(TKP2021-EGA-23) Támogató: Innovációs és Technológiai Minisztérium
(RRF-2.3.1-21-2022-00003)
(2019-2.1.7-ERA-NET-2021-00015)
Cardiomyopathies are leading causes of human mortality. Recent data indicate that
the cardiomyocyte-derived extracellular vesicles (EVs) released upon cardiac injury
are present in circulation. This paper aimed to analyze EVs released under normal
and hypoxic conditions by H9c2 (rat), AC16 (human) and HL1 (mouse) cardiac cell lines.
Small (sEVs), medium (mEVs) and large EVs (lEVs) were separated from a conditioned
medium by a combination of gravity filtration, differential centrifugation and tangential
flow filtration. The EVs were characterized by microBCA, SPV lipid assay, nanoparticle
tracking analysis, transmission and immunogold electron microscopy, flow cytometry
and Western blotting. Proteomic profiles of the EVs were determined. Surprisingly,
an endoplasmic reticulum chaperone, endoplasmin (ENPL, grp94 or gp96), was identified
in the EV samples, and its association with EVs was validated. The secretion and uptake
of ENPL was followed by confocal microscopy using GFP-ENPL fusion protein expressing
HL1 cells. We identified ENPL as an internal cargo of cardiomyocyte-derived mEVs and
sEVs. Based on our proteomic analysis, its presence in EVs was linked to hypoxia in
HL1 and H9c2 cells, and we hypothesize that EV-associated ENPL may have a cardioprotective
role by reducing cardiomyocyte ER stress.
