Endotoxin activity and leukocytic STAT3 mRNA alterations differ according to age in lipopolysaccharide-challenged calves.

The objective of this study was to investigate the age difference in the response to endotoxin in calves related to the plasma endotoxin activity and mRNA expression of cytokines. Fifteen calves were divided into three groups: control (191 ± 21 days), weaning (162.4 ± 17.5 days), and calf (22.4 ± 8.2 days). The weaning and calf groups received 2.5 µg/kg of ultrapure O111:B4 LPS in 10 mL of each autologous serum, whereas the control calves received a similar volume of saline. Blood samples were collected at 0, 0.5, 1, 2, 4, 8, 12, and 24 h. Liver samples were collected by ultrasound-guided liver biopsy at 0, 2, 4, and 24 h. Plasma endotoxin activity was measured by the limulus amebocyte lysate kinetic turbidimetric assay. The mRNA expression level of GAPDH, TLR-4, NF-κB2, TNF-α, IL-6, and STAT3 in leukocytes and the liver was measured by real-time PCR. Following LPS challenge, the maximal plasma endotoxin activity, and leukocytic expression of TLR4, NF-κB2, TNF-α, and STAT3 were reached at 0.5, 4, 2-4, 2-4, and 4 h, respectively, in weaning and calf groups. The endotoxin activity in calf remained high until 2 h. Furthermore, the expression of leukocytic STAT3 mRNA in calf was not significantly different from the pre-value. In contrast, STAT3 mRNA in weaning markedly increased at 2 and 4 h. Therefore, this study provides new information to the literature of immune and inflammatory responses in calves.

Malignant female adnexal tumor of Wolffian origin (FATWO) positive for CD56: a possible diagnostic role for the biomarker.

Female adnexal tumors of Wolffian origin (FATWOs) are rare tumors that arise in the broad ligament from the remnants of the mesonephric duct. Most FATWOs behave in a benign fashion, and there are only 14 case reports worldwide describing malignant FATWOs. The authors report herein the case of a 69-year-old woman with a malignant FATWO, positive for CD56. The mass was composed mainly of solid neoplastic epithelial cells, closely packed, branching, and anastomosing in slender tubules. There was an eosinophilic secretion within the lumens of some of the cysts and tubules. The number of mitoses was somewhat high in the active areas, numbering five to seven per ten high-power fields. The tumor cells were strongly positive for glutathione S-transferase π, and positive for cal- retinin, vimentin, c-Kit, CD99, and CD56; neuron-specific enolase was also partially expressed. The tumor cells were negative for inhibin α, estrogen receptors, progesterone receptors, B-cell lymphoma 2, and S100. Taken together, these immunohistochemical and pathological findings gave the diagnosis of malignant FATWO. The patient experienced a recurrence one year after her initial surgery. CD56 immunostaining was negative in two benign FATWO cases at the present institution. These findings suggest that CD56-positivity may be a diagnostic biomarker to differentiate malignant FATWOs from benign lesions.

Lymph node metastasis as a significant prognostic factor in gastric cancer: a multiple logistic regression analysis.

BACKGROUND: In Japan, the standard treatment policy for all potentially curable patients with gastric cancer is radical resection including extensive lymphadenectomy. This treatment strategy has been used for both early and advanced gastric cancers, and substantial increases in survival time have been reported. In advanced gastric cancer, lymphatic spread is reported to be one of the most relevant prognostic factors for gastric cancer resected for cure. The purpose of this study was to determine the factors affecting lymph node involvement and to establish guidelines for the extent of lymph node dissection most appropriate for the treatment of gastric cancer. METHODS: The clinicopathological features of 926 patients with gastric cancer were reviewed. Information on the clinicopathological features was obtained from the database of gastric cancer at the Department of Gastroenterological Surgery, Sendai National Hospital. Univariate and multivariate analyses of data for patients with gastric cancer tumors were performed to evaluate the prognostic significance of clinicopathological features. The independent risk factors influencing lymph node metastasis were determined by multiple logistic regression analysis. RESULTS: The following clinicopathologic factors were found to be correlated with prognosis of gastric cancer: (1) macroscopic type, (2) depth of invasion, (3) cancer-stromal relationship, (4) histological growth pattern, (5) lymph node involvement, (6) lymphatic invasion, (7) vascular invasion and (8) tumor site. However, a multivariate analysis revealed that macroscopic type, depth of invasion, lymph node involvement and tumor site are independent risk factors for the prognosis of gastric cancer patients. Among these factors, the prognosis of patients with gastric cancer was most strongly influenced by lymph node involvement (odds ratio, 4.632). According to a multiple logistic regression model, depth of cancer invasion and lymphatic invasion was significantly correlated with lymph node metastases. CONCLUSIONS: Lymph node involvement has the strongest influence on the prognosis of gastric cancer. Among the clinicopathological factors, depth of invasion and microscopically lymphatic invasion are important factors in predicting lymph node metastases. Thus, the ability to perform gastrectomy with dissection of lymph nodes is a basic requirement for gastric cancer surgeons.

