[IgA vasculitis with massive hemorrhage from the jejunum after steroid administration].

A 45-year-old Japanese man presenting with leg purpura, abdominal pain, and arthralgia was diagnosed with IgA vasculitis. His symptoms resolved after the intravenous administration of prednisolone. However, on day 20 of admission, he experienced bloody discharge and hypovolemic shock. The bleeding point was not identified on contrast-enhanced computed tomography scanning. The blood loss was approximately 10800ml and the patient received transfusions of 48 units of concentrated red blood cells, 18 units of fresh frozen plasma, and 30 units of concentrated platelets. Laparotomy and enteroscopy were performed through the incision of the jejunum to detect the bleeding source. Spurting bleeding was observed during the enteroscopy and partial resection of the jejunum was performed. Histopathological examination of the resected specimen revealed large vessels beneath the jejunal ulcer scar, suggesting bleeding from a Dieulafoy's lesion. Leukocytoclastic vasculitis or cytomegalovirus infection was not observed in the resected specimen. Gastrointestinal symptoms in patients with IgA vasculitis usually improve with bowel rest and conservative treatment. Administration of steroids or factor XIII is recommended for patients with severe abdominal pain refractory to conservative management. Rarely, massive bleeding, perforation, intussusception, and/or intestinal obstruction occur in the gastrointestinal tract and these complications affect patients' prognoses. The clinical course in the present patient indicated that severe bleeding from the gastrointestinal tract can occur even after symptom remission in patients with IgA vasculitis. In such cases, prompt treatment, including laparotomy and/or enteroscopy, is essential.

Preoperative Pulmonary Function Tests Predict Aspiration Pneumonia After Gastric Endoscopic Submucosal Dissection.

BACKGROUND: ESD allows higher rates of en-bloc and R0 resections, but has occasionally complications such as aspiration pneumonia. Factors associated with aspiration pneumonia are not completely understood. AIMS: To analyze the relationship between aspiration pneumonia and preoperative factors including pulmonary function tests. METHODS: A total of 978 patients with gastric tumors who had received pulmonary function tests were treated by ESD between June 2006 and May 2014. Pulmonary function tests were assessed using a spirometer. The patients were categorized into four groups according to the predicted vital capacity (%VC) and forced expiratory volume in 1 s as a percentage of forced vital capacity (FEV1.0%): normal; restrictive pulmonary dysfunction; obstructive; and mixed. The factors associated with aspiration pneumonia were retrospectively analyzed. RESULTS: Among the 268 cases with abnormal pulmonary function, 10 cases (3.7%) developed aspiration pneumonia. On the other hand, 7 cases (1.0%) with normal pulmonary function developed pneumonia. There was a significant correlation between pulmonary function and aspiration pneumonia (p = 0.010). When the pulmonary function cases were stratified into subgroups, 2.5% of cases with obstructive pulmonary dysfunction developed pneumonia, 5.5% with restrictive and 5.3% with mixed. By logistic regression analysis, pulmonary function, the presence of cerebral vascular disease, and procedure time were identified as significant independent risk factors associated with aspiration pneumonia. The odds ratios for pulmonary function, cerebral vascular disease, and procedure time were 3.6, 5.1, and 5.2, respectively. CONCLUSIONS: Preoperative pulmonary function tests may be useful markers to evaluate the risk for aspiration pneumonia after gastric ESD.

Synergistic anti-pancreatic cancer immunological effects by treatment with reduced expression in immortalized cells/dickkopf-3 protein and peripheral blood mononuclear cells.

