Recognition of endoscopic diagnosis in differentiated-type early gastric cancer by flexible spectral imaging color enhancement with indigo carmine.

BACKGROUND/AIMS: To evaluate the usefulness of flexible spectral imaging color enhancement with indigo carmine (I-FICE) in early gastric cancer (EGC) demarcation. METHODS: The study participants were 29 patients with differentiated-type EGC. The endoscope was fixed and images of the same area of EGC demarcations in each lesion were obtained using four different methods (WLE, flexible spectral imaging color enhancement (FICE), CE, and I-FICE). FICE mode at R 550 nm (Gain: 2), G 500 nm (Gain: 4), and B 470 nm (Gain: 4) was used. Four endoscopists ranked the images obtained by each method on the basis of the ease of recognition of demarcation using a 4-point system. We calculated the standard deviation of pixel values based on L*, a*, and b* color spaces in the demarcation region (Lab-SD score). RESULTS: The median ranking score for I-FICE images was significantly higher than that obtained from the other methods. Further, the average Lab-SD score was significantly higher for I-FICE images than for images obtained by the other methods. There was a good correlation between the ranking score and Lab-SD score. CONCLUSION: EGC demarcations were most easily recognized both subjectively and objectively using I-FICE image, followed by CE, FICE and WLE images.

Interleukin-8 production via protease-activated receptor 2 in human esophageal epithelial cells.

Interaction between proteases and protease-activated receptor (PAR) 2 has been proposed to mediate inflammatory and immune response in the gastrointestinal tract. Recently, increase in interleukin (IL)-8 in the esophageal mucosa has been associated with the pathogenesis of esophagitis induced by reflux of gastric acids, bile acids or trypsin. The aims of the present study were to determine PAR2 expression in normal human esophageal epithelial cells (HEEC) and to evaluate the mediation of IL-8 production by trypsin-PAR2 interaction in HEEC. Reverse transcription polymerase chain reaction (RT-PCR) and Western blot analysis revealed that PAR2 mRNA and protein were constitutively expressed in HEEC without upregulation by the stimulation with tumor necrosis factor alpha or trypsin. IL-8 was produced in a dose-dependent fashion when cells were stimulated with a PAR2 agonist such as trypsin or SLIGKV-amide. Blocking antibody to PAR2, camostat mesilate (a trypsin inhibitor), p-38 mitogen-activated protein kinase (MAPK) inhibitors or ERK1/2 inhibitors reduced IL-8 production from trypsin-stimulated HEEC. Mutation of the NFkappaB-, AP-1- and NF-IL-6-binding site on the IL-8 gene promoter abrogated the induction of luciferase activities stimulated with trypsin by 100, 80 and 50%, respectively. These results indicate that PAR2 activation in HEEC by trypsin induces NFkappaB- and AP-1-dependent IL-8 production in association with activation of p38 MAPK and ERK1/2, suggesting that esophageal inflammation may be induced by PAR2 activation via reflux of trypsin.

T-lymphocyte-derived tumor necrosis factor exacerbates anoxia-reoxygenation-induced neutrophil-endothelial cell adhesion.

The overall objective of this study was to determine whether T lymphocytes can modulate the increased neutrophil adherence and upregulation of endothelial cell adhesion molecules in human umbilical vein endothelial cells (HUVECs) exposed to anoxia/reoxygenation (A/R). HUVEC monolayers were exposed to 60 minutes of anoxia, followed by 4 hours of reoxygenation in the absence or presence of human T lymphocytes. The A/R-induced neutrophil adhesion was significantly enhanced when T lymphocytes and HUVECs were cocultured for the first 45 minutes of reoxygenation. This was accompanied by a more pronounced increase in E-selectin expression. When T lymphocytes were cocultured with HUVECs by use of inserts that prevented direct cell-cell contact, a comparable A/R-induced enhancement of neutrophil adhesion and of E-selectin expression was observed, indicating that soluble factors produced by T lymphocytes mediate the exaggerated A/R-induced inflammatory responses. Treatment with either an anti-tumor necrosis factor-alpha antibody or catalase attenuated the T-cell-mediated responses in postanoxic HUVECs. Moreover, the T-cell-mediated neutrophil adhesion response was mimicked by exposure of naive HUVECs to H(2)O(2). These findings indicate that H(2)O(2) produced by postanoxic endothelial cells stimulates T cells to produce tumor necrosis factor-alpha, which in turn elicits endothelial cell adhesion molecule expression and a corresponding increase in neutrophil adhesion.

