The predominance of a naive T helper cell subset in the immune response of experimental acute pancreatitis.

INTRODUCTION: In necrotizing acute pancreatitis (NAP), systemic inflammatory response syndrome (SIRS) and the compensatory anti-inflammatory response syndrome (CARS) decide overall outcome and mortality. In patients, low lymphocyte counts were found, but T-helper cells seemed to conversely increase. Our aim was to further categorize T-helper cells within the context of NAP induced SIRS and CARS. METHODS: NAP was induced by injection of sodium-taurocholate into the common bile duct of male BALB/c mice; sham treated animals received saline infusion. The animals were sacrificed at 6, 12, 24 and 48 h later. Lymphocytes from blood, liver and spleen were isolated and examined by flow cytometry. Staining was performed for CD4, CD8, CD19, CD45RB, CD25, CD69, and CD152. CD4+ cells were sorted for their CD45RB expression and sought for gene regulation associated to TH1/TH2 cells by quantitative RT-PCR. RESULTS: In NAP, CD4+ was solely increased in all compartments. CD8+ remained without substantial alterations. CD45RB showed significant expression in RBhigh in T-helper cells, confirmed by the CD45RBhigh/low ratio (Liver, 24 h: NAP 2.2, SHAM 0.6; p < 0.001). CD45RBhigh and -low cells were not associated to patterns of TH1/TH2 expression. In NAP, CCR4 expression was significantly decreased within RBhigh cells (fold change: 0.04, p < 0.05), while TLR6 showed significant overexpression (fold change: 2.36, p < 0.05). CONCLUSION: T-helper cells increase in NAP, leaning towards CD45RBhigh expression. They resemble naive T-cells, in which NAP leads to expression profiles associated with an innate immune response. This suggests new findings in immunological pathomechanisms of NAP.

The chemokine ligand CXCL16 is an indicator of bacterial infection in necrotizing pancreatitis.

OBJECTIVES: Current guidelines tell us that intervention in severe necrotizing pancreatitis ought to be performed as late as possible. However, when pancreatic necrosis becomes infected, the necrotic tissue needs to be removed. Unfortunately, bacterial infection can only be proven by invasive methods. METHODS: Necrotizing pancreatitis with sterile or infected necrosis was induced in mice. Mice serum samples were examined by antibody-based protein array. After identifying candidate proteins that showed strong regulation, the serum concentration of these proteins was examined by sandwich ELISA. Then, human serum samples were collected from patients with mild pancreatitis, severe pancreatitis with and without pancreatic necrosis and patients with microbiologically proven infection of pancreatic necrosis. These serum samples were then analyzed by sandwich ELISA. RESULTS: In mice 6 proteins were strongly up-regulated and were further investigated by ELISAs. Of these proteins, CXCL16 and TRANCE (RANKL) concentrations were analyzed in human serum samples. CXCL16 and TRANCE were increased in patients with pancreatic necrosis and abdominal infection. Receiver operated characteristics showed that CXCL16 was superior in predicting infected pancreatic necrosis when compared to C-reactive protein and TRANCE. CONCLUSIONS: Serum CXCL16 is increased in severe pancreatitis with infected pancreatic necrosis and identifies patients who benefit from surgical necrosectomy.

Organ-specific monocyte activation in necrotizing pancreatitis in mice.

BACKGROUND: Acute necrotizing pancreatitis (NAP) induces a systemic inflammatory response syndrome. We investigated the underlying changes of monocytes using different activation markers. MATERIALS AND METHODS: A retrograde injection of 2 mL/kg bodyweight of sodium taurocholate into the common bile duct of BALB/c mice induced NAP, whereas sham-operated animals (SOP) were treated with saline. After 6, 12, 24, and 48 h, histologic alterations, pancreatic enzymes, and interleukin 6 in serum, albumin, and myeloperoxidase (MPO) in bronchoalveolar lavage fluid were examined. Isolation of mononuclear cells from the blood, spleen, and liver and the subsequent determination of macrophages (F4/80) and their activation marker CD121b and MHCII (1Ad) were performed by fluorescence-activated cell sorting (FACS analyses). RESULTS: After pancreatitis induction, pancreatic enzymes (amylase: SOP 7008 U/L, NAP 37,044 U/L, P < 0.001) and histologic pancreatic damage (SOP 0.80 ± 1.92, NAP 19.6 ± 0.64, P < 0.001) developed instantly. Pulmonary vascular damage and MPO were detected between 6 and 12 h after onset (6 h albumin SOP 132.0 ± 12.0 µg/mL, NAP 267.2 ± 49.6 µg/mL; P < 0.05; MPO SOP 0.23 ± 0.20 ng/mL, NAP 11.29 ± 3.12 ng/mL, P < 0.01). Blood levels of interleukin 6 increased after 12-24 h (12 h SOP 584 ± 300 pg/mL; NAP 2169 ± 942 pg/mL, P < 0.05), whereas monocytes increased fourfold within 48 h (P < 0.05). Furthermore, pancreatitis increased the percentage of activated monocytes in the blood (6 h and/or 48 h: MHCII (1Ad) 2196%/5.65%; CD121b 51,654%/82,146%). Similar observations were made for monocytes from the liver and spleen. CONCLUSIONS: Although inflammatory mediators increased during 24 h after pancreatitis induction, monocyte activation lasted for at least 48 h. The latter is not limited to blood but also detected in isolated liver and spleen monocytes.

Metformin Decreases the Incidence of Pancreatic Ductal Adenocarcinoma Promoted by Diet-induced Obesity in the Conditional KrasG12D Mouse Model.

Pancreatic ductal adenocarcinoma (PDAC) is a particularly deadly disease. Chronic conditions, including obesity and type-2 diabetes are risk factors, thus making PDAC amenable to preventive strategies. We aimed to characterize the chemo-preventive effects of metformin, a widely used anti-diabetic drug, on PDAC development using the KrasG12D mouse model subjected to a diet high in fats and calories (HFCD). LSL-KrasG12D/+;p48-Cre (KC) mice were given control diet (CD), HFCD, or HFCD with 5 mg/ml metformin in drinking water for 3 or 9 months. After 3 months, metformin prevented HFCD-induced weight gain, hepatic steatosis, depletion of intact acini, formation of advanced PanIN lesions, and stimulation of ERK and mTORC1 in pancreas. In addition to reversing hepatic and pancreatic histopathology, metformin normalized HFCD-induced hyperinsulinemia and hyperleptinemia among the 9-month cohort. Importantly, the HFCD-increased PDAC incidence was completely abrogated by metformin (p < 0.01). The obesogenic diet also induced a marked increase in the expression of TAZ in pancreas, an effect abrogated by metformin. In conclusion, administration of metformin improved the metabolic profile and eliminated the promoting effects of diet-induced obesity on PDAC formation in KC mice. Given the established safety profile of metformin, our findings have a strong translational potential for novel chemo-preventive strategies for PDAC.