer than 50 days
ERR
estrogen-related receptor
ETC
electron transport chain
EV
extracellular vesicle
FCM
flow cytometry
HF
heart failure
iCM
induced pluripotent stem cell–derived cardiomyocyte
iPSC
induced pluripotent stem cell
Ld-EV
large vesicle-depleted extracellular vesicle
LV
left ventricular
M-EV
mitochondria-rich extracellular vesicle
MI
myocardial infarction
mRNA
messenger ribonucleic acid
mtDNA
mitochondrial deoxyribonucleic acid
PBS
phosphate-buffered saline
PGC
proliferator-activated receptor γ coactivator
RFP
red fluorescent protein

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