BACKGROUND AND AIM: Reduced expression in immortalized cells/dickkopf-3 (REIC/DKK3) is a reported tumor suppressor gene and has potential to become an innovative therapy for various cancers. We examined the antitumor immunological effects of human REIC/DKK3 protein against pancreatic cancer. METHODS: Activation of extracellular signal-regulated kinases 1 and 2, mammalian target of rapamycin, and signal transducer and activator of transcription 3 by REIC/DKK3 protein was assessed in human peripheral blood mononuclear cells using immunoblotting. Pancreatic cancer cell lines (AsPC-1 and MIA Paca-2) were cocultured with peripheral blood mononuclear cells, and the anticancer effects of REIC/DKK3 protein were assessed using the methyl thiazole tetrazolium, cytotoxicity, and enzyme-linked immunospot assays. The antitumor immunological effects of the combined treatment with REIC/DKK3 protein and peripheral blood mononuclear cells were also assessed in a pancreatic cancer model using non-obese diabetic/severe combined immunodeficiency mice. RESULTS: The REIC/DKK3 protein activated extracellular signal-regulated kinases 1 and 2, mammalian target of rapamycin, and signal transducer and activator of transcription 3 in peripheral blood mononuclear cells. REIC/DKK3 protein inhibited in vitro cancer cell viability and enhanced cytotoxicity when incubated with peripheral blood mononuclear cells. REIC/DKK3 protein induced significant production of interferon gamma from lymphocytes incubated with pancreatic cancer cells, indicating that CD8+ T cells were activated in the peripheral blood mononuclear cells when cocultured with AsPC-1 and MIA Paca-2 in the presence of REIC/DKK3 protein. Combined treatment with REIC/DKK3 protein and peripheral blood mononuclear cells produced in vivo anticancer immunostimulatory effects on pancreatic cancer cells. CONCLUSIONS: The REIC/DKK3 protein and peripheral blood mononuclear cells synergistically enhanced anticancer immunological effects against pancreatic cancer cells. The observed immunomodulatory effect of combined treatment likely occurs in adenovirus-mediated REIC/DKK3 gene therapy and provides important clues to the therapeutic mechanisms involving immune cells.

Potential of adenovirus-mediated REIC/Dkk-3 gene therapy for use in the treatment of pancreatic cancer.

BACKGROUND AND AIM: The reduced expression in immortalized cells REIC/the dickkopf 3 (Dkk-3) gene, tumor suppressor gene, is downregulated in various malignant tumors. In a prostate cancer study, an adenovirus vector carrying the REIC/Dkk-3 gene (Ad-REIC) induces apoptosis. In the current study, we examined the effects of REIC/Dkk-3 gene therapy in pancreatic cancer. METHODS: REIC/Dkk-3 expression was assessed by immunoblotting and immunohistochemistry in the pancreatic cancer cell lines (ASPC1, MIAPaCa2, Panc1, BxPC3, SUIT-2, KLM1, and T3M4) and pancreatic cancer tissues. The Ad-REIC agent was used to investigate the apoptotic effect in vitro and antitumor effects in vivo. We also assessed the therapeutic effects of Ad-REIC therapy with gemcitabine. RESULTS: The REIC/Dkk-3 expression was lost in the pancreatic cancer cell lines and decreased in pancreatic cancer tissues. Ad-REIC induced apoptosis and inhibited cell growth in the ASPC1 and MIAPaCa2 lines in vitro, and Ad-REIC inhibited tumor growth in the mouse xenograft model using ASPC1 cells. The antitumor effect was further enhanced in combination with gemcitabine. This synergistic effect may be caused by the suppression of autophagy via the enhancement of mammalian target of rapamycin signaling. CONCLUSIONS: Ad-REIC induces apoptosis and inhibits tumor growth in pancreatic cancer cell lines. REIC/Dkk-3 gene therapy is an attractive therapeutic tool for pancreatic cancer.

Runt-related transcription factor 3 reverses epithelial-mesenchymal transition in hepatocellular carcinoma.

Loss or decreased expression of runt-related transcription factor 3 (RUNX3), a tumor suppressor gene involved in gastric and other cancers, has been frequently observed in hepatocellular carcinoma (HCC). The objective of this study was to identify the regulatory mechanism of the epithelial-mesenchymal transition (EMT) by RUNX3 in HCC. Human HCC cell lines, Hep3B, Huh7, HLF and SK-Hep1, were divided into low- and high-EMT lines, based on their expression of TWIST1 and SNAI2, and were used in this in vitro study. Ectopic RUNX3 expression had an anti-EMT effect in low-EMT HCC cell lines characterized by increased E-cadherin expression and decreased N-cadherin and vimentin expression. RUNX3 expression has previously been reported to reduce jagged-1 (JAG1) expression; therefore, JAG1 ligand peptide was used to reinduce EMT in RUNX3-expressing low-EMT HCC cells. Immunohistochemical analyses were performed for RUNX3, E-cadherin, N-cadherin and TWIST1 in 33 human HCC tissues, also divided into low- and high-EMT HCC, based on TWIST1 expression. E-cadherin expression was correlated positively and N-cadherin expression was correlated negatively with RUNX3 expression in low-EMT HCC tissues. Correlations between EMT markers and RUNX3 mRNA expression were analyzed using Oncomine datasets. Similarly, mRNA expression of E-cadherin was also significantly correlated with that of RUNX3 in low-EMT HCC, while mRNA expression of JAG1 was negatively correlated with that of RUNX3. These results suggest a novel mechanism by which loss or decreased expression of RUNX3 induces EMT via induction of JAG1 expression in low-EMT HCC.