Postanoxic T lymphocyte-endothelial cell interactions induce tumor necrosis factor-alpha production and neutrophil adhesion: role of very late antigen-4/vascular cell adhesion molecule-1.

The objective of this study was to define the influence of postanoxic T-lymphocyte-endothelial cell interactions on anoxia-reoxygenation (A/R)-induced neutrophil-endothelial cell adhesion and cell adhesion molecule (CAM) expression on human umbilical vein endothelial cells (HUVECs). HUVEC monolayers were exposed to 60 minutes of anoxia, followed by 24 hours of reoxygenation, wherein freshly isolated human T lymphocytes were added at 6 hours during reoxygenation. After an additional 18 hours of incubation (ie, total of 24 hours of reoxygenation), the T-cell/endothelial cell (TC/EC) coculture media were collected and added to naive HUVEC monolayers incubated with neutrophils. Although the A/R-conditioned media per se had no effect on neutrophil adhesion, the media from TC/EC cocultures significantly increased the adhesion response. This enhanced adhesive interaction was associated with significant increases in tumor necrosis factor-alpha (TNF-alpha) and interleukin-8 (IL-8) levels in the TC/EC coculture media and was accompanied by a pronounced increase in endothelial E-selectin expression. Treatment of the TC/EC coculture media with anti-TNF-alpha or anti-IL-8 antibodies reduced the media-induced neutrophil adhesion response. The enhanced neutrophil adhesion and the elevated medium levels of TNF-alpha, but not IL-8, were markedly reduced by inserts that prevented direct TC/EC contact and by monoclonal antibodies directed against vascular cell adhesion molecule-1 (VCAM-1) or very late antigen-4 (VLA-4). Collectively, these findings show that VLA-4-/VCAM-1-mediated interactions between T lymphocytes and postanoxic endothelial cells stimulates TNF-alpha production, which in turn elicits endothelial cell adhesion molecule expression and a corresponding increase in neutrophil adhesion.

A novel cancer therapy based on oxygen radicals.

The antitumor effect of oxygen radicals produced by hypoxanthine and xanthine oxidase reaction was studied in an experimental rabbit model. VX2 carcinomas were transplanted into rabbit hind legs. Hypoxanthine was administered continuously through the ear vein, while xanthine oxidase was administered simultaneously through the femoral artery. As a result, hypoxanthine and xanthine oxidase reacted only in the hind leg, and superoxide was produced in that area. The volume of the VX2 carcinoma was measured immediately prior to treatment and 7 days later. As an index of lipid peroxidation, thiobarbituric acid-reactive substances in the tumor tissue were measured 60 min following infusion of hypoxanthine and xanthine oxidase. Tumor growth was suppressed significantly by the hypoxanthine-xanthine oxidase reaction, and thiobarbituric acid-reactive substances in the tumor tissue infused with hypoxanthine and xanthine oxidase were significantly increased. In addition, the antitumor effect of the hypoxanthine and xanthine oxidase reaction was significantly inhibited by the administration of superoxide dismutase and catalase. Pathological examination showed that oxygen radicals produced by hypoxanthine and xanthine oxidase reaction were selectively more destructive for VX2 carcinoma tissue than muscle tissue surrounding the tumor region. These results suggest that oxygen radicals produced by hypoxanthine and xanthine oxidase reaction produce an anticancer effect and that the VX2 carcinoma used in this study was more sensitive to oxygen radicals than normal muscle tissue.

Antitumor effect of ischemia-reperfusion injury induced by transient embolization.

