Fig. 4: Lactate production is required for epigenetic and functional polarization.
a, Extracellular lactate production from BMDMs stimulated with control vehicle (Ctrl) or FGK45 mAb in the absence or presence of rotenone or oligomycin under glucose-deplete media. b, ECAR measurement (left) and the average of etomoxir (Eto)-sensitive ECAR (right) in BMDMs stimulated with either Ctrl or FGK45 mAb under glucose-deplete media. c, Extracellular lactate production from Cas9-expressing BMDMs harboring control sgRNA (Ctrl) and sgRNAs targeting Cpt1a (Cpt1a sgRNA-1 and sgRNA-2) under glucose-deplete media. d, qPCR analysis of mRNA expression of pro-inflammatory marker genes in BMDMs harboring control sgRNA (Ctrl) and sgRNAs targeting LdhA (LDHA sgRNA-1 and sgRNA-2) under glucose-deplete media. e,f, Tumor growth (e) and tumor weight measurement (f) in tumor-bearing WT or LdhAcKO mice treated with PBS or FGK45 mAb. g, qPCR analysis of mRNA expression of pro-inflammatory marker genes in TAM isolated from tumor-bearing WT or LdhAcKO mice. h, Representative immunoblots of indicated proteins in lysates of Cas9-expressing BMDMs harboring indicated sgRNAs. i, Chromatin immunoprecipitation (ChIP) analysis of H3K27ac in Cas9-expressing BMDMs harboring indicated sgRNAs. Data were normalized to input. j, The concentration of indicated metabolites in Cas9-expressing BMDMs harboring indicated sgRNAs upon FGK45 stimulation for indicated periods. k, qPCR analysis of mRNA expression of pro-inflammatory marker genes in Cas9-expressing BMDMs harboring indicated sgRNAs in the absence or presence of sodium acetate (Ace, 5 mM) under glucose-deplete media. l,m, Representative immunoblots of indicated proteins (l) and qPCR analysis of mRNA expression of pro-inflammatory marker genes (m) in of Cas9-expressing BMDMs harboring indicated sgRNAs upon FGK45 mAb under glucose-deplete media with or without acetate (ace). a–d,l,m, Data are representative of three individual experiments with n = 3 per group. e–g, Representative data or pooled data from two individual experiments with n = 4 mice per group in e; n = 6 for WT-PBS, n = 8 for WT-FGK45, n = 5 for LDHAcKO-PBS and n = 8 for LDHAcKO-PBS in f; n = 4 for WT-PBS, n = 6 for WT-FGK45, n = 3 for LDHAcKO-PBS and n = 5 for LDHAcKO-PBS in g. h–k, Data are representative of three individual experiments (n = 3 per group). All data are the mean ± s.d. and analyzed by a two-tailed Student’s t-test. NS, not significant.
