A, Confocal microscopy image that demonstrates the mitochondrial localization (TOM20) of the Flag tag in dox-treated, in vitro-differentiated adipocytes from adipo-mitoFlag mice. B and C, mitoFlag detection by gene expression (B) and immunoprecipitation (C) in heart or sWAT tissue from adipo-mitoFlag mice fed dox-HFD for 11 weeks. D, Confocal microscopy images demonstrating the localization of the Flag tag in cardiomyocytes (CTN1 positive) with no primary antibody control and no mitoFlag transgene control. Arrow specifies non-specific signal and asterisks are supplied for orientation. E, Immunoprecipitated Flag tag in isolated cardiac mitochondria or whole sWAT from the adipo-mitoFlag mice on dox-HFD for 11 weeks. F, sEVs from in vitro differentiated adipocytes expressing mitoFlag and treated with palmitate were injected into WT mice. Where indicated, mice were injected with chloroquine (CQ) 3 hours prior to sEV injection. The Flag tag was immunoprecipitated from heart tissue 1 hour following sEV injection. G, sEVs from primary human (Hu) adipocytes treated with palmitate were injected into wild type mice with or without a CQ pre-injection. Human mtDNA was quantified in mouse heart tissue 1 hour following sEV injection and extensive perfusion with PBS. Data are presented as mean ± s.e.m. *P < 0.05, ** P < 0.01, *** P < 0.001. See also Figure S7.