The effect of ischemia-reperfusion, induced by the transient embolic agent degradable starch microspheres (DSMs) on tumor tissue was investigated from the standpoint of active oxygen species. Rabbits with VX2 carcinoma received regional infusion of DSMs by transcatheter angiography, and it was confirmed that DSMs occluded tumor vessels. Blood flow in the tumors decreased rapidly immediately after the DSM treatment and returned to the original level within 40 min. The size of tumors did not change after a single infusion of DMS, while five repeated DSM treatments led to a significant reduction in tumor size. This reduction in tumor size was prevented by the treatment of rabbits with superoxide dismutase and catalase, indicating that the generation of active oxygen species in the tumor was involved in the mechanism of action of DSMs. Thiobarbituric acid-reactive substances also increased in the tumors after DSM infusion, and this increase was also inhibited by treatment with superoxide dismutase and catalase. In conclusion, the antitumor effect of the transient embolic agent DSM is secondary to the phenomenon of ischemia-reperfusion injury. In addition, active oxygen species and lipid peroxidation are possible causes of ischemia-reperfusion injury.

The role of active oxygen species and lipid peroxidation in the antitumor effect of hyperthermia.

The role of active oxygen species and lipid peroxidation in the antitumor effect of hyperthermia was studied in an experimental rabbit model. VX2 tumors were transplanted into rabbit hind legs, and the effect of hyperthermia on tumor growth was measured at 7 and 14 days after heating. As an index of lipid peroxidation, thiobarbituric acid-reactive substances in the tumor tissue were measured prior to hyperthermia and 3, 6, 12, and 24 h after hyperthermia. Tumor growth in rabbits treated with hyperthermia was significantly reduced, and thiobarbituric acid-reactive substances in the tumor tissue treated with hyperthermia were significantly increased until 6 h after hyperthermia. In addition, alpha-tocopherol in the tumor tissue was significantly decreased after hyperthermia. The antitumor effect of hyperthermia and the increase of thiobarbituric acid-reactive substances in the tumor tissue treated with hyperthermia were significantly inhibited by the administration of superoxide dismutase and catalase or dimethyl sulfoxide. These results suggest that lipid peroxidation mediated by active oxygen species plays an important role in the antitumor effect of hyperthermia.

Efficacy of hyperthermia and polyunsaturated fatty acids on experimental carcinoma.

We investigated the efficacy of hyperthermia and gamma-linolenic acid on experimental carcinoma. This study focused on polyunsaturated fatty acids that are substrates for free radical reactions. Oleic acid, linolenic acid, alpha-linolenic acid, or gamma-linolenic acid was injected into the arteries feeding AH109A carcinoma implanted into rat hind limbs. Among these, gamma-linolenic acid had the greatest effect on tumor tissue lipid peroxidation and demonstrated an antitumor effect. Consequently, gamma-linolenic acid injection into the feeding artery of a tumor was performed immediately prior to hyperthermia. This combination therapy induced a high level of lipid peroxidation in tumor tissue and a significant antitumor effect. Hyperthermia combined with gamma-linolenic acid produces free radical reactions by increasing the radical reaction substrate and may be an effective anticancer modality.

The effect of rebamipide on Helicobacter pylori extract-mediated changes of gene expression in gastric epithelial cells.

BACKGROUND: Recent studies have shown that Helicobacter pylori affects intracellular signal transduction in host cells, leading to the activation of transcriptional factors and the induction of pro-inflammatory cytokines. On the other hand, rebamipide, an anti-gastritis and anti-ulcer agent, could scavenge reactive oxygen species and reduce interleukin-8 (IL-8) expression in gastric epithelial cells induced by H. pylori-stimulation through the attenuated activation of nuclear factor-kappaB (NF-kappaB). AIMS: In this study, we investigated the effects of rebamipide on gene expression in H. pylori-stimulated epithelial cells using DNA chip. METHODS: H. pylori water extract (HPE) was prepared from NCTC11637, the type strain of H. pylori. Total RNA was extracted from MKN45 cells, a human gastric cancer cell line, following HPE-stimulation with and without rebamipide for 3 h, and differences in gene expression profiles were observed using GeneChip and Human 6800 probe array. RESULTS: The GeneChip analysis demonstrated that 132 up-regulated genes and 873 down-regulated genes, such as growth factors, chemokines and transcription factors, were detected in MKN45 cells 3 h after stimulation of H. pylori. Among them, several genes, including bFGF, RANTES and MIP-2beta, were previously unknown to be expressed in H. pylori-stimulated human gastric cells. Rebamipide reduced expression of 119 genes encoding cytokines, growth factors and their receptors and transcription factors. CONCLUSIONS: These findings suggest that rebamipide could inhibit inflammatory reactions and tumour progression by modifying H. pylori infection-induced gene expression in gastric epithelial cells.

