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. 2020 Nov 20;218(2):e20200675. doi: 10.1084/jem.20200675

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© 2020 Richard et al.

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Figure S4. — Metabolic analysis of WT and TLR4-SNP macrophages. (A) Diagram of glycolytic stress test measurements (reproduced with permission; courtesy of Agilent Technologies). (B) Glycolytic capacity ([ECAR after inhibition of mitochondrial ATP synthase with oligomycin forcing all cellular energy production to the glycolytic pathway] − [ECAR after inhibition of glycolysis with excess 2-deoxyglucose]) from the same experiments as in Fig. 4 B (combined data from nine separate experiments). Data were analyzed by two-way ANOVA (α = 0.10), with Sidak’s multiple comparison post-tests to compare WT and TLR4-SNP responses as follows: untreated, not significant (N.S.); 1 ng/ml LPS, N.S.; 100 ng/ml LPS, N.S.; WT and TLR4^−/− responses: untreated, N.S.; 1 ng/ml LPS, ***, P = 0.0001; 100 ng/ml LPS, ***, P = 0.0011; TLR4-SNP and TLR4^−/− responses: 100 ng/ml LPS, **, P = 0.010 (adjusted for multiple comparisons). (C) Diagram of mitochondrial stress test measurements (reproduced with permission; courtesy of Agilent Technologies). (D) ATP production ([OCR before any injection] − [OCR after injecting ATP synthase inhibitor oligomycin]) from the same experiments as in Fig. 4 (E and F; combined data from seven separate experiments). Data were analyzed by two-way ANOVA (α = 0.10), with Sidak’s multiple comparison post-tests to compare WT and TLR4-SNP responses as follows: untreated, N.S.; 1 ng/ml LPS, N.S.; 100 ng/ml LPS, N.S.; WT and TLR4^−/− responses: untreated, N.S.; 1 ng/ml LPS, **, P = 0.0067; 100 ng/ml LPS, **, P = 0.0023; TLR4-SNP and TLR4^−/− responses at 100 ng/ml LPS, N.S. (adjusted for multiple comparisons). (E) Proton leak ([OCR after ATP synthase inhibition by oligomycin] − [OCR after electron transport inhibition by AA]) from the same experiments as in Fig. 4 (E and F). Data were analyzed by two-way ANOVA (α = 0.10), with Sidak’s multiple comparison post-tests to compare WT vs. TLR4-SNP responses as follows: untreated, N.S.; 1 ng/ml LPS, **, P = 0.014; 100 ng/ml LPS, N.S.; WT vs. TLR4^−/− responses: untreated, N.S.; 1 ng/ml LPS, ***, P = 0.0002; 100 ng/ml LPS, ***, P < 0.0001; TLR4-SNP vs. TLR4^−/− responses at 100 ng/ml LPS, ***, P < 0.0001. (adjusted for multiple comparisons). (F) Mitochondrial coupling efficiency (100 × [ATP production rate]/[basal respiration rate]) from the same experiments as in Fig. 4 (E and F) (combined data from seven separate experiments). Data were analyzed by two-way ANOVA (α = 0.10), with Sidak’s multiple comparison post-tests to compare WT vs. TLR4-SNP responses: untreated, N.S.; 1 ng/ml LPS, N.S.; 100 ng/ml LPS, N.S.; WT vs. TLR4^−/− responses: untreated, N.S.; 1 ng/ml LPS, **, P = 0.0025; 100 ng/ml LPS,***, P < 0.0001; TLR4-SNP vs. TLR4^−/− responses at 100 ng/ml LPS, ***, P < 0.0001 (adjusted for multiple comparisons).