Fig. 3: Infusion of ex vivo polarized BMDMs alters macrophage polarization and CER metabolism in the liver after partial hepatectomy.

Mouse livers after hepatectomy were infused with phosphate buffer saline (PBS) (Veh), M0, M1, and M2 BMDMs. The remnant liver tissues were collected at postoperative day 2 (POD2) to examine macrophage polarization and levels of CER metabolites. A Liver tissues were harvested at 2 days and 4 days after GFP-positive polarized BMDM infusion. Liver sections were stained with anti-GFP (green), anti-F4/80 (macrophage marker, red), anti-Inos (M1 marker, yellow), and anti-CD206 antibodies (M2 marker, yellow) to validate the infiltration of polarized BMDM in the remnant liver after hepatectomy by confocal scanning. DAPI was used to visualize nuclei (blue). B, C Liver sections were co-stained with F4/80 (red) and iNos (green) (B) or CD206 (green) (C) to examine M1 or M2 macrophage polarization in response to BMDM infusion in the remnant liver after hepatectomy. DAPI was used to visualize nuclei (blue). D M1 and M2 macrophages in the remnant liver tissues were counted in five random 20× fields to quantify alteration in M1 and M2 polarization by BMDM infusion in the remnant liver after hepatectomy. E, F Total RNA was extracted from the remnant liver tissues, then mRNA levels of M1-macrophage-associated genes (Tnf-α, Inos, and Il-6) (E) and M2-macrophage-associated genes (Fizzl, Arg1, Il-4, and Il-10) (F) were determined by qPCR. G, H Levels of CER metabolites were determined in the remnant liver tissues with BMDM infusion after hepatectomy, including d18:1-CER (G), d18:1-SPH (H), and d18:1- S1P (H) by targeted UHPLC-ESI-MS/MS. Images in A–C represent results from five individual mice in each group. Data in E–H are demonstrated as mean ± SD, n = 5–6; *p < 0.05, **p < 0.01, ***p < 0.001.