The novel Aryl hydrocarbon receptor inhibitor biseugenol inhibits gastric tumor growth and peritoneal dissemination.

Biseugenol (Eug) is known to antiproliferative of cancer cells; however, to date, the antiperitoneal dissemination effects have not been studied in any mouse cancer model. In this study, Aryl hydrocarbon receptor (AhR) expression was associated with lymph node and distant metastasis in patients with gastric cancer and was correlated with clinicolpathological pattern. We evaluated the antiperitoneal dissemination potential of knockdown AhR and Biseugenol in cancer mouse model and assessed mesenchymal characteristics. Our results demonstrate that tumor growth, peritoneal dissemination and peritoneum or organ metastasis implanted MKN45 cells were significantly decreased in shAhR and Biseugenol-treated mice and that endoplasmic reticulum (ER) stress was caused. Biseugenol-exposure tumors showed acquired epithelial features such as phosphorylation of E-cadherin, cytokeratin-18 and loss mesenchymal signature Snail, but not vimentin regulation. Snail expression, through AhR activation, is an epithelial-to-mesenchymal transition (EMT) determinant. Moreover, Biseugenol enhanced Calpain-10 (Calp-10) and AhR interaction results in Snail downregulation. The effect of shCalpain-10 in cancer cells was associated with inactivation of AhR/Snail promoter binding activity. Inhibition of Calpain-10 in gastric cancer cells by short hairpin RNA or pharmacological inhibitor was found to effectively reduced growth ability and vessel density in vivo. Importantly, knockdown of AhR completed abrogated peritoneal dissemination. Herein, Biseugenol targeting ER stress provokes Calpain-10 activity, sequentially induces reversal of EMT and apoptosis via AhR may involve the paralleling processes. Taken together, these data suggest that Calpain-10 activation and AhR inhibition by Biseugenol impedes both gastric tumor growth and peritoneal dissemination by inducing ER stress and inhibiting EMT.

Tpl2 inhibitors thwart endothelial cell function in angiogenesis and peritoneal dissemination.

Angiogenesis is critical in the development of cancer, which involves several angiogenic factors in its peritoneal dissemination. The role of protein tumor progression locus 2 (Tpl2) in angiogenic factor-related endothelial cell angiogenesis is still unclear. To understand the precise mechanism(s) of Tpl2 inhibition in endothelial cells, this study investigated the role of Tpl2 in mediating angiogenic signals using in vitro, in vivo, and ex vivo models. Results showed that inhibition of Tpl2 inhibitor significantly reduced peritoneal dissemination in a mouse model by positron emission tomography/computed tomography imaging. Simultaneously, inhibiting Tpl2 blocked angiogenesis in tumor nodules and prevented angiogenic factor-induced proliferating cell nuclear antigen (PCNA) in endothelial cells. Vascular endothelial growth factor (VEGF) or chemokine (C-X-C motif) ligand 1 (CXCL1) increased Tpl2 kinase activity and phosphorylation in a dose- and time-dependent manner. Furthermore, Tpl2 inhibition or ablation by siRNA prevented the angiogenic signal-induced tube formation in Matrigel plug assay or aortic ring assay. Inhibiting Tpl2 also prevented the angiogenic factor-induced chemotactic motility and migration of endothelial cells. Tpl2 inhibition by CXCL1 or epidermal growth factor in endothelial cells was associated with inactivation of CCAAT/enhancer binding protein ß, nuclear factor κ light-chain enhancer of activated B cells, and activating protein 1 and suppression of VEGF expression. Thus, Tpl2 inhibitors thwart Tpl2-regulated VEGF by inactivating transcription factors involved in angiogenic factor-triggered endothelial cell angiogenesis. These results suggest that the therapeutic inhibition of Tpl2 may extend beyond cancer and include the treatment of other diseases involving pathologic angiogenesis.

Honokiol thwarts gastric tumor growth and peritoneal dissemination by inhibiting Tpl2 in an orthotopic model.

