Fig. 3: Search for active molecule driving healthy fibroblast cell state.

A Left: Schematic illustrating generation and application of fat-conditioned media (FCM). Created with Biorender. Right: FCM induces healthy fibroblast signature. n = 4 technical replicates, representative of two independent experiments. B Left: Schematic illustrating generation and application of adipocyte-conditioned media (ACM). Created with Biorender. Right: ACM induces the healthy fibroblast gene signature. n3 technical replicates, representative of two independent experiments. C Bligh and dyer separation of adipocyte-conditioned media (ACM). n = 3 technical replicates, representative of two independent experiments. D Left: Schematic detailing fractionation approach. ACM was separated using the Bligh and Dyer method into aqueous and organic phases. Then, the organic phase was taken for solid phase separation and eluted based on polarity using chloroform, acetone, methanol, and water. Right: Testing activity of each fraction. C, D Statistical comparisons: all groups were compared to basal/ctl. Basal: n = 6, ACM: n = 3, CHCL3: n = 5, Acetone: n = 5, Methanol: n = 5, H2O: n = 5 technical replicates, representative of two independent experiments. E MS intensity values were shown as the ion chromatogram areas extracted at m/z 303.233 ([M–H]-) for arachidonic acid (AA), m/z 356.070 ([M–H]-) for indomethacin (INDO), m/z 221.104 ([M–H]-) for isobutylmethylxanthine (IBMX), and m/z 437.198 ([M + COO]-) for dexamethasone in all three acetone fractions. n = 3 technical replicates of three SPE column purifications from one ACM, one independent experiment. All data are presented as mean ± standard deviation. A, B P values were calculated using a two-tailed Student’s T test; C, D were calculated using an ordinary one-way ANOVA followed by Dunnet’s multiple comparison post hoc testing. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. Source data are provided as a Source Data file. A, B Created with BioRender.com released under a Creative Commons Attribution-NonCommercial-NoDerivs 4.0 International license (https://creativecommons.org/licenses/by-nc-nd/4.0/deed.en).