Figure 2.

Intervenolin (ITV) inhibits mitochondrial complex I activity and induces metabolic shift in cancer cells

(A) Results of seahorse flux analyzer assays showing the OCR of MKN-74 cells. Assay buffer (Ctrl) or the indicated concentrations of ITV were introduced at 24 min; FCCP, 64 min; and rotenone (ROT) and antimycin A (AA), 88 min.

(B) Activity of the mitochondrial complex I as measured using an NADH assay. Reduction rate of NADH that normalized by non-treated value was shown as complex I activity. The activity of mitochondria without compounds was defined as 100%.

(C) Seahorse flux analyzer assay results showing the extracellular acidification rate (ECAR) in MKN-74 cells as in (A).

(D) Energy map of MKN-74 cells based on OCR and ECAR values. The status of the metabolic pathways is classified as aerobic, glycolytic, energetic, or quiescent.

(E–G). Intracellular metabolite concentrations were determined using mass spectrometric analysis (E and F). Relative amounts of the indicated metabolites in MKN-74 cells treated by 1 μg/mL ITV, 1 μg/mL AS-1936 (AS), or 0.01 μg/mL rotenone (ROT) for 3 h are shown. B.D., below detection limit (n = 3; ∗p < 0.05 versus Ctrl, ∗∗p < 0.005 versus Ctrl).

(G) Relative amounts of the indicated metabolites in MKN-74 cells treated by ITV for 3 h are shown (n = 3; ∗p < 0.05 versus non-treated cells). Data are presented as the mean ± SD (n = 3) and normalized according to the baseline and analyzed using two-sided Student's t -test.