Honokiol is known to suppress the growth of cancer cells; however, to date, its antiperitoneal dissemination effects have not been studied in an orthotopic mouse model. In the present study, we evaluated the antiperitoneal dissemination potential of Honokiol in an orthotopic mouse model and assessed associations with tumor growth factor-ß1 (TGFß1) and cells stimulated by a carcinogen, N-methyl-N'-nitro-N-nitrosoguanidine (MNNG). Our results demonstrate that tumor growth, peritoneal dissemination and peritoneum or organ metastasis of orthotopically implanted MKN45 cells were significantly decreased in Honokiol-treated mice and that endoplasmic reticulum (ER) stress was induced. Honokiol-treated tumors showed increased epithelial signatures such as E-cadherin, cytokeratin-18 and ER stress marker. In contrast, decreased expression of vimentin, Snail and tumor progression locus 2 (Tpl2) was also noted. TGFß1 and MNNG-induced downregulation of E-cadherin and upregulation of Tpl2 were abrogated by Honokiol treatment. The effect of Tpl2 inhibition in cancer cells or endothelial cells was associated with inactivation of CCAAT/enhancer binding protein B, nuclear factor kappa-light-chain-enhancer of activated B cell and activator protein-1 and suppression of vascular endothelial growth factor. Inhibition of Tpl2 in gastric cancer cells by small interfering RNA or pharmacological inhibitor was found to effectively reduce growth ability and vessel density in vivo. Honokiol-induced reversal of epithelial-to-mesenchymal transition (EMT) and ER stress-induced apoptosis via Tp12 may involve the paralleling processes. Taken together, our results suggest that the therapeutic inhibition of Tpl2 by Honokiol thwarts both gastric tumor growth and peritoneal dissemination by inducing ER stress and inhibiting EMT.

The thermostable direct hemolysin from Grimontia hollisae causes acute hepatotoxicity in vitro and in vivo.

BACKGROUND: G. hollisae thermostable direct hemolysin (Gh-TDH) is produced by most strains of G. hollisae. This toxin has been reported to be absorbed in the intestines in humans. Secondary liver injury might be caused by venous return of the toxin through the portal system. We aimed to firstly analyze the in vitro and in vivo hepatotoxicity of Gh-TDH. METHODS: Liver cells (primary human non-cancer cell and FL83B mouse cells) were treated and mice (BALB/c) were fed with this toxin to investigate its hepatotoxicity. Morphological examination and cytotoxicity assays using liver cells were also performed. Fluorescein isothiocyanate-conjugated toxin was used to analyze the localization of this protein in liver cells. Mice were subjected to liver function measurements and liver biopsies following toxin treatment and wild-type bacterial infection. PET (positron emission tomography)/CT (computed tomography) images were taken to assess liver metabolism during acute injury and recovery. RESULTS: The effect of hepatotoxicity was dose and time dependent. Cellular localization showed that the toxin was initially located around the cellular margins and subsequently entered the nucleus. Liver function measurements and liver biopsies of the mice following treatment with toxin or infection with wild-type Grimontia hollisae showed elevated levels of transaminases and damage to the periportal area, respectively. The PET/CT images revealed that the reconstruction of the liver continued for at least one week after exposure to a single dose of the toxin or bacterial infection. CONCLUSIONS: The hepatotoxicity of Gh-TDH was firstly demonstrated. The damage was located in the periportal area of the liver, and the liver became functionally insufficient.

Calpain/SHP-1 interaction by honokiol dampening peritoneal dissemination of gastric cancer in nu/nu mice.

BACKGROUND: Honokiol, a small-molecular weight natural product, has previously been reported to activate apoptosis and inhibit gastric tumorigenesis. Whether honokiol inhibits the angiogenesis and metastasis of gastric cancer cells remains unknown. METHODOLOGY/PRINCIPAL FINDINGS: We tested the effects of honokiol on angiogenic activity and peritoneal dissemination using in vivo, ex vivo and in vitro assay systems. The signaling responses in human gastric cancer cells, human umbilical vascular endothelial cells (HUVECs), and isolated tumors were detected and analyzed. In a xenograft gastric tumor mouse model, honokiol significantly inhibited the peritoneal dissemination detected by PET/CT technique. Honokiol also effectively attenuated the angiogenesis detected by chick chorioallantoic membrane assay, mouse matrigel plug assay, rat aortic ring endothelial cell sprouting assay, and endothelial cell tube formation assay. Furthermore, honokiol effectively enhanced signal transducer and activator of transcription (STAT-3) dephosphorylation and inhibited STAT-3 DNA binding activity in human gastric cancer cells and HUVECs, which was correlated with the up-regulation of the activity and protein expression of Src homology 2 (SH2)-containing tyrosine phosphatase-1 (SHP-1). Calpain-II inhibitor and siRNA transfection significantly reversed the honokiol-induced SHP-1 activity. The decreased STAT-3 phosphorylation and increased SHP-1 expression were also shown in isolated peritoneal metastatic tumors. Honokiol was also capable of inhibiting VEGF generation, which could be reversed by SHP-1 siRNA transfection. CONCLUSIONS/SIGNIFICANCE: Honokiol increases expression and activity of SPH-1 that further deactivates STAT3 pathway. These findings also suggest that honokiol is a novel and potent inhibitor of angiogenesis and peritoneal dissemination of gastric cancer cells, providing support for the application potential of honokiol in gastric cancer therapy.

Detection of gastroesophageal reflux esophagitis using 2-fluoro-2-deoxy-d-glucose positron emission tomography.

