RCAN1 is a marker of oxidative stress, induced in acute pancreatitis.

BACKGROUND: To date, there still is a lack of specific acute pancreatitis markers and specifically an early marker that can reliably predict disease severity. The inflammatory response in acute pancreatitis is mediated in part through oxidative stress and calcineurin-NFAT (Nuclear Factor of Activated T-cells) signaling, which is inducing its own negative regulator, regulator of calcineurin 1 (RCAN1). Caerulein induction is a commonly used in vivo model of experimental acute pancreatitis. Caerulein induces CN-NFAT signaling, reactive oxygen species and inflammation. METHODS: To screen for potential markers of acute pancreatitis, we used the caerulein model of experimental acute pancreatitis (AP) in C57Bl/6 J mice. Pancreata from treated and control mice were used for expression profiling. Promising gene candidates were validated in cell culture experiments using primary murine acinar cells and rat AR42J cells. These candidates were then further tested for their usefulness as biomarkers in mouse and human plasma. RESULTS: We identified a number of novel genes, including Regulator of calcineurin 1 (Rcan1) and Sestrin 2 (Sesn2) and demonstrated that they are induced by oxidative stress, by stimulation with H2O2 and by inhibiting caerulein stimulated expression with the antioxidant N-acetylcysteine. We found Rcan1 protein to be significantly elevated in AP-induced mouse plasma as well as in plasma from AP patients. CONCLUSION: We demonstrated that Rcan1 is regulated by oxidative stress and identified RCAN1 as a potential diagnostic marker of AP.

Cerulein-induced pancreatic fibrosis is modulated by Smad7, the major negative regulator of transforming growth factor-ß signaling.

Chronic pancreatitis is the most common disease of the exocrine pancreas, characterized by progressive inflammation, acinar atrophy and fibrosis. Transforming growth factor-ß signaling (TGFß) is the most potent fibrogenic cytokine known, and its increased expression is a common denominator for fibrosis in chronic pancreatitis. Smad7 is induced by the TGFß superfamily members as an intracellular inhibitory feedback antagonizing TGFß signaling. To investigate the functional role of Smad7 in vivo, we induced chronic pancreatitis by repeated administration of cerulein in mice that are deficient in exon-I of Smad7. The response to chronic pancreatitis induction was significantly more severe in Smad7 mutant mice as indicated by a stronger accumulation of extracellular matrix, increased levels of inflammatory cells and an elevated number of mesenchymal cells/myofibroblasts in Smad7 mutant pancreata. Taken together, we conclude that lack of a functional Smad7 gene results in more severe damage in chronic pancreatitis. Therefore, Smad7 could be envisaged as a promising target in antifibrotic therapy of the pancreas.

Targeting of Smad7 in Mesenchymal Cells Does Not Exacerbate Fibrosis During Experimental Chronic Pancreatitis.

OBJECTIVES: Transforming growth factor-ß (TGF-ß)-mediated accumulation of extracellular matrix proteins such as collagen I is a common feature of fibrosis. Pancreatic stellate cells play an integral role in the pathogenesis of pancreatitis, and their profibrotic ability is mainly mediated by TGF-ß signaling. To specifically address the role of fibrogenic cells in experimental pancreatic fibrosis, we deleted Smad7, the main feedback inhibitor of TGF-ß signaling in this cell type in mice. METHODS: A mouse strain harboring a conditional knockout allele of Smad7 (Smad7fl/fl) with the tamoxifen-inducible inducible Col1a2-CreERT allele was generated and compared with wild-type mice challenged with the cerulein-based model of chronic pancreatitis. RESULTS: Pancreatic stellate cells lacking Smad7 had significantly increased collagen I and fibronectin production and showed a higher activation level in vitro. Surprisingly, the fibrotic index in the pancreata of treated conditional knockout mice was only slightly increased, without statistical significance. Except for fibronectin, the expression of different extracellular matrix proteins and the numbers of fibroblasts and inflammatory cells were similar between Smad7-mutant and control mice. CONCLUSIONS: There was no clear evidence that the lack of Smad7 in pancreatic stellate cells plays a major role in experimental pancreatitis, at least in the mouse model investigated here.

Real-time assessment of tissue hypoxia in vivo with combined photoacoustics and high-frequency ultrasound.

PURPOSE: In preclinical cancer studies, non-invasive functional imaging has become an important tool to assess tumor development and therapeutic effects. Tumor hypoxia is closely associated with tumor aggressiveness and is therefore a key parameter to be monitored. Recently, photoacoustic (PA) imaging with inherently co-registered high-frequency ultrasound (US) has reached preclinical applicability, allowing parallel collection of anatomical and functional information. Dual-wavelength PA imaging can be used to quantify tissue oxygen saturation based on the absorbance spectrum differences between hemoglobin and deoxyhemoglobin. EXPERIMENTAL DESIGN: A new bi-modal PA/US system for small animal imaging was employed to test feasibility and reliability of dual-wavelength PA for measuring relative tissue oxygenation. Murine models of pancreatic and colon cancer were imaged, and differences in tissue oxygenation were compared to immunohistochemistry for hypoxia in the corresponding tissue regions. RESULTS: Functional studies proved feasibility and reliability of oxygenation detection in murine tissue in vivo. Tumor models exhibited different levels of hypoxia in localized regions, which positively correlated with immunohistochemical staining for hypoxia. Contrast-enhanced imaging yielded complementary information on tissue perfusion using the same system. CONCLUSION: Bimodal PA/US imaging can be utilized to reliably detect hypoxic tumor regions in murine tumor models, thus providing the possibility to collect anatomical and functional information on tumor growth and treatment response live in longitudinal preclinical studies.