Transplantation of Mitochondria isolated from iPSC-MSCs mitigates doxorubicin-induced cardiomyopathy by inhibiting cardiomyocyte senescence.

Mitochondria isolated from iPSC-MSCs exhibited intact mitochondrial morphology and quality as confirmed by characterization methods.

• Mitochondria were isolated using ultracentrifugation from induced pluripotent stem cell-derived mesenchymal stem cells (iPSC-MSCs).
• Characterization was performed by transmission electron microscopy and Western blotting.
• The isolated iPSC-MSC-Mito demonstrated intact mitochondrial morphology and quality suitable for transplantation.

iPSC-MSC-Mito were successfully internalized by neonatal mouse cardiomyocytes (NMCMs) under doxorubicin challenge in vitro.

• The cellular model used neonatal mouse cardiomyocytes (NMCMs) exposed to doxorubicin (DOX).
• Internalization of exogenous iPSC-MSC-Mito by DOX-challenged NMCMs was demonstrated in vitro.
• This confirmed the feasibility of mitochondrial transfer into cardiomyocytes under pathological conditions.

Administration of iPSC-MSC-Mito improved the respiratory capacity of cardiomyocytes under doxorubicin challenge.

• Mitochondrial function was evaluated using the Seahorse assay in DOX-treated NMCMs exposed to different treatments.
• iPSC-MSC-Mito treatment improved respiratory capacity of DOX-challenged cardiomyocytes.
• The improvement in respiratory capacity was associated with downregulated lactate levels in treated cardiomyocytes.

iPSC-MSC-Mito inhibited cardiomyocyte senescence in vitro through reduction of lactate levels, and this effect was partially abrogated by exogenous lactate.

• Cellular senescence of NMCMs was examined by senescence-associated-β-galactosidase (SA-β-gal) assay.
• Administration of iPSC-MSC-Mito led to inhibition of cardiomyocyte senescence, mediated through downregulated lactate levels.
• The anti-senescence effect of iPSC-MSC-Mito was partially abrogated by addition of exogenous lactate, confirming the role of lactate in this pathway.

Lactate regulated SIRT2 expression by binding to the ARG97 and HIS187 residues in the PH domain of SIRT2.

• The mechanism was identified using molecular docking and site-directed mutation assays.
• Lactate was found to bind specifically to ARG97 and HIS187 residues located in the PH domain of SIRT2.
• This binding interaction regulated SIRT2 expression, linking metabolic changes (lactate levels) to the senescence pathway.

In vivo transplantation of iPSC-MSC-Mito attenuated doxorubicin-induced cardiomyopathy, improving cardiac function and decreasing cardiac fibrosis and cardiomyocyte senescence.

• A mouse model of DIC was induced by intraperitoneal injection of DOX, followed by intraperitoneal injection of iPSC-MSC-Mito.
• Cardiac function, fibrosis, and cardiomyocyte senescence were examined in each group.
• Transplantation of iPSC-MSC-Mito resulted in improved cardiac function, decreased cardiac fibrosis, and reduced cardiomyocyte senescence in vivo.