Prognostic factors of gastric cancer tumours of less than 2 cm in diameter: rationale for limited surgery.

BACKGROUND: A recent trend in the surgical treatment of patients with early gastric cancer in Japan has been to limit surgery to an extent that ensures complete cure and improvement in the patient's quality of life. If a gastric cancer tumour can be completely eradicated by laparoscopic surgery, the patient can be cured of cancer without major operative stress. A small gastric cancer tumour of less than 2 cm in diameter is an indication for laparoscopic surgery, but little is known about what protocol of surgical treatment is appropriate for this type of tumour. PATIENTS AND METHODS: The clinicopathological features of 150 patients with gastric cancer tumour of less than 2 cm in diameter were reviewed retrospectively from hospital records between 1985 and 1995. The results of retrospective analysis of clinicopathological data of 24 patients with advanced cancer were compared with those of 126 patients with early cancer. Univariate and multivariate analyses of patients with small gastric cancer tumours were performed to evaluate the prognostic significance of clinicopathological features. RESULTS: A significant difference was seen between the gross tumour appearances in the two groups; Borrmann type-4 tumours were more common in the advanced group. Lymph-node metastasis, lymphatic vessel invasion and vascular invasion were found more frequently in the advanced cancer group than in the early cancer group. Scirrhous type was more common in the advanced cancer group. In univariate analysis, unfavourable prognostic factors included deep cancer invasion, presence of lymph-node metastasis, lymphatic invasion and vascular invasion. Using Cox's proportional hazard regression model, only nodal involvement emerged as an independent statistically significant prognostic parameter associated with long-term survival. CONCLUSION: Laparoscopic surgery should not be performed on tumours that are Borrmann type in macroscopic appearance and scirrhous-type histologically. Lymph-node metastasis is an independent prognostic factor. We recommend laparoscopic surgery involving local resection of the stomach without lymphadenectomy for small, early gastric cancer tumours that satisfy the criteria mentioned above. However, the validity of this recommendation should be tested by a prospective randomized control trial in the future.

Helicobacter pylori lipopolysaccharide induces apoptosis of cultured guinea pig gastric mucosal cells.

Helicobacter pylori lipopolysaccharide (LPS) is generally accepted as a low-toxicity virulence. Primary cultures of guinea pig gastric mucosal cells expressed the Toll-like receptor 4 and were sensitive to H. pylori LPS as well as Escherichia coli LPS. H. pylori LPS stimulated phosphorylation of transforming growth factor-beta-activated kinase 1 (TAK1), TAK1-binding protein 1 (TAB1), and c-Jun NH(2)-terminal kinase (JNK) 2. H. pylori LPS at >2.1 endotoxin unit/ml (>1 ng/ml) activated caspase-8, stimulated cytochrome c release from mitochondria, and subsequently activated caspases-9 and -3, leading to apoptosis. Epidermal growth factor blocked all of these apoptotic processes and inhibited apoptosis, whereas it did not modify the phosphorylation of TAK1, TAB1, and JNK2. A comparatively specific inhibitor of caspase-8 or -9 blocked apoptosis, whereas cytochrome c release was prevented only with a caspase-8-like inhibitor. Our results suggest that caspase-8 and mitochondria may play crucial roles in H. pylori LPS-induced apoptosis and that this accelerated apoptosis may be involved in abnormal cell turnover of H. pylori-infected gastric mucosa.

Type I Helicobacter pylori lipopolysaccharide stimulates toll-like receptor 4 and activates mitogen oxidase 1 in gastric pit cells.

Guinea pig gastric pit cells express an isozyme of gp91-phox, mitogen oxidase 1 (Mox1), and essential components for the phagocyte NADPH oxidase (p67-, p47-, p40-, and p22-phox). Helicobacter pylori lipopolysaccharide (LPS) and Escherichia coli LPS have been shown to function as potent activators for the Mox1 oxidase. These cells spontaneously secreted about 10 nmol of superoxide anion (O(2)(-))/mg of protein/h under LPS-free conditions. They expressed the mRNA and protein of Toll-like receptor 4 (TLR4) but not those of TLR2. LPS from type I H. pylori at 2.1 endotoxin units/ml or higher stimulated TLR4-mediated phosphorylations of transforming growth factor beta-activated kinase 1 and its binding protein 1 induced TLR4 and p67-phox and up-regulated O(2)(-) production 10-fold. In contrast, none of these events occurred with H. pylori LPS from complete or partial deletion mutants of the cag pathogenicity island. Lipid A was confirmed to be a bioactive component for the priming effects, while removal of bisphosphates from lipid A completely eliminated the effects, suggesting the importance of the phosphorylation pattern besides the acylation pattern for the bioactivity. H. pylori LPS is generally accepted as having low toxicity; however, our results suggest that type I H. pylori lipid A may be a potent stimulator for innate immune responses of gastric mucosa by stimulating the TLR4 cascade and Mox1 oxidase in pit cells.

