MiR-146b-3p protects against AR42J cell injury in cerulein-induced acute pancreatitis model through targeting Anxa2

Background Acute pancreatitis (AP) is a common inflammatory disorder. MicroRNAs play crucial roles in the pathogenesis of AP. In this article, we explored the detailed role and molecular mechanisms of miR-146b-3p in AP progression. Methods The rat AR42J cells were treated with cerulein to establish the AP model in vitro. The miR-146b-3p and Annexin A2 (Anxa2) mRNA levels were assessed by quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR). Cell viability and apoptosis were tested using the Cell Counting Kit-8 (CCK-8) and flow cytometry assays, respectively. Caspase-3 activity and the production of interleukin-1β (IL-1β), IL-6, and tumor necrosis factor-α (TNF-α) were measured by enzyme-linked immunosorbent assay and qRT-PCR. Targeted interaction between miR-146b-3p and Anxa2 was verified by the dual-luciferase reporter and RNA immunoprecipitation assays. Western blot analysis was performed to detect the expression of Anxa2 protein. Results Our data revealed that miR-146b-3p was significantly downregulated in AP samples. The enforced expression of miR-146b-3p alleviated cerulein-induced injury in AR42J cells, as evidenced by the promotion in cell viability and the repression in cell apoptosis, as well as the reduction in IL-1β, IL-6, and TNF-α production. Anxa2 was directly targeted and inhibited by miR-146b-3p. Moreover, the alleviative effect of miR-146b-3p overexpression on cerulein-induced AR42J cell injury was mediated by Anxa2. Conclusions The current work had led to the identification of miR-146b-3p overexpression that protected against cerulein-induced injury in AR42J cells at least in part by targeting Anxa2, revealing a promising target for AP diagnosis and treatment.

Low Expression of Mas1 Receptor in Pancreas Related with Acute Pancreatitis

Background：Acute pancreatitis (AP) continues to be one of the most common causes of hospitalization among all gastrointestinal disease. There is a lack of therapies directed to its molecular pathogenesis. The aim of this study was to investigate the role and major mechanisms of Mas1 receptor in AP. Methods: AP was induced in C57BL/6 mice by administration of caerulein and lipopolysaccharide. The effects of intervening Mas1 receptor on the severity of AP, trypsinogen activation peptide (TAP), zymogens distribution and autophagy activity were detected in vivo. Then Mas1-lentivirus transfected AR42J cell and incubated with caerulein. TAP, autophagy activity and co-location of trypsin and autophagic were further detected in vitro. Human pancreas tissue from patients with AP and without AP were also used for Mas1 receptor protein and mRNA expression levels.Results: Mas1 receptor proteins were down-regulated in AP mice pancreas tissue as well as in human patients with AP. Compared with Mas1 receptor inhibited in AP mice, AP mice with Mas1 receptor activated decreased severe pathological changes in pancreata, lower levels of trypsinogen activation concomitant with zymogens basolateral distribution, and autophagy down-regulation. Mas1 receptor knockdown and over-expression further verified those results in vitro and showed Mas1 receptor knockdown in AR42J cells had an increased colocalization of LC3II with trypsin.Conclusions: Mas1 receptor decreased expresssion in pancreas may promote zymogens premature activation relating to AP.

Melatonin Induces Anti-Inflammatory Effects to Play a Protective Role via Endoplasmic Reticulum Stress in Acute Pancreatitis

Background/Aims: Melatonin, which is mainly secreted by the pineal gland and released into blood, has anti-inflammatory properties in acute pancreatitis. Many studies show that melatonin can relieve inflammation in taurocholate-induced acute pancreatitis. However, the mechanisms of its anti-inflammatory effects are still undefined, especially the relationship between melatonin and endoplasmic reticulum stress. We explored the anti-inflammatory activity of melatonin in AR42J and rat models. Methods: The CCK-8 assay was used to assess effects of melatonin on AR42J cell viability. Inflammatory degree and the expressions of endoplasmic reticulum stress related molecules were examined by quantitative RT-PCR and western blotting. The degree of inflammation in the tissue was also accessed by pathological grading. Finally, we used the western blotting method to verify apoptosis and autophagy. Results: Endoplasmic reticulum stress was obviously activated in early stage inflammation in AR42J and rat models. Melatonin could induce anti-inflammatory effects via endoplasmic reticulum stress. Melatonin significantly inhibited inflammatory cytokines and the expression of ERS-related molecules. Finally, it played a protective role by promoting apoptosis and autophagy of the cells, which were damaged in the process of inflammatory reaction. Conclusion: Melatonin induces anti-inflammatory effects via endoplasmic reticulum stress in acute pancreatitis to play a protective role.

Ptf1a Binds to and Activates Area III, a Highly Conserved Region of the Pdx1 Promoter That Mediates Early Pancreas-Wide Pdx1 Expression

ABSTRACT The critical pancreatic transcription factor Pdx1 is expressed throughout the pancreas early but enriched in insulin-producing β cells postnatally. Previous studies showed that the 5′ conserved promoter regions areas I and II (Pdx1 PB) direct endocrine cell expression, while an adjacent region (Pdx1 XB) containing conserved area III directs transient β-cell expression. In this study, we used Cre-mediated lineage tracing to track cells that activated these regions. Pdx1 PBCre mediated only endocrine cell recombination, while Pdx1 XBCre directed broad and early recombination in the developing pancreas. Also, a reporter transgene containing areas I, II, and III was expressed throughout the embryonic day 10.5 (E10.5) pancreas and gradually became β cell enriched, similar to endogenous Pdx1. These data suggested that sequences within area III mediate early pancreas-wide Pdx1 expression. Area III contains a binding site for PTF1, a transcription factor complex essential for pancreas development. This site contributed to area III-dependent reporter gene expression in the acinar AR42J cell line, while PTF1 specifically trans-activated area III-containing reporter expression in a nonpancreatic cell line. Importantly, Ptf1a occupied sequences spanning the endogenous PTF1 site in area III of E11.5 pancreatic buds. These data strongly suggest that PTF1 is an important early activator of Pdx1 in acinar and endocrine progenitor cells during pancreas development.