Fig 2: Systemic administration of mMSCs, BMDMCs, CD34-, CD3-, CD19-, CD31-, and Sca-1-depleted BMDMCs, but not HLFs, or CD11b-, and CD45-depleted BMDMCs, significantly reduced histologic lung inflammation provoked by AHE sensitization and challenge. (A): Inflammation score of the airway in N and A mice treated with HLF, mMSCs, BMDMCs, CD45-, CD34-, CD11b, CD3, CD19, CD31-, and Sca-1-depleted BMDMCs (N, n = 17; A-P, n = 15; A-HLF, n = 6; A-mMSC, n = 6; A-BMDMC, n = 13; A-CD45, n = 6; A-CD34, n = 6; A-CD11b, n = 9; A-CD3, n = 6; A-CD19, n = 6; A-CD31, n = 6; A-Sca-1, n = 6). Data are presented as median and interquartile ranges. (B): Representative photomicrographs stained with hematoxylin and eosin. Original magnification, ×10. Scale bar = 25 µm. *, p = .05 compared with N; #, compared with A-P; **, compared with A-BMDMC. Abbreviations: A, Aspergillus hyphal extract-exposed mice; A-CD3 (or A-CD11b-, A-CD19-, A-CD31-, A-CD34-, A-CD45-, A-Sca-1-), Aspergillus hyphal extract-exposed mice treated with CD-3- (or CD11b-, CD19-, CD31-, CD34-, CD45-, and Sca-1-) depleted bone marrow-derived mononuclear cells; A-HLF, Aspergillus hyphal extract-exposed mice treated with human lung fibroblasts; A-P, Aspergillus hyphal extract-exposed mice treated with phosphate-buffered saline; BMDMC, bone marrow-derived mononuclear cell; HLF, human lung fibroblast; mMSC, mouse mesenchymal stem/stromal cell; N, naïve; P, phosphate-buffered saline.

Fig 3: Ndrg1 is associated with B cell anergy.a Top panel: differential gene expression between IgHEL naïve follicular (B220+CD19+IgMa+CD93-CD21midCD23hi) and IgHEL/sHEL anergic (B220+IgMaloIgDa+) splenic B cell populations, represented in FACS plots. Lower Panel: Genes differentially expressed >1 and <-1 log2 fold change (FC) and below a significance threshold of adjusted p value, 10e-32, are shown in red, with the top 30 differentially expressed genes filtered by adjusted p value labeled. b Heatmap of the relative abundance of the top 30 differentially expressed genes by adjusted p value between naïve (left) and anergic (right) B cells. Columns represent samples from individual mice.

Fig 4: Study design. (A): Protocol of allergic airway inflammation induction. OP administration of PBS (naïve) or AHE solution on days 0 and 7 to initiate the immune response, then challenged for 3 days on days 14, 15, and16 with OP inoculations of AHE. Cell administration was performed on day 14; harvest was on day 19. (B): Experimental groups. Animals exposed to AHE solution (A) were randomly divided to receive treatment with the vehicle PBS, HLFs, mMSCs, BMDMCs, and BMDMCs depleted of CD45, CD34, CD11b, CD3, CD19, CD31, and Sca-1 positive cells. Abbreviations: #, oropharyngeal Aspergillus hyphal extract solution administered; ?, human lung fibroblasts, mouse mesenchymal stem/stromal cells, bone marrow-derived mononuclear cells and depleted bone marrow-derived mononuclear cells administered; A, animals exposed to AHE solution; AHE, Aspergillus hyphal extract; BMDMC, bone marrow-derived mononuclear cell; HLF, human lung fibroblast; mMSC, mouse mesenchymal stem/stromal cell; OP, oropharyngeal; P, group receiving phosphate-buffered saline; PBS, phosphate-buffered saline.

Fig 5: NDRG1-deficient B cells exposed to self-antigen in the periphery are subject to follicular exclusion and cell death.a Representative flow cytometry plots of 50:50 mixtures of WT CD45.1+ IgHEL and WT or Ndrg1-/- CD45.2+ IgHEL splenocytes labeled with CTV dye and transferred by i.v. injection into non-tg or sHEL-tg CD45.2+ recipients for 48 h, showing live B220+CD19+ gated populations and % transferred CTV+ cells. b Percentage of total lymphocytes that were either CD45.1+ or CD45.2+ CTV+ as gated in (a), with white points showing paired CD45.1+ IgHEL cells from donor, colored points are CD45.2+. c MFI of surface marker expression CD86, CD23, CXCR5, IgMa, BAFF-R and CD95, and intracellular Bim and Bcl2, in transferred IgHEL B cells, gated as in (a). Data points are individual mice and lines show means with 95% CI error bars and data are representative from 1 to 3 independent experiments with 4–6 mice per group. d, e Ratio of B220+ cells surviving from 50:50 mixtures of WT CD45.1+ and WT or Ndrg1-/- CD45.2+ non-tg B cells stimulated with anti-IgM at the indicated concentrations for 16 h at 37 °C ex vivo (d) and CD86 surface expression as MFI on the CD45.2 + B220+ cells (e). Data are representative of 3 independent experiments with columns or circles as means with 95% CI error bars.