[Case of von Hippel-Lindau disease diagnosed by detection of multiple pancreatic endocrine tumors and renal tumor 13 years after bilateral adrenalectomy].

von Hippel-Lindau (VHL) syndrome is an inherited neoplastic syndrome caused by abnormity of the VHL gene found on the short arm of the chromosome 3. We reported a case of VHL disease diagnosed by the detection of multiple pancreatic endocrine tumors and renal tumor 13 years after bilateral adrenalectomy. A 40-year-old man presented with multiple pancreas tumors (maximum size 42 mm in diameter) detected by screening abdominal ultrasonography. A 23 mm renal tumor was detected by contrast computed tomography scan at that time. His past history included left retinal angioma (age 15) and bilateral adrenal pheochromocytoma (age 27). VHL was diagnosed by genetic testing. Endoscopic ultrasound-guided fine-needle aspiration biopsy of the pancreas tumor was performed, and tumor was diagnosed as an endocrine tumor. After diagnosis, distal pancreatectomy (body-tail) was performed. This was a didactic case indicating that we should suspect VHL syndrome based on past history and family history and follow such cases up strictly.

[A case of duodenal gangliocytic paraganglioma with lymph node metastasis].

A 28-year-old man complained of tarry stool. A series of examinations showed a submucosal tumor with bleeding at the papilla of Vater and a swollen # 17b lymph node, both of which indicated a hypervascular tumor. The pathological findings of the enucleated tumor specimens revealed gangliocytic paraganglioma with metastasis to the # 17b lymph node. Additional pancreaticoduodenectomy revealed another # 17b lymph node metastasis 7-mm in diameter. Although the majority of gangliocytic paragangliomas are benign, 7% of reported cases have lymph node metastases, as shown in the present case. These findings are important in treating patients with gangliocytic paraganglioma.

[Clinical study of irinotecan plus infusional fluorouracil/l-leucovorin (FOLFIRI) in patients with fluoropyrimidine-resistant metastatic colorectal cancer].

The clinical efficacy and safety of irinotecan plus infusional fluorouracil/l-leucovorin (FOLFIRI) in patients with fluoropyrimidine-resistant metastatic colorectal cancer were studied retrospectively. 20 patients were treated with FOLFIRI at our hospital between October 2003 and November 2005. The objective overall response rate was 28% (5/18; 95% confidence interval, 9.7-54%). The disease control rate (CR+PR+SD) was 61%. The most frequent grade 3 to 4 adverse event was neutropenia, and it was seen in 50%. Non-hematological toxicities were mild. The median survival time was 11 months. The 1-year survival rate was 34%. This study showed the activity and safety of FOLFIRI in practice.

[A case of alpha-fetoprotein-producing minute mucosal gastric cancer].

The endoscopic examination of a 63-year-old man revealed a IIc lesion, 5 mm in diameter, in the lesser curvature of the gastric body. His serum alpha-fetoprotein (AFP) level was high at 14.3 ng/ml. Immunohistological studies on the biopsy specimens showed a positive reaction for AFP. Endoscopic ultrasonography revealed that the tumor was limited to the second layer of the gastric wall and this tumor was diagnosed as mucosal cancer. The final diagnosis was AFP-producing mucosal gastric cancer (AFPMGC). Cases of AFPMGC are rarely encountered. The present case suggested that even minute mucosal gastric cancer could produce AFP.

[Corrigendum] Loss of Runt­related transcription factor 3 induces resistance to 5­fluorouracil and cisplatin in hepatocellular carcinoma.

