Ambient fine particulate matter (PM2.5) is a leading global environmental risk factor for cardiovascular disease. Although PM2.5 exposure is known to lead to mitochondrial dysfunction, the precise metabolic mechanisms underlying this effect remain incompletely understood. In this study, we investigated the role of the mitochondrial deacetylase Sirt3 in PM2.5-induced cardiotoxicity using human induced pluripotent stem cell-derived cardiac organoids (hiPSC-COs), a physiologically relevant multicellular model. Our results demonstrated that PM2.5 exposure compromised the contractile function of COs. Specifically, it induced the expression of myocardial injury markers and triggered a series of mitochondrial impairments, including structural damage, inflammatory response, oxidative stress and loss of membrane potential. Mechanistically, exposure of COs to PM2.5 suppressed Sirt3 expression, leading to increased acetylation and decreased activity of malate dehydrogenase 2 (Mdh2), a key enzyme in the tricarboxylic acid (TCA) cycle. This disruption resulted in reduced levels of TCA cycle metabolites and a severe ATP deficit. Pharmacological activation of Sirt3 with dihydromyricetin attenuated the PM2.5-induced mitochondrial dysfunction, restored ATP levels and improved contractility. Our findings elucidate that PM2.5 impairs cardiac energy metabolism via the Sirt3/Mdh2 pathway, identifying Sirt3 as a potential therapeutic target for mitigating PM2.5-related cardiovascular dysfunction.