Fig 1: Pattern of intraepithelial mononuclear phagocyte recruitment following viral exposure of the colonic mucosa(A–L) Human (A–C, G–I) and macaque (D–F, J–L) colonic tissues incubated for 30 min either in medium with HIV-1 AD8 (A–C, with brightfield image superimposed to the fluorescence) or SIVmac251 (D–F), or medium only (G–L), fixed with 4% PFA and cryopreserved are shown. Labeling of CD11c-FITC (green) in combination with either CD64 (A, B, D, E) followed by goat-anti-mouse Alexa Fluor 594 (red) or CX3CR1-PE (C, F) was performed. Arrows point to subsets of CD11c+ CD64+ (A and D), CD11c+ CD64− (B and E), and CD11c+ CX3CR1+ (C and F) cells that migrate from the lamina propria to in between the epithelial cells following viral exposure. A magnification of the intraepithelial cells is shown in panels A–F (the yellow signal identifies double-positive cells). Magnification of selected regions (indicated by arrows) is shown in the boxed areas of H, J, and K panels. DAPI was used to stain the nuclei. Scale bars indicate the magnification.
Fig 2: HIV-1 coreceptor expression by mononuclear phagocytesImmune phenotyping was performed as described in STAR Methods and Table S1.(A–D) Proportion of CD11c+ CD64- DCs, CD11c+ CD64+ Mfs, and CD11c- CD64+ Mfs expressing the CCR5 receptor in the colonic mucosa from five macaques CCR5 and CXCR4 expression by (B) total CD11c+ HLA-DR+ lineage- MNPs, (C) CD11c+ CD103+ MNPs, and (D) CD11c+ CX3CR1+ MNPs in human (n = 5) and macaque (n = 6) colon samples is presented. The mean ± the SD is plotted. Wilcoxon rank pairs tests were applied in panel A, whereas Friedman tests with FDR correction were used in panels B–D. p-values < 0.05 were considered significant (*p < 0.05, **p < 0.01, ***p < 0.001).
Fig 3: Distribution of mononuclear phagocyte subsets in human and macaque colonic mucosa at steady state(A–L) Sections from untreated human (A–F) and macaque (G–L) colonic tissues fixed with 4% PFA and cryopreserved are shown. DC and Mf subsets were identified by labeling with CD11c-FITC (green) in combination with either CX3CR1-PE, CD103, or CD64, followed by goat-anti-mouse Alexa Fluor 594 (red). DAPI (blue) was used to stain the nuclei. The bright field image (BF, gray) was superimposed with the fluorescence images to visualize the contour of the intestinal barrier. The lower panels (D–F and J–L) are magnifications of the boxed area in the upper panel. Scale bars indicate the magnification.
Fig 4: Intraepithelial CD103+ cells are CD3+ T lymphocytes and not CD11c+ DCs(A–F) Distribution of CD3+ CD103+ T lymphocytes (yellow cells in the upper panels) and CD11c+ CD103+ DCs (yellow cells in the lower panels) and following 30 min of incubation with medium, HIV-1 AD8, or SIV, respectively, are shown. DAPI was used to stain the nuclei (blue). Brightfield images (BF) were superimposed to visualize the intestinal barrier contour. Arrows indicate the intraepithelial cells (or cells partially interposed to epithelium), i.e. CD3+ CD103+ (A–C), CD3+ CD103− (D–F) and CD11c− CD103+ (D–F). Scale bars indicate the magnification.
Fig 5: Phenotypic characterization of lamina propria mononuclear phagocytes in the human and macaque colon(A and B) Representative flow cytometry plots of MNPs in untreated and morphologically preserved human (A) and macaque (B) colon. Leukocytes are identified among live cells as CD45+. Three subsets of MNPs were identified based on the expression of CD11c and CD64 within single, viable, CD45+ HLA-DR+ lineage- cells: CD11c+ CD64- DCs, CD11c+ CD64+ Mfs, and CD11c- CD64+ Mfs.(C and D) The frequency of (C) CD45+ cells and (D) HLA-DR+/lineage- cells (E, F) are shown. Symbols: DCs and Mfs in humans = round symbols; macaques = squared symbols; colon sample number: n = 12 for human and n = 10 for macaque tissues. The mean ± the SD is plotted. Mann-Whitney tests were used to compare the frequency of cells between human and macaque colon, whereas Friedman tests with FDR correction were used to compare DC and Mf numbers within the same colon donor. p-values < 0.05 were considered significant (*p < 0.05, **p < 0.01, ***p < 0.001).
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