Figure 1. CRIg⁺TRMs form a protective barrier to prevent tissue autoimmune infiltration and activation.

(A) The distribution of CRIg⁺ TRMs within the pancreas. Immunostaining of pancreatic frozen sections of 10-week-old NOD mice. (left) Representative images depicting an intact (upper) and an insulitic (lower) islet, respectively. Red: CRIg; Blue: DAPI. The border of the islet was marked by dotted lines; immune infiltration was highlighted by higher density of DAPI⁺ dots (n = 9 for intact and n = 8 for insulitic). Bar, 50 um. (right) The numbers of CRIg⁺ TRMs per islet counted from immunostaining sections of pancreas as in (A). Islets were categorized into intact (free from infiltration) and insulitic groups. (B) The severity of insulitis in 10-week-old NOD and NOD/CRIg KO mice. Hematoxylin and eosin staining of pancreatic paraffin sections. Arrowheads depict massive immune infiltration. Bar, 50 um. (C) Flow cytometric analyses of digested pancreases of NOD/CRIg KO mice and littermate controls (n = 7 in each group). (Left) The percentages of pancreatic Treg cells in age-matched NOD and NOD/CRIg KO mice (mixed of both females and males). (Right) The ratio between Treg cells and CD8⁺ T cells (n = 7 in each group). (D) The production of IFN-γ in CD4⁺ Tconv and CD8⁺ T cells. Pancreatic digestions were the same as in (C). Data are representative of three (A, B) or four (C, D) experiments. Student’s t-test was used. *p<0.05; **p<0.01; ***p<0.001.

Figure 1—figure supplement 1. Tissue-distribution of CRIg⁺ TRMs.

(A) Immunostaining for CRIg on frozen sections of the pancreas, liver, small and large intestines and lung from 7-week-old B6 mice. lower panel: second Ab only. LP, lamina propria. Bar: 50 um. (B) Flow cytometric analyses of CRIg expression in different tissues. The strain and age of the mice were the same as in (A). Bar: 50 um. Data are representative of at least three experiments. (C) The expression of CRIg in human pancreas. Red, CRIg; green, insulin. Dotted lines depicted islets. Data are representative of four human pancreases. (D) Flow cytometric analyses of CRIg in lymphoid-lineage cells. None of these cells expresses CRIg.

Figure 1—figure supplement 2. CRIg deficiency does not affect T cells in lymphoid organs.

(A) The percentages of Treg cells in age-matched wildtype and CRIg KO mice (mixed of both females and males) in spleen and pancreatic draining LNs (panLNs). (B) The ratios between Treg cells and CD8⁺ T cells in age-matched wildtype and CRIg KO mice (mixed of both females and males) in spleen and panLNs. (C) The production of IFN-γ in CD4⁺ Tconv from spleen or panLNs of wildtype and CRIg KO mice. (D) The production of IFN-γ in CD8⁺ T cells from spleen or panLNs of wildtype and CRIg KO mice. Data are representative of four experiments. Student’s t-test was used. n.s., non-significant. WT, wildtype; KO, CRIg knockout.

Figure 1—figure supplement 3. Cell-cell contact between CRIg⁺TRMs and T cells in pancreas.

(A) Cell-cell contact between CRIg⁺ macrophages (green) and islet-infiltrating CD4 T cells (red). Arrowheads, CD4⁺ T cells; long arrows, CRIg⁺ TRMs. Bar, 50 um.