Following the publication of this article, we realize that there were some errors in the manuscript. Details of the experiments describing the gene silencing of RUNX3 with small interfering RNA (siRNA) were erroneously included in this paper, and all references to siRNA should have been deleted from the manuscript prior to publication. In the subsection entitled 'Cell lines and cell culture' on page 2577, the left­hand column, the text should have indicated that the human HCC cell lines Hep3B and Huh7 were obtained from the American Type Culture Collection (ATCC; Manassas, VA, USA), whereas HLF cells were obtained from the Japanese Cancer Resources Bank (Tokyo, Japan). Lastly, an error was made in describing the calculation of the IC50 values, which did not correlate with the data shown in Fig. 2. Therefore, the subsection entitled 'Ectopic RUNX3 protein expression suppresses cell growth...' should have been entitled 'Ectopic RUNX3 protein expression increases 5­FU and CDDP sensitivity', and the text herein should have read as follows: We analyzed the effects of RUNX3 on chemosensitivity in the RUNX3­ or CAT (mock)­transfected Hep3B and Huh7 cells. RUNX3 expression enhanced 5­FU sensitivity in both cell lines; the cell viability with 5­FU (100 nM) decreased from 66.3±4.6 to 34.3±5.0%, and from 71.0±4.7% to 27.0±5.5% in the Hep3B and Huh7 cells, respectively (Fig. 2A). RUNX3 expression also enhanced CDDP sensitivity in both cell lines; the cell viability with CDDP (100 nM) decreased from 58.7±2.6% to 25.7±4.9%, and from 67.7±4.1% to 25.7±7.5% in the Hep3B and Huh7 cells, respectively (Fig. 2B). We sincerely apologize for these errors and oversights, which have not affected any of the overall conclusions reported in the study, and regret any inconvenience they may have caused. [the original article was published in the Oncology Reports 35: 2576-2582, 2016; DOI: 10.3892/or.2016.4681].

[A case of gastric hamartomatous inverted polyp (submucosal tumor type) accompanied by gastric cancer].

Endoscopic examination of a 39-year-old woman revealed a flat-elevated submucosal tumor (SMT) in the upper third area of the stomach and a IIc lesion in the anterior wall of the lower third of the stomach. Endoscopic ultrasonography findings suggested an aberrant pancreas. Both lesions were resected by distal gastrectomy. Histological examination of the SMT revealed the dilated glands with cyst formation and proliferation of smooth muscle bundles in the submucosal layer, and it was diagnosed as a gastric hamartomatous inverted polyp. The IIc lesion was a signet ring cell carcinoma. Cases of gastric hamartomatous inverted polyps coexistent with gastric cancer have been rarely reported in Japan.

Loss of Runt-related transcription factor 3 induces resistance to 5-fluorouracil and cisplatin in hepatocellular carcinoma.

Runt-related transcription factor 3 (RUNX3) is known to function as a tumor suppressor in gastric cancer and other types of cancers, including hepatocellular carcinoma (HCC). However, its role has not been fully elucidated. In the present study, we aimed to evaluate the role of RUNX3 in HCC. We used the human HCC cell lines Hep3B, Huh7 and HLF; RUNX3 cDNA was introduced into Hep3B and Huh7 cells, which were negative for endogenous RUNX3 expression, and RUNX3 siRNA was transfected into HLF cells, which were positive for endogenous RUNX3. We analyzed the expression of RUNX3 and multidrug resistance-associated protein (MRP) by immunoblotting. MTT assays were used to determine the effects of RUNX3 expression on 5-fluorouracil (5-FU) and cisplatin (CDDP) sensitivity. Finally, 23 HCC specimens resected from patients with HCC at Okayama University Hospital were analyzed, and correlations among immunohistochemical expression of RUNX3 protein and MRP protein were evaluated in these specimens. Exogenous RUNX3 expression reduced the expression of MRP1, MRP2, MRP3 and MRP5 in the RUNX3-negative cells, whereas knockdown of RUNX3 in the HLF cells stimulated the expression of these MRPs. An inverse correlation between RUNX3 and MRP expression was observed in the HCC tissues. Importantly, loss of RUNX3 expression contributed to 5-FU and CDDP resistance by inducing MRP expression. These data have important implications in the study of chemotherapy resistance in HCC.

Unusual metastasis of hepatocellular carcinoma to the esophagus.

This report describes a case of metastatic hepatocellular carcinoma (HCC) presenting as a polypoid mass in the lower esophagus. The patient was a 63-year-old man with HCC. An endoscopic examination revealed a pedunculated polypoid mass, about 3 cm in diameter, at the lower part of the esophagus. The biopsy specimen obtained from the tumor revealed that the mass consisted of a pseudoglandular arrangement of tumor cells, and the tumor was diagnosed as metastatic HCC. There were no symptoms due to esophageal tumor. He died of progressive hepatic failure. Cases of premortem-diagnosed esophageal metastasis from HCC are extremely rare.

