Fig 2: A model of de novo beige fat biogenesisCold, ß3-AR agonist, and burn injury trigger WAT lipolysis and the release of free fatty acids. CD81+ beige progenitor cells actively uptake linoleic acid through the plasma membrane fatty acid transporter CD36 for mitochondrial ß-oxidation and also for the synthesis of arachidonic-acid-derived metabolites, including PGD2 via the Cox2 pathway. PGD2 stimulates CD81+ cell proliferation by paracrine or autocrine manner via the receptor DP1.

Fig 3: CD36 is required for CD81+ APC proliferation(A) Relative expression levels of indicated genes enriched in inguinal WAT-derived CD81+ cells relative to CD81- cells.(B) Relative mRNA levels of Cd36 in CD81+ cells and CD81- cells. n = 4.(C) Relative Cd36 protein expression in CD81+ cells and CD81- cells. n = 4.(D) Schematic illustration of the experiments.(E) Relative mRNA levels of Cd36 in the inguinal WAT-derived CD81+ cells. n = 4.(F) Representative FACS histogram of Ki67+ CD81+ cells in (D).(G) Quantification of Ki67+ CD81+ cells in (F). n = 6.(H) Representative H&E staining in the inguinal WAT of mice in (D). LN, lymph node. Scale bars, 100 µm.(I) Relative mRNA levels of indicated genes in the inguinal WAT. n = 6.(J) The number of primary CD81+ cells treated with vehicle or linoleic acid for 4 days. n = 4. ***p < 0.001 by one-way ANOVA followed by the Tukey-Kramer’s post hoc test.(K) Quantitative analysis of Ki67+ CD81+ cells in the inguinal WAT following linoleic acid supplementation at 30°C. n = 6.(L) Relative mRNA levels of indicated genes in the inguinal WAT following linoleic acid supplementation and cold exposure. n = 6.(M) Representative H&E staining in the inguinal WAT. Scale bars, 100 µm.(B, C, E, G, I, K, and L) *p < 0.05, **p < 0.01, ****p < 0.0001, by two-tailed unpaired Student’s t test. ns, not significant.

Fig 4: Lactate promotes the release of extracellular FFAs from TG via lactyl‐proteins. A) Global lactylome profiling showing the numbers of lactylated‐proteins in various pathways, as identified by KEGG map analysis in tumors. B) Top 20 intracellular and extracellular small molecule metabolites in tumor cells showing significant changes (p < 0.05) after 14 h lactate treatment, as analyzed by LC‐MS/MS. C) Change in intracellular and extracellular 19‐methyl‐heneicosanoic acid in tumor cells after lactate treatment. D–K) Analysis of intracellular or extracellular FFAs in the indicated tumor cells. Data are presented as mean ± SD from n ≥ 3 independent experiments, *p < 0.05, **p < 0.01, Student's t‐test. D) Analysis of FFAs in H1299 tumor cells after 14 h treatment with 20 mM lactate. (E and F) Analysis of intracellular or extracellular FFAs and oil red staining in HepG2 cells treated with 30 mM lactate for 24 h. Representative images show cells stained with oil red. G) H1299 cells were infected with either control shRNA, CD36‐Sh1, or Sh2 lentiviruses for 72 h, followed by another 14 h lactate treatment. CD36 knockdown was confirmed via Western blotting. H and I) Intracellular lactate promotes the increase of extracellular FFAs. H1299 cells were infected with control shRNA, LDHA‐Sh1, LDHA‐Sh2, MCT4‐Sh1, or MCT4‐Sh2 lentiviruses for 72 h, or treated with specific inhibitors for 12 h, followed by a 14 h pyruvic acid (5 mM) treatment. Extracellular FFAs were analyzed in the indicated cells. J and K) H1299 cells (J) and A549 cells (K) were treated with lactate (20 mM), A‐485[10 nM, (J); 3/10 nM, (K)], KAT8‐IN‐1 (300 µM), NU9056 (2 µM), Butyrolactone 3 (MB‐3, 300 µM), or Gln‐AMS (10 µM) for 14 h. Quantitative analysis of extracellular FFAs was conducted post‐treatment.

Fig 5: Western blots of brain homogenates to assess neuroinflammation. This is a representative Western blot of brain homogenates for mouse groups as described in Figure 1 with mean ± SD values shown above each band expressed as fold-change from control for replicate studies. Normalization of band densities to β actin in the same samples was performed for daily experiments and then values for experimental groups were expressed as a ratio against the control value (thus control is shown as 1.0). Bold numbers indicate values statistically significantly different from control (p ≤ 0.01, Kruskal–Wallis test). Blots were probed for thrombospondin-1 (TSP1), Ly6G, myeloperoxidase (MPO), Aquaporin-4 (AQP4), NF-κB, p65-phosphorylated NF-κB (pNF-κB), CD36, phosphatidyl serine, and IL-1β proteins.