Nobuyuki Ozaki

Serine protease inhibitor Kazal type 1 promotes proliferation of pancreatic cancer cells through the epidermal growth factor receptor.

Serine protease inhibitor, Kazal type 1 (SPINK1) is expressed not only in normal human pancreatic acinar cells but also in a variety of pancreatic ductal neoplasms. There are structural similarities between SPINK1 and epidermal growth factor (EGF). Hence, we hypothesized that SPINK1 binds to EGF receptor (EGFR) to activate its downstream signaling. We first showed that SPINK1 induced proliferation of NIH 3T3 cells and pancreatic cancer cell lines. We showed that SPINK1 coprecipitated with EGFR in an immunoprecipitation experiment and that the binding affinity of SPINK1 to EGFR was about half of that of EGF using quartz-crystal microbalance (QCM) technique. As expected, EGFR and its downstream molecules, signal transducer and activator of transcription 3, v-Akt murine thymoma viral oncogene homologue, and extracellular signal-regulated kinase 1/2, were phosphorylated by SPINK1 as well as EGF. To determine which pathway is the most important for cell growth, we further analyzed the effect of inhibitors. Growth stimulation by EGF or SPINK1 was completely inhibited by EGFR and mitogen-activated protein kinase/extracellular signal-regulated kinase kinase inhibitor but not by Janus-activated kinase and phosphoinositide 3-kinase inhibitors. To further analyze the clinical importance of SPINK1 in the development of pancreatic cancer, we examined the expression of SPINK1 and EGFR in pancreatic tubular adenocarcinomas and pancreatic intraepithelial neoplasm. Both SPNK1 and EGFR were coexpressed not only in the early stage of cancer, PanIN-1A, but also in advanced stages. Taken together, these results suggest that SPINK1 stimulates the proliferation of pancreatic cancer cells through the EGFR/mitogen-activated protein kinase cascade. (Mol Cancer Res 2009;7(9):1572–81)

C/EBP homologous protein is crucial for the acceleration of experimental pancreatitis.

C/EBP homologous protein (CHOP) is one of the main mediating factors in the ER stress pathway. To elucidate the role of the ER stress-CHOP pathway in experimental pancreatitis, wild-type (Chop(+/+)) and Chop deficient (Chop(-/-)) mice were administered cerulein, a cholecystokinin analogue, or both cerulein and lipopolysaccharide (LPS). In cerulein-induced acute pancreatitis, ER stress, serum amylase elevation and histological interstitial edema were induced. However, there was no remarkable activation downstream of the CHOP pathway regardless of the presence or absence of CHOP. Whereas, in the cerulein and LPS model, inflammation-associated caspases (caspase-11, caspase-1) and IL-1beta, but not apoptosis-associated caspases, were activated. In Chop(-/-) mice, the expression levels of these mediators returned to basal levels resulting in a milder pancreatitis and decreased serum amylase level. These results indicated that the ER stress-CHOP pathway has a pivotal role in the acceleration of pancreatitis through the induction of inflammation-associated caspases and IL-1beta.

Relationship of strain-dependent susceptibility to experimentally induced acute pancreatitis with regulation of Prss1 and Spink3 expression

To analyze susceptibility to acute pancreatitis, five mouse strains including Japanese Fancy Mouse 1 (JF1), C57BL/6J, BALB/c, CBA/J, and C3H/HeJ were treated with either a cholecystokinin analog, cerulein, or a choline-deficient, ethionine-supplemented (CDE) diet. The severity of acute pancreatitis induced by cerulein was highest in C3H/HeJ and CBA/J, moderate in BALB/c, and mildest in C57BL/6J and JF1. Basal protein expression levels of the serine protease inhibitor, Kazal type 3 (Spink3) were higher in JF1 and C57BL/6J mice than those of the other three strains under normal feeding conditions. After treatment with cerulein, expression level of Spink3 increased remarkably in JF1 and mildly in C57BL/6J, BALB/c, CBA/J, and C3H/HeJ strains. Increased proteinase, serine, 1 (Prss1) protein expression accompanied by increased trypsin activity with cerulein treatment was observed in susceptible strains such as CBA/J and C3H/HeJ. Similar results were obtained with a CDE diet. In the 3 kb Spink3 promoter region, 92 or 8 nucleotide changes were found in JF1 or C3H vs C57BL/6J, respectively, whereas in the Prss1 promoter region 39 or 46 nucleotide changes were found in JF1 or C3H vs C57BL/6J, respectively. These results suggest that regulation of Prss1 and Spink3 expression is involved in the susceptibility to experimentally induced pancreatitis. The JF1 strain, which is derived from the Japanese wild mouse, will be useful to examine new mechanisms that may not be found in other laboratory mouse strains.

