Fig. 6

CX3CR1⁺ macrophages inhibit the growth of NR5A1 ⁺ tumor cells through INHBA/activin A—ACVR1B axis. A Histogram plot revealed the gene expression alteration of the tumor cells co-cultured with CX3CR1⁺ macrophages (n = 3). B–D HE images with pie charts in each spot colored by annotation showing the cellular composition in the ST sample (B). The spatial feature plot shows the location and proportions of macrophages in ST spots using deconvolution results (C). Spatial feature plots show the expression of INHBA and its receptor ACVR1B in the macrophage-containing spots (D). E Histogram plot revealed the gene expression alteration of the tumor cells co-cultured with activin A. F–G The representative image of immunofluorescence revealing the specific expression of INHBA in CX3CR1⁺ and CX3CR1⁻ macrophages (F) and the statistics of INHBA mean fluorescence intensity (MFI) in CX3CR1⁺ and CX3CR1⁻ macrophages in three patients (G). H–I The cell viability (H) and apoptosis (I) in the four NR5A1⁺ primary cell treated activin A (10 ng/ml) with or without SB-505124 (1 μM) or follistatin (100 ng/ml). The histogram shows the cells' apoptosis in different conditions for the primary cell (I). J Cell viability assessment in the AtT20 cell line treated with Activin A (10 ng/ml) in the presence or absence of SB-505124 (1 μM), A 83–01 (1 μM), or follistatin (100 ng/ml). K Cell viability analysis in AtT20 cell line with shEV-GFP or shAcvr1b treated with Activin A (10 ng/ml). L–M. Apoptosis assessment in the AtT20 cell line treated with Activin A (10 ng/ml) in the presence or absence of SB-505124 (1 μM), A 83–01 (1 μM), or follistatin (100 ng/ml). The histogram illustrates the apoptosis levels of cells under various conditions in the AtT20 cell line (M). N–P Tumor growth curve (N). The xenografts harboring mice were injected intraperitoneally with activin A release preparations, 5 mg/kg weight, every other day with PBS. Representative images of H&E and IF staining (Ki-67 and Cleaved-caspase 3) of resected AtT20 tumors from an experimental study between different groups (O). Scale bar as indicated. Statistical analysis of Ki67 and Cleaved-caspase 3 percentages in control and treated Att20 tumors (P). Q The staining revealed the co-localization of CD68 and CX3CR1 in xenografts harboring mice. Arrow indicated the CD68 and CX3CR1 positive staining. Scale bar as indicated. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. P-values were calculated using the Wilcoxon rank-sum test. Results are presented as mean ± standard error of the mean in bar plots