Figure 6. Ch25h-deficient and cholesterol loaded macrophages have impaired mitochondrial metabolism.

(A) (Left) Representative histogram of Mitotracker Deep Red staining on Ch25h^+/− and Ch25h^−/− BMDMs stimulated with LPS for 8 hr. (Right) Summary mean fluorescence intensity (MFI) data from 3 experiments (mean +/− SD). Representative of 8–10 independent experiments. (B) MFI of Mitotracker Deep Red staining on Ch25h^−/− BMDMs transduced with MSCV-vector, −Ch25h, or −Ch25hmut. Data from 4 independent experiments (mean+/−SD). (C) (Left) Representative histogram of Mitotracker Green staining on Ch25h^+/− and Ch25h^−/− BMDMs stimulated as above. (Right) Summary MFI data from 3 experiments (mean+/−SD). Representative of 8–10 independent experiments (mean +/− SD). (D) MFI of Mitotracker Green staining on Ch25h^−/− BMDMs transduced with MSCV-vector, −Ch25h, or −Ch25hmut. Data from 4 independent experiments (mean+/−SD). (E) Representative histogram of TMRM staining on Ch25h^+/− and Ch25h^−/− BMDMs stimulated with LPS for 8 hr. Graph shows summary MFI data from 3 experiments (mean+/−SD). (F) MFI of TMRM staining on Ch25h^−/− BMDMs transduced with MSCV-vector, −Ch25h, or −Ch25hmut. Data from 2 independent experiments (mean+/−SD). (G) Seahorse analysis of oxygen consumption rate (OCR) over time from BMDMs pretreated for 8 hr with LPS and after the indicated additional treatments. Data are representative of 3 independent experiments (mean+/−SD). (H) Seahorse analysis of basal OCR and extracellular acidification rate (ECAR) of Ch25h^+/− and Ch25h^−/− BMDMs stimulated with LPS for 8 hr. Data from 3 independent experiments (mean+/−SD). (I, J) Mitotracker Deep Red (I) and Mitotracker Green (J) staining of Ch25h^+/− and Ch25h^−/− peritoneal macrophages 8 hr after I.P. saline or LPS injection. Data from 3 independent (n=6–7 mice per group). See also Figure S4.