[Effect of intra-arterial chemotherapy combined with degradable starch microspheres against liver cancer--a study by experimental VX2 liver tumor model].

We investigated the anticancer effect and pharmacokinetics of doxorubicin hydrochloride(ADR, 1 mg/kg) and mitomycin C (MMC, 0.2 mg/kg) given intra-arterially in combination with a temporary embolizing material, degradable starch microspheres (DSM, 5 mg/kg), to rabbits in which VX2 tumors had been implanted into the liver. The results revealed that the anticancer effect of ADR and MMC was potentiated by concomitant treatment with DSM in comparison to intra-arterial drug therapy alone. The tumor growth inhibition rate in both groups was about 87%. There was a tendency of higher intra-tumour ADR concentrations after ADR and DSM than after ADR alone. When we measured the serum levels of ADR and MMC over 45 min., it was indicated that exposure of the drugs to the systemic circulation was inhibited by the combination with DSM. In these experiments, GOT and GPT values increased temporarily after intra-arterial chemotherapy combined with DSM, but decreased gradually later to normal or close to the normal levels. These results indicate that intra-arterial chemotherapy combined with DSM causes a temporary blood flow blockade and the resulting regional retention of the cytotoxic agents, provides a more potent anticancer effect than drug therapy alone.

[Role of oxygen species and lipid peroxidation for anti-tumor effect of intra-arterial injection with degradable starch microspheres].

The role of superoxide and lipid peroxidation for anti-tumor effect of intra-arterial injection with degradable starch microspheres (DSM) was investigated in rabbits. The anti-tumor effect of intra-arterial injection with DSM was studied in rabbits with VX2 carcinoma of the hind leg. The tumor growth in rabbit treated with DSM 5 times was completely suppressed, and thiobarbituric acid (TBA)-reactive substances in the tumor tissue treated with DSM were significantly increased. But the anti-tumor effect of DSM and the increase in TBA reactive substances in the tumor tissue treated with DSM were significantly inhibited by treatment with superoxide dismutase combined with catalase. These results suggest that the anti-tumor effect of intra-arterial injection with DSM may be due to ischemia-reperfusion injury and that active oxygen species and lipid peroxidation may play an important role in the anti-tumor effect of intra-arterial injection with DSM.

[Chemoembolization with degradable starch microspheres for liver metastases of colorectal cancer].

Transcatheter chemoembolization using degradable starch microspheres (DSM) was performed in 17 patients with liver metastases of colorectal cancer. DSM, 45 microns in diameter, which are easily degraded by serum amylase, and therefore obstruct arterial blood flow temporarily at the arteriolar capillary bed. DSM mixed with mitomycin C were administered through the catheter introduced by Seldinger's method in 13 cases, and through subcutaneously implanted drug delivery system (Port-A-Cath) in 4 cases. The treatment was repeated 5 times on the average at intervals of 2 to 4 weeks. The therapeutic effect of this chemoembolization was evaluated by the change in tumor size measured by angiography or computed tomography. Tumor regression of over 50% was observed in 9 of 17 cases (53%). Elevated serum CEA levels (greater than 10 ng/ml) decreased in 10 of 15 cases (67%). One-year survival rate was 48% in 17 cases, and among them it was 100% in 7 cases with the extent of several liver metastases (H2). Extrahepatic metastasis was observed in 3 of 17 cases (18%) before the treatment, and in 8 cases (47%) after the treatment. Abdominal pain occurred in 36% of the cases by the administration of DSM, but the pain disappeared within 2 hours. No major side effects such as bone marrow suppression or hepatotoxicity were observed. Our results suggest that chemoembolization using DSM is effective and safe in the treatment of liver metastases from colorectal cancer.