Hepatic stellate cells promote upregulation of epithelial cell adhesion molecule and epithelial-mesenchymal transition in hepatic cancer cells.

Microenvironment plays an important role in epithelial-mesenchymal transition (EMT) and stemness of cells in hepatocellular carcinoma (HCC). Epithelial cell adhesion molecule (EpCAM) is known as a tumor stemness marker of HCC. To investigate the relationship between microenvironment and stemness, we performed an in vitro co-culture assay. Four HCC cell lines (HepG2, Hep3B, HuH-7 and PLC/PRF/5) were co-cultured with the TWNT-1 immortalized hepatic stellate cells (HSCs), which create a microenvironment with HCC. Cell proliferation ability was analyzed by flow cytometry (FCM) and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, while migration ability was assessed by a wound healing assay. Expression of EpCAM was analyzed by immunoblotting and FCM. HCC cell lines were co-cultured with TWNT-1 treated with small interfering RNA (siRNA) for TGF-ß and HB-EGF; we then analyzed proliferation, migration ability and protein expression using the methods described above. Proliferation ability was unchanged in HCC cell lines co-cultured with TWNT-1. Migration ability was increased in HCC cell lines (HepG2, Hep3B, HuH-7 and PLC/PRF/5) directly (216.2±67.0, 61.0±22.0, 124.0±66.2 and 51.5±40.3%) and indirectly (102.5±22.0, 84.6±30.9, 86.1±25.7 and 73.9±29.7%) co-cultured with TWNT-1 compared with the HCC uni-culture. Immunoblot analysis revealed increased EpCAM expression in the HCC cell lines co-cultured with TWNT-1. Flow cytometry revealed that the population of E-cadherin-/N-cadherin+ and EpCAM-positive cells increased and accordingly, EMT and stemness in the HCC cell line were activated. These results were similar in the directly and indirectly co-cultured samples, indicating that humoral factors were at play. Conversely, HCC cell lines co-cultured with siRNA­treated TWNT-1 showed decreased migration ability, a decreased population of EpCAM-positive and E-cadherin-/N-cadherin+ cells. Taken together, humoral factors secreted from TWNT-1 promote upregulation of EpCAM and EMT in hepatic cancer cells.

Restored expression of the tumor suppressor gene RUNX3 reduces cancer stem cells in hepatocellular carcinoma by suppressing Jagged1-Notch signaling.

Runt-related transcription factor 3 (RUNX3) is a candidate tumor suppressor gene that is downregulated in various cancers. In the present study, we analyzed the regulatory function of RUNX3 on Jagged-1 (JAG1) expression and cancer stem cell (CSC) signaling in hepatocellular carcinoma (HCC). Eleven HCC cell lines and 30 human HCC tissues were used. RUNX3 and JAG1 expression levels were analyzed by immunoblotting and immunohistochemistry. Ectopic RUNX3 expression was induced by introducing RUNX3 cDNA into the RUNX3-negative HCC cell line Hep3B and Huh7 cells. Furthermore endogenous RUNX3 expression was knocked down by RUNX3 siRNA in SK-Hep-1 cells. In order to analyze JAG1 transcriptional regulation, we conducted reporter assays, chromatin immunoprecipitation (ChIP) assays and electrophoretic mobility shift assays (EMSAs). Tumorigenicity was analyzed using a SCID mouse liver injection model. An inverse correlation was observed between RUNX3 expression and JAG1 expression in most HCC cell lines and tissues. Restoring RUNX3 expression decreased the expression of JAG1 in Hep3B and Huh7 cells, whereas JAG1 expression was upregulated in RUNX3 siRNA-treated SK-Hep-1 cells. Reporter assays, ChIP assays and EMSAs revealed that RUNX3 directly bound to the transcriptional regulatory region of JAG1 and suppressed JAG1 transcription. Moreover, RUNX3 restoration downregulated CSCs by suppressing JAG1-mediated Notch signaling. The tumorigenic capacity of RUNX3-expressing Hep3B cells was lower compared to that of control Hep3B cells. RUNX3 expression suppressed JAG1 expression and resulted in downregulation of tumorigenesis by suppression of JAG1-mediated CSCs.