Fig 2: Inflammation induces changes in macrophage subtypes and the development of IUA(A–D) H&E staining (scale bars: 200 µm), Masson’s trichrome staining, and immunohistochemical staining of a-SMA and collagen 1 (scale bars of Masson, a-SMA and Col1: 50 µm) in the endometria of mice after sham operation (sham, n = 8) and on Day 5 (day 5, n = 8), Day 7 (day 7, n = 8) and Day 10 (day 10, n = 8) after modeling.(E) The relative integrated optical density (IOD) was analyzed by Image-pro plus.(F–K) H&E staining (scale bars: 200 µm), immunohistochemical staining of CD45, immunofluorescence analysis of F4/80 (scale bars of CD45 and F4/80: 100 µm), and immunohistochemical staining of CD3, Eomes and CD301 (scale bars of CD3, Eomes and CD301: 50 µm) in the endometria of mice after sham operation (sham, n = 8) and on Day 1 (Day 1, n = 8), Day 2 (Day 2, n = 8), Day 3 (Day 3, n = 8), Day 4 (Day 4, n = 8) and Day 5 (Day 5, n = 8) after modeling.(L) Double immunofluorescence staining of F4/80 (green) and CD301 (red) in the endometria of mice. Scale bars: 50 µm. Arrow: F4/80+ CD301+ macrophage.(M) The mean number of CD45+ leukocytes, F4/80+ macrophages, CD3+ T cells and Eomes+ NK cells per field at a magnification of 200×. Each point represents one mouse.(N) The mean number of CD301+ macrophages per field at a magnification of 400×. Each point represents one mouse. (E) and (M-N) Error bars indicate the mean ± SD. No significant difference (ns), *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.

Fig 3: 38–2 mAb induces M2 polarization through activation of SFK in peritoneal macrophages.(A) Uncropped, full picture of Western blotting for PrPC with 38–2 mAb in the lungs (Lg) and peritoneal macrophages (PM) from WT and Prnp0/0 mice. Actb, ß-actin. (B) Western blotting for total SFK, phosphorylated SFK (Tyr416), and MGL1/2 in peritoneal macrophages from WT and Prnp0/0 mice 3 hrs after treatment with control IgG and 38–2 mAb together with DS or control DMSO. (C) ELISA for INF-?, TNF-a, IL-4, and IL-10 in the culture medium from WT peritoneal macrophages at 0.5, 4, 16, and 24 hours (hrs) after treatment with control IgG and 38–2 mAb.*, p<0.05;** p<0.01. (D) Western blotting for total SFK, phosphorylated SFK (Tyr416), and MGL1/2 in peritoneal macrophages (PM) from WT and Prnp0/0 mice 1 and 3 days after peritoneal injection with control IgG and 38–2 mAb together with control DMSO and DS.

Fig 4: ITI-hUC-MSCs secrete C1INH to inhibit CD301+ macrophage polarization(A and B) The proportion of CD301+ macrophages and mean fluorescence intensity (MFI) of CD301 in PMA-treated THP-1 cells stimulated with LPS and IFN-? for 24 h and then treated with conditioned media (CM) from hUC-MSCs (n = 3) and ITI-hUC-MSCs (n = 3) for another 24 h was measured by flow cytometry.(C and D) Heatmap and volcano plot showing DEGs in hUC-MSCs (n = 3) and ITI-hUC-MSCs (n = 3). The red dots represent the upregulated transcripts, and the blue dots represent the downregulated transcripts (FDR (false discovery rate) < 0.05 and fold change >2).(E) Venn analysis of the top 200 genes by FC (fold change), top 200 genes by FDR and top 200 genes by expression abundance.(F) Heatmap showing the 43 genes filtered out through Venn analysis.(G) The mRNA levels of Serping1, Cxcl5, Il32, Il32a, Il32ß, Il32? and Mmp3 in hUC-MSCs (n = 4) and ITI-hUC-MSCs (n = 4) were measured by qRT?PCR.(H) The protein level of C1INH was examined by western blotting in hUC-MSCs (n = 3) and ITI-hUC-MSCs (n = 3).(I) The relative band intensities of C1INH were analyzed by ImageJ.(J and K) The proportion of CD301+ macrophages and MFI of CD301 in PMA-treated THP-1 cells stimulated with LPS and IFN-? for 24 h and treated with 100 ng/mL recombinant C1INH for another 24 h (n = 3) were measured by flow cytometry.(L) The mRNA level of Serping1 in hUC-MSCs (n = 5) transfected with three different small interfering RNA-seq of Serping1 (si-1, si-2, si-3) or negative control (si-NC) for 48 h was examined by qRT?PCR.(M) The protein level of C1INH in hUC-MSCs (n = 3) transfected with si-1, si-2, si-3 or si-NC for 48 h was examined by western blotting.(N) The relative band intensities of C1INH were analyzed by ImageJ.(O and P) The proportion of CD301+ macrophages and MFI of CD301 in PMA-treated THP-1 cells stimulated with LPS and IFN-? for 24 h and treated with conditioned media of ITI-hUC-MSCs transfected with si-NC (n = 3) and si-Serping1 (si-2; n = 3) for another 24 h was measured by flow cytometry. (A-B), (G), (I-L) and (N-P) Error bars indicate the mean ± SD. No significant difference (ns), *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.

Fig 5: 3S9 and 2H9 anti-PrP mAbs evokes protection against IAV infection through a similar mechanism.(A) Western blotting for total SFK, phosphorylated SFK (Tyr416), and MGL1/2 in peritoneal macrophages from WT and Prnp0/0 mice 3 hrs after treatment with control IgG and 3S9 and 2H9 mAbs together with control DMSO and DS. (B) Survival rate (%, upper panels) and body weight loss (%, lower panels) of WT mice intraperitoneally administered with control IgG and 3S9 and 2H9 mAbs 1 day before intranasal infection with 100 (left panels) or 200 IFU (right panels) of IAV/PR8. Error bars, SD. *, p<0.05; **, p<0.01; IgG vs 3S9. #, <0.05; IgG vs 2H9.