Significance of endothelial molecular markers in the evaluation of the severity of acute pancreatitis

PurposeIn acute pancreatitis, neutrophil elastase is secreted which damages the endothelial cells. This study was designed to demonstrate that the plasma levels of soluble E-selectin (sES) and soluble thrombomodulin (sTM) serve as endothelial molecular markers; the former is used as an endothelial activation marker, while the latter, as an endothelial injury marker.MethodsA total of 27 acute pancreatitis patients were enrolled. The plasma sES and sTM levels were assessed for 10 days after admission.ResultsThe plasma sES levels of all the patients in different disease stages were elevated at the time of admission day (day 1). The plasma sTM levels correlated with the severity and prognosis of acute pancreatitis. The required cutoff to predict a fatal outcome was set as 32 Teijin Units (TU)/ml (sensitivity, 80%; specificity, 91%). On day 1, the mortality rate of patients with the sTM levels of ≥32 TU/ml (67%, 4/6) was significantly higher than of those with the sTM levels of <32 TU/ml (5%, 1/21).ConclusionThese results indicated that (1) the activation of the vascular endothelial cells and the resultant increase in the plasma sES levels might be evoked in all disease stages, and (2) an elevation of the plasma sTM level, which indicates the presence of vascular endothelial injury, might therefore result in a poor prognosis.

SPINK1 status in colorectal cancer, impact on proliferation, and role in colitis-associated cancer

Colorectal cancer is a major cause of deaths due to cancer; therefore, research into its etiology is urgently needed. Although it is clear that chronic inflammation is a risk factor for colorectal cancer, the details remain uncertain. Serine protease inhibitor, Kazal type 1 (SPINK1) is mainly produced in pancreatic acinar cells. However, SPINK1 is expressed in various cancers and in inflammatory states, such as colon cancer and inflammatory bowel disease. There are structural similarities between SPINK1 and epidermal growth factor (EGF). Hence, it was hypothesized that SPINK1 functions as a growth factor for tissue repair in inflammatory states, and if prolonged, acts as a promoter for cell proliferation in cancerous tissues. Here, immunohistochemical staining for SPINK1 was observed in a high percentage of colorectal cancer patient specimens and SPINK1 induced proliferation of human colon cancer cell lines. To clarify its role in colon cancer in vivo, a mouse model exposed to the colon carcinogen azoxymethane and nongenotoxic carcinogen dextran sodium sulfate revealed that Spink3 (mouse homolog of SPINK1) is overexpressed in cancerous tissues. In Spink3 heterozygous mice, tumor multiplicity and tumor volume were significantly decreased compared with wild-type mice. These results suggest that SPINK1/Spink3 stimulates the proliferation of colon cancer cells and is involved in colorectal cancer progression. Implications: Evidence suggests that SPINK1 is an important growth factor that connects chronic inflammation and cancer. Mol Cancer Res; 13(7); 1130–8. ©2015 AACR.

Sister Mary Joseph’s nodule derived from pancreatic cancer

Background/PurposeUmbilical metastases known as Sister Mary Joseph’s nodules (SMJNs) occur rarely. They are an uncommon clinical or radiographic finding, and are rare as the first sign of a malignant disease. Pancreatic cancer presenting as an SMJN is a rare phenomenon, and is the source of an SMJN in 9% of cases.Case presentationWe experienced four cases of SMJN derived from pancreatic cancer. All cases originated in the tail and/or body of the pancreas.DiscussionHerein, we introduce a case with pathognomonic imaging findings. We also present the results of our review of recently published papers about SMJN, done by conducting a PubMed search.

