Skip to main page content
Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2017 May 16;19(7):1322-1333.
doi: 10.1016/j.celrep.2017.04.052.

The Presence of Interleukin-13 at Pancreatic ADM/PanIN Lesions Alters Macrophage Populations and Mediates Pancreatic Tumorigenesis

Affiliations
Free PMC article

The Presence of Interleukin-13 at Pancreatic ADM/PanIN Lesions Alters Macrophage Populations and Mediates Pancreatic Tumorigenesis

Geou-Yarh Liou et al. Cell Rep. .
Free PMC article

Abstract

The contributions of the innate immune system to the development of pancreatic cancer are still ill defined. Inflammatory macrophages can initiate metaplasia of pancreatic acinar cells to a duct-like phenotype (acinar-to-ductal metaplasia [ADM]), which then gives rise to pancreatic intraepithelial neoplasia (PanIN) when oncogenic KRas is present. However, it remains unclear when and how this inflammatory macrophage population is replaced by tumor-promoting macrophages. Here, we demonstrate the presence of interleukin-13 (IL-13), which can convert inflammatory into Ym1+ alternatively activated macrophages, at ADM/PanIN lesions. We further show that Ym1+ macrophages release factors, such as IL-1ra and CCL2, to drive pancreatic fibrogenesis and tumorigenesis. Treatment of mice expressing oncogenic KRas under an acinar cell-specific promoter with a neutralizing antibody for IL-13 significantly decreased the accumulation of alternatively activated macrophages at these lesions, resulting in decreased fibrosis and lesion growth.

Keywords: CCL-2; IL-13; IL-1ra; PanIN; Tuft cells; interleukin-13; macrophages; metaplasia; pancreatic cancer; polarization.

Figures

Figure 1
Figure 1. ADM and PanIN show different predominance of macrophage polarization types
A: Patient samples that contain PanIN1/2 lesions were stained with H&E (left panel), or analyzed by immunofluorescence for presence of macrophages (anti-CD68, middle panel). The inset marks the zoom-in area for which presence of different macrophage subtypes were determined by using iNOS or CD163 as markers. DAPI stains nuclei. Scale bar: 50 μm. B: Quantification of iNOS+ or CD163+ macrophages localized at human PanIN1/2 areas (n=10). The asterisk indicates statistical significance (p < 0.05) of the increase in CD163+ as compared to iNOS+ cells. C: H&E staining of pancreatic tissue of 14 week old KC mice showing typical areas of PanIN1 and ADM. Immunofluorescence pictures show analysis of presence of macrophage subtypes by labeling with F4/80 in combination with pSTAT1 or YM1. Scale bar: 50 μm. D: Quantification of F4/80/pSTAT1+ or F4/80/Ym1+ macrophages in ADM (n=8) and PanIN areas (n=9). * indicates statistical significance (p < 0.05) as compared to F4/80/pSTAT1+ macrophages in ADM regions; ** indicates statistical significance (p < 0.05) as compared to F4/80/Ym1+ macrophages in ADM regions.
Figure 2
Figure 2. Expression of IL-13 is increased in PanIN and pancreatic cancer
A, H: Pancreatic tissue of 18 week old KC or control mice (A) or pancreatic cancer patient samples (H) were analyzed by immunohistochemistry for expression of IL-13. Scale bar: 50 μm. B: Quantification of samples (from A) for IL-13 expression. * indicates statistical significance (p < 0.05) as compared to normal acinar cells; ** indicates statistical significance (p < 0.05) as compared to ADM regions. C: PanIN1A/B (n=50) or adjacent “normal” acinar cells (n=50) were microdissected from KC mice and analyzed by q-PCR for IL-13 mRNA expression. D: Indicated pancreatic tissues from KC mice were analyzed for IL-13 mRNA expression (brown dots) using RNAscope in situ hybridization (ISH). E, F: PanIN areas from KC mice were analyzed for IL-13 mRNA expression (brown dots) using RNAscope ISH and DCLK1 or Desmin protein expression (pink staining) using IHC. G–I: Pancreatic tissue of 14 week old KC or control mice (G) or pancreatic cancer patient samples (H, I) were analyzed by immunofluorescence labeling (G, I) for IL-13 and acetyl-α-tubulin. Nuclei were visualized by DAPI staining. Samples in H were stained by IHC for IL-13. In I, the rectangle window represents an area for which single channels are shown. Scale bars in GI: 50 μm.
Figure 3
Figure 3. Neutralization of IL-13 decreases the presence of Ym1+ macrophages at PanIN lesions and results in an overall decrease in abnormal pancreatic structures
A: Treatment scheme. KC or control mice at an age of 3 weeks were treated with indicated antibodies for 7 weeks. At 10 weeks pancreas tissues were harvested. B: Representative H&E staining of pancreas tissue samples from KC mice treated as indicated in A. Scale bar: 200 μm. C: Quantification of total abnormal structures including ADM and PanIN (grey) and grading of different pancreatic lesions. The asterisk indicates statistical significance (p<0.05) as compared to isotype control antibody-treated KC mice. D–F: IHC for F4/80+ macrophages and quantification of their presence in regions of ADM and PanIN1 in KC mice either treated with mIL-13-NAB or isotype control AB. Scale bar: 100 μm. G–I: Further analyses of pSTAT1+ and Ym1+ macrophages at PanIN1 regions of KC mice either treated with mIL-13-NAB or isotype control AB using immunofluorescent labeling and quantification. Scale bar: 50 μm.
Figure 4
Figure 4. Neutralization of IL-13 decreases fibrosis in pancreatic lesions and the proliferation of PanIN lesion cells
A–C: Indicated pancreatic tissues were analyzed for presence of collagen using trichrome staining (A), smooth muscle actin (SMA) (B) or desmin (C). D–G: Indicated pancreatic tissues were analyzed for Ki-67 (D) and quantified for Ki-67+ cells per PanIN1 (E); or analyzed for expression of phospho-T202/Y204-ERK1/2 as proliferation marker (F, G). Scale bars: 50 μm.
Figure 5
Figure 5. CCL2 and IL-1ra are expressed by Ym1+ cells and induce the growth of PanIN lesions
A, B: PanIN areas from KC mice were analyzed for CCL2 (A) or IL-1ra (B) mRNA expression (brown dots) using RNAscope ISH and Ym1 expression (pink) using IHC. The arrows indicate CCL2 or IL-1ra mRNA C: Quantification of CCL2 or IL-1ra mRNA expression from pancreata from KC or control mice treated as indicated. The asterisk indicates statistical significance (p<0.05) as compared to isotype control antibody group. D–G: Detection (D, F) and quantification (E, G) of CCL2 and IL-1ra protein in pancreas tissues of KC mice treated with indicated antibodies. Scale bars: 50 μm. The asterisk indicates statistical significance (p<0.05) as compared to the isotype control antibody group. H: Ductal structures of SM3 cells seeded on Matrigel and treated with control vehicle or indicated factors (50 ng/ml) at day 2. Scale bar: 50 μm. I: Ductal area covered by n=60 structures (sorted from smallest to largest) from three independent experiments for each condition at day 2. The asterisk indicates statistical significance (p<0.0001; One-way ANOVA test). J: SM3 cells were treated as indicated for 48 hours and analyzed by immunoblotting for pERK1/2, ERK1/2 and Akt. The fold increase was determined by quantification (Image J software) of three independent experiments.

Similar articles

See all similar articles

Cited by 20 articles

See all "Cited by" articles

Publication types

Feedback