Fig. 5

Figure 5. Cxcl8a is necessary for recruiting neutrophils to the islet (A) Quantification of islet visits by neutrophils in live imaging videos from 64–72 h. Videos were recorded at 30 s/frame. Data are mean ± SEM. n = 3/group, one-way ANOVA followed by Tukey’s multiple comparisons test; ∗p < 0.05, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001. (B) Quantification of islet visits by neutrophils in 30 s/frame live imaging videos of control and zMIR fish from 66–67 h. Unpaired t test; n = 4/group; bar graphs represent mean ± SEM; ∗∗∗p < 0.001. See also Videos S1A and S1B. (C) Representative tracks of neutrophil movement in the islet vicinity in a zMIR fish from 66–68 h. Scale bars, 50 μm. See also Video S2. (D) β cell loss during the 67–68 hpt interval in a representative islet. White arrows point to the β cells lost during the interval. Scale bars, 15 μm. See also Video S3C. (E) Quantification of islet visits by neutrophils in live imaging videos of tnfa−/−, zMIR fish; irf8−/−, zMIR fish; cxcl8a−/−, zMIR fish, and control zMIR fish from 66–67 h. Videos were recorded at 30 s/frame. Data are mean ± SEM. n = 3/group, one-way ANOVA followed by Tukey’s multiple comparisons test; ∗∗∗p < 0.001. (F) Representative video frames showing neutrophils tracking a macrophage in the principal islet in zMIR fish from 66–67 hpt. Yellow arrows point to macrophages, and blue arrows point to neutrophils. Videos were recorded at 30-s intervals. Macrophages were distinguished from neutrophils by their morphological irregularity and lower velocity. See also Video S3D. (G) Representative immunofluorescence images of neutrophil elastase and CXCl15 in pancreas sections of 8-week-old db/+ (B6.BKS(D)-Leprdb/J) and db/db mice. Scale bars, 50 μm. Inset: contact between neutrophils and CXCl15-positive β cells. Scale bars, 10 μm. (H) Quantification of intra-islet neutrophil numbers of db/+ and db/db mice (n = 4; at least 50 islets were quantified in each animal). Unpaired t test, bar graphs represent mean ± SEM, ∗∗∗p < 0.001. (I) Quantification of CXCL15-positive β cells in db/+ and db/db mice (n = 4; at least 50 islets were quantified in each animal). Unpaired t test, bar graphs represent mean ± SEM, ∗∗∗p < 0.001. (J) Representative immunofluorescence images of neutrophil elastase and CXCl15 in pancreas sections of 8-week-old control (C57BL/6) and Akita (C57BL/6-Ins2Akita/J) mice. Scale bars, 50 μm. Inset: contacts between neutrophils and CXCl15-positive β cells. Scale bars, 10 μm. (K) Quantification of intra-islet neutrophil numbers of control and Akita mice (n = 4; at least 50 islets were quantified in each animal). Unpaired t test, bar graphs represent mean ± SEM, ∗∗p < 0.01. (L) Quantification of CXCL15-positive β cells in control and Akita mice (n = 4; at least 50 islets were quantified in each animal). Unpaired t test, bar graphs represent mean ± SEM, ∗∗∗p < 0.001.