Simultaneous total laparoscopic curative resection for synchronous gastric, cecal and rectal cancer: Report of a case

Highlights • Simultaneous total laparoscopic curative resection for synchronous GC, cecal and rectal cancer.• Our ingenious technical attempts can lead to the successful completion of simultaneous total laparoscopic curative resection.• Simultaneous laparoscopic surgery for synchronous GI cancers is a minimally invasive, feasible treatment option.

Autophagy regulation in pancreatic acinar cells is independent of epidermal growth factor receptor signaling.

Autophagy is an intracellular degradation system in eukaryotic cells that occurs at a basal level. It can also be induced in response to environmental signals including nutrients, hormones, microbial pathogens, and growth factors, although the mechanism is not known in detail. We previously demonstrated that excessive autophagy is induced within pancreatic acinar cells deficient in Spink3, which is a trypsin inhibitor. SPINK1, the human homolog of murine Spink3, has structural similarity to epidermal growth factor (EGF), and can bind and stimulate the EGF receptor (EGFR). To analyze the role of the EGFR in pancreatic development, in the regulation of autophagy in pancreatic acinar cells, and in cerulein-induced pancreatitis, we generated and examined acinar cell-specific Egfr-deficient (Egfr(-/-)) mice. Egfr(-/-) mice showed no abnormalities in pancreatic development, induction of autophagy, or cerulein-induced pancreatitis, suggesting that Egfr is dispensable for autophagy regulation in pancreatic acinar cells.

Gastrointestinal perforation during regorafenib administration in a case with hepatic metastases of colon cancer

Although common side effects of regorafenib include hand-and-foot syndrome and diarrhoea, the incidence of gastrointestinal perforation is reportedly unknown. We describe our experience with the case of a 65-year-old woman treated with regorafenib as a third-line therapy for progressive caecal cancer with multiple hepatic metastases after 4 and 6 courses of systemic mFOLFOX6 + bevacizumab (BV) and FOLFIRI + BV chemotherapy, respectively. The patient used regorafenib for 32 days but visited our hospital with abdominal pain during the second course. She was diagnosed with acute appendicitis and treated conservatively with antibiotics. The abdominal findings did not improve, and a computed tomography evaluation on day 4 of hospitalization revealed free air lateral to the caecal tumour, liver surface, and epigastric region. The patient underwent same-day emergency surgery based on a diagnosis of gastrointestinal perforation with generalized peritonitis. Upon observing digestive fluid leakage into the peri-ileocaecal area and a 5-mm perforation in the appendix, the patient was diagnosed with peritonitis due to gastrointestinal perforation. Ileocaecal resection with D2 debridement was performed, and a colostomy was opened into the ileum and ascending colon. We conclude that our patient developed gastrointestinal perforation during regorafenib therapy and note that clinicians should be aware of this possible complication in patients with a history of prior treatment with BV.

Definitive diagnosis of a duplicate gallbladder can only be made intraoperatively: report of a case

Duplicated gallbladders are rare congenital anomalies that are important in clinical practice as they may cause clinical, surgical, and diagnostic problems. Here, we describe the case of a 79-year-old female patient who presented with acute cholangitis. Abdominal ultrasonography, endoscopic ultrasonography, computed tomography, and magnetic resonance imaging revealed an intrahepatic cystic lesion, suggesting communication with the intrahepatic bile duct; no evidence of a polypoid lesion within the cystic lesion was observed. Based on these findings, intrahepatic cholangiectasis, intrahepatic bile duct cystadenoma, and the presence of a duplicated gallbladder were suspected, and surgery was performed. During surgery, a tube inserted into the common bile duct from a cystic duct facilitated intraoperative cholangiography, which indicated the presence of a duplicated gallbladder. Thus, we believe that a duplicated gallbladder should be an additional consideration when typical gallbladder disease symptoms are present under certain circumstances. A multimodal imaging approach can help to establish the diagnosis preoperatively or intraoperatively.