BACKGROUND: Gastroesophageal reflux disease (GERD) is a common disease and a major upper gastrointestinal problem. The purpose of the present study is to evaluate the use of noninvasive 2-fluoro-2-deoxy-d-glucose positron emission tomography (FDG-PET) to detect gastroesophageal reflux esophagitis. MATERIALS AND METHODS: This is a retrospective study reviewing 408 healthy check-up subjects (169 females and 239 men), who underwent both FDG-PET and upper gastrointestinal endoscopy during September 2008 to December 2009. Quantitative analysis of FDG uptake in the distal part of the esophagus was performed by calculating the maximum standard uptake value (SUVmax). This indicated the degree of esophagitis. FDG-PET findings were compared with endoscopic (modified version of the Los Angeles classification) diagnoses as the gold standard. RESULTS: The SUVmax ranged from 1.30 to 3.40 in normal subjects and from 1.30 to 4.00 in subjects with gastroesophageal reflux esophagitis. In the esophagitis group, the SUVmax was 2.13 ± 0.42 in subjects with modified LA grade M, 2.21 ± 0.45 in subjects with LA grade A, and 2.48 ± 0.44 in subjects with LA grade B and C gastroesophageal reflux esophagitis. One-way ANOVA and post-hoc comparison with Bonferroni correction (P value = 0.003) identified statistical differences between the three groups. CONCLUSION: Noninvasive FDG-PET may be useful in the detection and evaluation of various degrees of gastroesophageal reflux esophagitis.

Unexpected accumulation of F-18 FDG in the urinary bladder after bladder irrigation and retrograde filling with sterile saline: a possible pitfall in PET examination.

BACKGROUND: : Bladder irrigation and retrograde filling technique has been used to reduce urinary F-18 FDG (FDG) activity for better image interpretation in patients with pelvic tumors. Despite the zealous use of this technique, FDG accumulation in the urinary bladder has been reported and might cause false-positive or false-negative results. In this study, we analyzed the pattern and estimated the incidence of the unexpected accumulation of urinary FDG activity after bladder irrigation and retrograde filling with sterile saline. METHODS: : We reviewed 53 patients with pelvic malignancies who underwent FDG-PET scans. All of them had bladder irrigation and retrograde filling with sterile saline via triple lumen Foley Catheters. If there was high FDG activity, either focal or diffuse, in the urinary bladder on the standard 1-hour image, bladder irrigation and retrograde filling were repeated and the delayed 3-hour images were acquired. If delayed 3-hour scanning was not possible due to time restraint or the patient's inability to cooperate, pelvic images were immediately acquired while the patient changed to the prone position. RESULTS: : On the 1-hour images, 43 of the 53 (81.1%) patients showed low urinary FDG activities and satisfactory image quality for interpretation. However, 10 (18.9%) patients had high FDG activity in the urinary bladder, which could interfere with pelvic image interpretation. Of these 10 patients, 3 (5.7%) patients had diffuse FDG activity in the urinary bladder, which could obliterate a nearby FDG-avid lesion. All of the diffuse urinary FDG activity was cleared with repeated bladder irrigation and retrograde filling. The remaining 7 (13.2%) patients had focal FDG-accumulation in the bladder, which might cause false-positive result. Five of them had repeated bladder irrigation and retrograde filling and all of the focal FDG-accumulations disappeared on the delayed images. Bladder irrigation and retrograde filling could not be performed in 2 patients and pelvic images were acquired after the patients changed to the prone position. The urinary FDG accumulation changed from the posterior portion of the bladder in the supine position to the anterior portion of the bladder in the prone position. CONCLUSION: : Unexpected FDG activity in the urinary bladder, either focal or diffuse, may occur after bladder irrigation and retrograde filling and interfere with pelvic image interpretation. Great caution is required to avoid misdiagnosis: focal FDG accumulation may mimic tumor uptake of FDG whereas diffuse FDG activity may obliterate a FDG-avid pelvic lesion. Repeated bladder irrigation and retrograde filling, and prone-position imaging are useful techniques to ascertain the nature of the FDG accumulation.

Calyceal diverticulum in FDG-PET imaging.

A 46-year-old female underwent a fluorodeoxyglucose F-18 (FDG) positron emission tomography (PET) whole-body scan for tumor screening after showing no symptoms or signs. The PET images showed a large FDG-avid lesion in the left renal pelvis, with a maximum standard uptake value (SUV) of 10.7 at 1 hour and 3.9 at 3 hours. An abdominal computed tomography (CT) scan showed a 3.7-cm cystic lesion with a slightly irregular wall in the left kidney. The immediate impression was that of a complicated cyst. A histopathologic report after surgery confirmed a calyceal diverticulum. To the best of our knowledge, this is the first report of FDG uptake in a case of calyceal diverticulum.