# An Exploratory Study of Kinase Activation Profiles in Hypoxic Human Cardiomyocytes Treated with Protective Extracellular Vesicles

## TL;DR

This study explores how different types of extracellular vesicles affect kinase activity in heart cells under low oxygen conditions, revealing distinct signaling patterns despite similar protective effects.

## Contribution

The study reveals distinct kinase activation profiles induced by different extracellular vesicle types in hypoxic cardiomyocytes, despite equivalent cardioprotection.

## Key findings

- All EV types significantly reduced LDH release compared to hypoxia alone.
- Each EV type induced unique kinase activation patterns, with S-EVs and PC-EVs increasing phosphorylation while CSC-EVs and MSC-EVs decreased it.
- AKT family kinases were implicated in all treatments, but phosphorylation of AKT1/2/3 at Ser473 was unchanged.

## Abstract

Myocardial infarction (MI) causes hypoxic cardiomyocyte death, and extracellular vesicles (EVs) offer therapeutic potential. This study aimed to compare kinase activation profiles induced by four human-derived EV types-serum-derived (S-EVs), platelet concentrate-derived (PC-EVs), cardiac stromal cell-derived (CSC-EVs), and bone marrow mesenchymal stromal cell-derived (MSC-EVs)- in hypoxic metabolically matured human iPSC-derived cardiomyocytes (iCMs).

Metabolically matured human induced pluripotent stem cell-derived cardiomyocytes (iCMs) were exposed to 12-hour hypoxia ± standardized doses of EVs. Kinase activity was profiled using the PamStation platform, and bioinformatic tools (KRSA, UKA, PTM-SEA, KEA3) identified differentially activated kinases. AKT phosphorylation (Ser473) was measured by ELISA.

All EVs significantly reduced LDH release versus hypoxia alone (P ≤ 0.0001), with no inter-group differences. Hypoxia globally suppressed kinase activity, while each EV type induced distinct patterns: S-EVs and PC-EVs increased total phosphorylation, whereas CSC-EVs and MSC-EVs further decreased it. Bioinformatics implicated the AKT family in all treatments, but ELISA revealed no change in AKT1/2/3 phosphorylation at Ser473 versus hypoxia controls.

Despite equivalent cardioprotection, each EV type elicited unique kinase activation profiles, suggesting distinct signaling mechanisms. Kinase activity was not a predictor of protection at the measured time point, highlighting the complexity of EV-mediated pathways.