Fig 1.
Characterization of untreated Ifnardel mice reveals no differences to wild type mice.
(A) qRT PCR analysis of mRNA levels of Ifnar1 from whole pancreatic tissue of untreated WT and Ifnardel mice. (n = 3 per group. Bars indicate mean +/- SD. Normalized on the mRNA of the housekeeping gene Ppib. **P<0.01, Mann-Whitney-test). (B) Representative H&E staining of the pancreas from 8 week old WT and Ifnardel mice without treatment (original magnification, 100x). (C) Pancreas weight/body weight ratio of 8 week old WT and Ifnardel mice without treatment (n = 7 per group. Bars indicate mean +/- SD). (D) Amylase, lipase and LDH (lactat-dehydrogenase) levels in the serum from of 8 week old WT and Ifnardel mice without treatment (n = 7 per group. Bars indicate mean +/- SD).
Fig 2.
Pancreatic inflammation after caerulein- induced injury is limited in Ifnardel mice.
(A) Representative H&E staining of the pancreas from WT and Ifnardel mice, 24 and 48 hours following caerulein-induced injury (original magnification, 100x). (B) Representative H&E staining of the pancreas from WT and Ifnardel mice, 3, 7 and 14 days following caerulein-induced injury (original magnification, 100x). (C-E) Combined and individual scoring of pancreatic histological parameters from WT and Ifnardel mice, 24, 48 hours, 3, 5, 7 and 14 days following caerulein-induced injury (n = 3–9 per group. Bars indicate mean +/- SD. *P<0.05, **P<0.01, ***P<0.001, ****P<0.0001, Mann-Whitney-test).
Fig 3.
Macrophages are the predominant immune cells infiltrating the pancreas following caerulein-induced injury in Ifnardel mice.
(A) The absolute number of CD45-positive immune cells, B220-positive B-lymphocytes, CD3-positive T-lymphocytes and MPO-positive neutrophils was counted in five separate high power fields for each section of untreated WT and Ifnardel mice and 24, 48 hours, 3, 5, 7 and 14 days following caerulein-induced injury (n = 3–9 per group. Bars indicate mean +/- SD. *P<0.05, **P<0.01, ****P<0.0001, Student´s t-test). (B) Immunohistochemical staining for F4/80-positive macrophages in the pancreas from WT and Ifnardel mice 24h following caerulein-induced injury and counting of the absolute number of positive cells in five separate high power fields for each section of untreated WT and Ifnardel mice and 24, 48 hours, 3, 5, 7 and 14 days following caerulein-induced injury (n = 3–9 per group. Bars indicate mean +/- SD. *P<0.05, ****P<0.0001, Student´s t-test. Original magnification, 200x and 650x).
Fig 4.
Depletion of macrophages rescues the focally restricted inflammation phenotype in Ifnardel mice.
(A) Representative H&E staining of the pancreas from WT and Ifnardel mice 48 hours after treatment. Mice were either treated with caerulein (regeneration model, left), caerulein plus clodronate filled liposomes (macrophage depletion, middle) or caerulein plus PBS filled liposomes (PBS control group, right) as described in the methods section (n = 3; original magnification, 100x). (B-D) Combined and individual scoring of pancreatic histological parameters from WT and Ifnardel mice 24 and 48 hours, and 5 days following caerulein-induced injury (plain bars) or caerulein-induced injury combined with macrophage depletion (striped bars) (n = 3–9 per group. Bars indicate mean +/- SD. *P<0.05, **P<0.01, ***P<0.001, ****P<0.0001, Mann-Whitney-test).
Fig 5.
Proinflammatory cytokine expression in WT and Ifnardel mice.
(A-B) qRT PCR analysis of mRNA levels of the proinflammatory cytokines Ifnγ, Tnfα, IL-6, Lbp, G-Csf, Ifnα, IL-1α and IL-1β (A) and the anti-inflammatory cytokines IL-10, IL-13, Tgfβ1–3 and Csf (B) from whole pancreatic tissue of WT and Ifnardel mice harvested 4 hours following caerulein-induced injury (n = 3 per group. Bars indicate mean +/- SD. Normalized on the mRNA of the housekeeping gene Ppib). (C-D) qRT PCR analysis of mRNA levels of the proinflammatory cytokines Ifnγ, Tnfα, IL-6, Lbp, G-Csf, Ifnα, IL-1α and IL-1β (C) and the anti- inflammatory cytokines IL-10, IL-13, Tgfβ1–3 and Csf (D) from whole pancreatic tissue of untreated WT and Ifnardel mice (n = 7 per group. Bars indicate mean +/- SD. Normalized on the mRNA of the housekeeping gene Ppib. *P<0.05, unpaired Student´s t-test).
Fig 6.
Release of the chemoattractant Ccl2/MCP1 in untreated Ifnardel mice.
(A) qRT PCR analysis of mRNA levels of the chemokines Cxcl9, Cxcl10, Cxcl11, Ccl2, Ccl4, Ccl5 and Ccl25 from whole pancreatic tissue of WT and Ifnardel mice harvested 4 hours following caerulein-induced injury (n = 3 per group. Bars indicate mean +/- SD. Normalized on the mRNA of the housekeeping gene Ppib). (B) qRT PCR analysis of mRNA levels of the chemokines Cxcl9, Cxcl10, Cxcl11, Ccl2, Ccl4, Ccl5 and Ccl25 from whole pancreatic tissue of untreated WT and Ifnardel mice (n = 7 per group. Bars indicate mean +/- SD. Normalized on the mRNA of the housekeeping gene Ppib. *P<0.05, unpaired Student´s t-test). (C) Immunohistochemical staining for MCP1-positive centro-acinar cells in the pancreas from WT and Ifnardel mice 24 hours following caerulein-induced injury or untreated and counting of the absolute number of positive cells on five separate high power fields for each section of untreated WT and Ifnardel mice and after 24 hours following caerulein-induced injury (n = 3–7 per group. Bars indicate mean +/- SD. ****P<0.0001, Mann-Whitney-test. Original magnification, 200x and 1000x).