[Role of polymorphonuclear leukocytes (PMN) and active oxygen species in hyperthermia--antitumor effect of hyperthermia combined with rhG-CSF].

We have reported that polymorphonuclear leukocytes (PMN) and active oxygen species from PMN may play an important role in the mechanism of the antitumor effect of hyperthermia. At this time, we focused our experimental studies on rat AH109A carcinoma treated with hyperthermia combined with arterial injection of rhG-CSF. Rats with transplantable AH109A carcinoma at the hind leg received hyperthermia. These tumors showed mild suppression of further development only by hyperthermia. However, when arterial injection of rhG-CSF was applied together with hyperthermia, marked suppression of tumor development was observed. Our data suggest that hyperthermia combined with rhG-CSF is closely related to the generation of free radical-mediated tumor cell killing, and it can be an effective treatment for cancer.

[Role of polymorphonuclear leukocytes (PMN) and active oxygen species in hyperthermia--enhancing effect of G-CSF on superoxide generation from PMN].

We examined in vitro the effect of G-CSF and temperature on superoxide (O2-) generation by Cypridina luciferin analog (CLA) dependent chemiluminescence. PMN significantly generated O2- at the concentration of G-CSF 25 ng/ml or more at 37 degrees C within the range of 0.1 from 1,000 ng/ml. O2- generation from PMN was remarkably enhanced, stimulated by opsonized zymosan (OZ) and phorbol myristate acetate (PMA), at 41 degrees C as compared with 37 degrees C. O2- generation was enhanced with the addition of 25 ng/ml of G-CSF at 41 degrees C as compared to without it at 41 degrees C. A significant enhancement of O2- generation from PMN was observed at 25 ng/ml G-CSF and 41 degrees C.

[Role of oxygen-derived free radicals for antitumor effects of intra-arterial injection with adriamycin].

This study examined the role of oxygen-derived free radicals for antitumor effects of intra-arterial injection with adriamycin (ADR). The effect of chemoembolization using ADR and degradable starch microspheres (DSM) was studied in rabbits with VX2 carcinoma of the hind leg. The tumor growth in rabbit treated with chemoembolization (ADR: 3.0 mg/kg + DSM 20mg/kg) was completely suppressed, but the therapeutic effect of chemoembolization combined with dimethylsulfoxide (DMSO) was reduced. On the other hand, the therapeutic effect of chemoembolization combined with superoxide dismutase and catalase was not reduced. These results indicate that the production of oxygen-derived free radicals, especially hydroxyl radical in the cancer cell, plays an important role in the antitumor effect of adriamycin.

Inhibition of Bach1 ameliorates indomethacin-induced intestinal injury in mice.

BTB and CNC homolog 1 (Bach1) is a transcriptional repressor of heme oxygenase-1 (HO-1). It plays an important role in the feedback regulation of HO-1 expression, which protects cells from various insults including oxidative stress and inflammatory cytokines. However, the role of Bach1 in intestinal inflammation remains unclear. In this study, the role of Bach1 in intestinal mucosal injury was elucidated using 8-week-old female C57BL/6 (wild-type) and homozygous Bach1-deficient C57BL/6 mice. Intestinal mucosal injuries induced by a single subcutaneous administration of indomethacin were evaluated macroscopically, histologically, and biochemically. Mucosal protein content and chemokine mRNA levels were determined by real-time PCR. Our results showed that the indomethacin-induced intestinal injury was remarkably improved in Bach1-deficient mice. Histological examination showed that the area of injured lesion was decreased in Bach1-deficient mice compared to wild-type mice. Administration of indomethacin induced expression of inflammatory chemokines such as KC, MIP1alpha and MCP1, which was suppressed in Bach1-deficient mice. Myeloperoxidase activity in the intestinal mucosa was also significantly decreased in Bach1-deficient mice. Additionally, Bach1 deficiency enhanced immunopositivity of HO-1 in the intestinal mucosa after indomethacin administration. Disruption of the Bach1 gene thus caused inhibition of mucosal injury, indicating that inhibition of Bach1 may be a novel therapeutic strategy for treating indomethacin-induced intestinal injury.

