Mitochondrial Function Evaluation in 3T3-L1 Cells

Mitochondrial function was assessed and analysed in live cells by Seahorse XF Cell Mito Stress Test Kit (Agilent Technologies #103015), according to the manufacturer’s protocol. Briefly, 2×10⁴ cells/well of 3T3-L1 cells were plated on a Seahorse XF24 cell culture microplate and transfected with siHif3α. At 24 h, the medium was replaced with differentiated medium in presence of pro-inflammatory cytokines. After 72 h, the plate was loaded into the by Seahorse XF24 Analyser (Agilent Technologies) and analysed as reported in (Carafa et al., 2020 (link)). Multiple parameters are evaluated including basal respiration, ATP-linked respiration, maximal and reserve capacities, and non-mitochondrial respiration.