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Noggin attenuates cerulein-induced acute pancreatitis and impaired autophagy.

ABSTRACT: To investigate the role of bone morphogenetic protein (BMP) signaling in acute pancreatitis (AP) by administration of noggin, an endogenous BMP antagonist, in a cerulein-induced AP model.Acute pancreatitis was induced by 9 hourly intraperitoneal injections of cerulein (50 ?g/kg). Control mice received phosphate-buffered saline injections. In a separate group, noggin (0.5 mg/kg) was given intraperitoneally at 1 hour before and 2, 4, and 6 hours after AP induction. The mice were euthanized at 1 hour after completion of AP induction. The blood samples and the pancreas were harvested for analysis. Isolated pancreatic acini from normal mice and AR42J cells were treated with BMP2 and cerulein. AR42J cells were also treated with noggin. Phosphorylation of Smad1/5/8 was measured.Bone morphogenetic protein signaling was up-regulated in AP mouse pancreas. Bone morphogenetic protein 2 and cerulein-induced phosphorylation of Smad1/5/8 in the acinar cells in vitro, which was blocked by noggin. Noggin administration in vivo attenuated AP induction, decreased vacuole formation in acinar cells, blocked LC3-II levels, and partially restored Beclin-1 and lysosomal-associated membrane protein 2 levels.Bone morphogenetic protein signaling seems to promote AP induction and autophagy, as suggested by our study showing that noggin ameliorates AP and partially restores autophagic homeostasis.

SUBMITTER: Cao Y

PROVIDER: S-EPMC3894737 | biostudies-literature | 2013 Mar

REPOSITORIES: biostudies-literature

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Noggin attenuates cerulein-induced acute pancreatitis and impaired autophagy.

Cao Yanna Y Yang Wenli W Tyler Matthew A MA Gao Xuxia X Duan Chaojun C Kim Sung O SO Aronson Judith F JF Popov Vsevolod V Takahashi Hitoshi H Saito Hiroshi H Evers Bernard Mark BM Chao Celia C Hellmich Mark R MR Ko Tien C TC

Pancreas 20130301 2

<h4>Objectives</h4>To investigate the role of bone morphogenetic protein (BMP) signaling in acute pancreatitis (AP) by administration of noggin, an endogenous BMP antagonist, in a cerulein-induced AP model.<h4>Methods</h4>Acute pancreatitis was induced by 9 hourly intraperitoneal injections of cerulein (50 μg/kg). Control mice received phosphate-buffered saline injections. In a separate group, noggin (0.5 mg/kg) was given intraperitoneally at 1 hour before and 2, 4, and 6 hours after AP inductio ...[more]

PMID: 22850625

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Project description:BackgroundAR42J are immortalized pancreatic adenocarcinoma cells that share similarities with pancreatic acinar cells. AR42J are often used as a cell-culture model of cerulein (CN)-induced acute pancreatitis (AP). Nevertheless, it is controversial how to treat AR42J for reliable induction of AP-like processes. Gene knockout and/or overexpression often remain challenging, as well. In this study, we demonstrate conditions for a reliable induction of proinflammatory markers upon CN treatment in AR42J and high transfection efficacy using Glyoxalase-I (Glo-I) as a target of interest.MethodsEffects of dexamethasone (dexa) and CN on cell morphology and amylase secretion were analyzed via ELISA of supernatant. IL-6, TNF-α and NF-κB-p65 were measured via qRT-PCR, ELISA and Western Blot (WB). Transfection efficacy was determined by WB, qRT-PCR and immune fluorescence of pEGFP-N1-Glo-I-Vector and Glo-I-siRNA.ResultsTreatment of AR42J with 100 nm dexa is mandatory for differentiation to an acinar-cell-like phenotype and amylase production. CN resulted in secretion of amylase but did not influence amylase production. High levels of CN-induced amylase secretion were detected between 3 and 24 hours of incubation. Treatment with LPS alone or in combination with CN did not influence amylase release compared to control or CN. CN treatment resulted in increased TNF-α production but not secretion and did not influence IL-6 mRNA. CN-induced stimulation of NF-κB was found to be highest on protein levels after 6h of incubation. Transient transfection was able to induce overexpression on protein and mRNA levels, with highest effect after 12 to 24 hours. Gene-knockdown was achieved by using 30 pmol of siRNA leading to effective reduction of protein levels after 72 hours. CN did not induce amylase secretion in AR42J cell passages beyond 35.ConclusionAR42J cells demonstrate a reliable in-vitro model of CN-induced AP but specific conditions are mandatory to obtain reproducible data.

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Noggin attenuates cerulein-induced acute pancreatitis and impaired autophagy.

Publications

Noggin attenuates cerulein-induced acute pancreatitis and impaired autophagy.

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