Hyperthermia ameliorates 2,4,6-trinitrobenzene sulphonic acid-induced colitis in rats: the role of heat shock proteins.

PURPOSE: Hyperthermia is known to protect against cellular injury through the expression of heat shock proteins. In this study, the therapeutic effects of hyperthermia on experimental colitis in the rat were evaluated. MATERIALS AND METHODS: Male Wistar rats were given a single intracolonic injection of 2,4,6-trinitrobenzene sulphonic acid (TNBS). Hyperthermia was induced in anesthetized rats by placing them in a temperature-controlled water bath. We started the hyperthermic treatment on the day after the enema. The severity of colitis was evaluated pathologically, and the activities of tissue myeloperoxidase were measured 6 days after the induction of colitis. Furthermore, cytokines, and hyperthermia-induced heat shock proteins in colonic mucosa were detected by enzyme-linked immunosorbent assay and Western blotting. We also investigated the effects of geranylgeranylacetone and zinc protoporphyrin IX on the therapeutic effect of hyperthermia. RESULTS: Hyperthermia significantly improved the macroscopic scores of colitis. The TNBS-induced increases in the activities of myeloperoxidase in the colonic tissue were blunted significantly in hyperthermia-treated animals. Furthermore, hyperthermia attenuated increases in cytokine-induced neutrophil chemoattractants-1 and tumor necrosis factor-alpha in the colon. Furthermore, hyperthermia induced the production of heat shock proteins in rat colonic mucosa, and the combination of geranylgeranylacetone with hyperthermia further induced the heat shock protein HSP70, which resulted in further improvement of TNBS-induced colitis. On the other hand, the combination of zinc protoporphyrin IX with hyperthermia attenuated the therapeutic effect of hyperthermia. CONCLUSIONS: Hyperthermia ameliorates TNBS-induced colitis in rats through the expression of HSP70 and HO-1. It is postulated that hyperthermia may be useful for the treatment of inflammatory bowel diseases.

Molecular mechanisms involved in anti-inflammatory effects of proton pump inhibitors.

OBJECTIVE: Interleukin (IL)-8 has been reported to participate in neutrophil infiltration in Helicobacter pylori (H. pylori)-induced gastritis in humans. In this study, we investigated the anti-inflammatory actions beyond the suppression of acid secretion by proton pump inhibitors (PPI), such as omeprazole and lansoprazole, on IL-8 production by gastric epithelial cells (MKN45) and human umbilical vein endothelial cells (HUVEC) and on the transendothelial migration of polymorphonuclear neutrophils (PMN). MATERIALS AND METHODS: MKN45 and HUVEC were stimulated with H. pylori water extract (HPE) and IL-1beta, respectively, and nuclear factor kappa B (NFkappaB) activation and subsequent IL-8 production was assessed in the absence or presence of PPI. We also assessed the effect of PPI on IL-8-induced PMN transendothelial migration and on the alteration of cytoplasmic calcium concentration in formyl-methionyl-leucyl-phenylalanine (fMLP)-stimulated PMN. RESULTS: HPE and IL-1beta induced a significant increase in IL-8 production by MKN45 and HUVEC, respectively, along with NFkappaB activation, which was significantly inhibited by PPI. PPI also inhibited the IL-8-induced transendothelial migration of PMN and the fMLP-induced cytosolic calcium increase in PMN. CONCLUSIONS: PPI attenuate PMN-dependent gastric mucosal inflammation partly by interfering with NFkappaB activation in vascular endothelial cells and gastric epithelial cells, and partly by modulating the calcium concentration of PMN.