Interleukin-1beta enhances retinoic acid-mediated expression of bone-type alkaline phosphatase in rat IEC-6 cells.

We previously showed that vitamin A upregulated the expression of bone-type alkaline phosphatase (ALP) in fetal rat small intestine and rat intestinal IEC-6 cells. In this study, we examined interactions between retinoic acid (RA) and several growth factors/cytokines on the isozyme expression in IEC-6 cells. Epidermal growth factor and interleukins (ILs)-2, -4, -5, and -6 completely blocked the RA-mediated increase in ALP activity. In contrast, IL-1beta markedly increased the activity, protein, and mRNA of the bone-type ALP only when RA was present. IL-1beta and/or RA did not change the type 1 IL-1 receptor transcript level, whereas IL-1beta enhanced the RA-induced expressions of retinoic acid receptor-beta (RAR-beta) and retinoid X receptor-beta (RXR-beta) mRNAs and RA-mediated RXR response element binding. The synergism of IL-1beta and RA on ALP activity was completely blocked by protein kinase C (PKC) inhibitors. Our results suggest that IL-1beta may modify the ALP isozyme expression in small intestinal epithelial cells by stimulating PKC-dependent, RAR-beta- and/or RXR-beta-mediated signaling pathways.

Probability of lymph node metastasis in small gastric cancer tumor: is it an indication for limited surgery?

The purpose of this study was to determine the factors that are predictive of lymph node metastasis in a small gastric cancer tumor <2 cm in diameter. The clinicopathological features of 17 patients with node-positive small gastric cancer were reviewed from the database of gastric cancer at the Department of Surgery, Sendai National Hospital, Sendai, Japan, and they were compared with those of 131 patients with node-negative cancer. The independent risk factors influencing the lymph node metastasis were determined by multiple logistic regression analysis. Depth of invasion, macroscopic appearance, cancer-stromal relationship, and lymphatic microinvasion were found to be associated with lymph node metastasis. The variables found to be significant risk factors for lymph node metastasis were depth of invasion (P = 0.0250) and lymphatic microinvasion (P = 0.0028). It is possible for even a small gastric cancer tumor to have lymph node metastasis. A surgeon treating a small gastric cancer tumor must consider that although the cure rate is high, >10% of these tumors have lymph node metastases. Because of the possibility of lymph node metastasis, even with accurate knowledge of the depth of cancer invasion, selective performance of local resection or limited surgery with incomplete lymph node dissection is not justified. Accurate preoperative diagnosis and the appropriate decision for surgical indication are important. Large-scale randomized, controlled trials should be performed to show the advantage of limited surgery for gastric cancer.

Regulation of growth and apoptosis of cultured guinea pig gastric mucosal cells by mitogenic oxidase 1.

We previously reported that primary cultures of guinea pig gastric pit cells expressed all of the phagocyte NADPH oxidase components (gp91-, p22-, p67-, p47-, and p40-phox) and could spontaneously release superoxide anion (O(2)(-)). We demonstrate here that pit cells express a nonphagocyte-specific gp91-phox homolog (Mox1) but not gp91-phox. Inclusion of catalase significantly inhibited [(3)H]thymidine uptake during the initial 2 days of culture. Pit cells, matured on day 2, slowly underwent spontaneous apoptosis. Scavenging O(2)(-) and related oxidants by superoxide dismutase plus catalase or N-acetyl cysteine (NAC) and inhibiting Mox1 oxidase by diphenylene iodonium activated caspase 3-like proteases and markedly enhanced chromatin condensation and DNA fragmentation. This accelerated apoptosis was completely blocked by a caspase inhibitor, z-Val-Ala-Asp-CH(2)F. Mox1-derived reactive oxygen intermediates constitutively activated nuclear factor-kappaB, and inhibition of this activity by nuclear factor-kappaB decoy oligodeoxynucleotide accelerated their spontaneous apoptosis. These results suggest that O(2)(-) produced by the pit cell Mox1 oxidase may play a crucial role in the regulation of their spontaneous apoptosis as well as cell proliferation.

[A case of locally advanced rectal carcinoma with liver metastasis treated with a combination of CRT, HAI, and systemic chemotherapy].

A 55-year-old man with locally advanced rectal carcinoma and liver metastasis was treated with a combination of chemo-radiotherapy (5-FU suppository 100 mg/day and 63 Gy of RT), hepatic arterial infusion chemotherapy (5-FU 1,000 mg/3 h, biweekly), and systemic chemotherapy (5'-DFUR 800 mg/day + cimetidine 800 mg/day). His rectal tumor was reduced and his symptoms such as pain and bleeding had markedly decreased. The river metastasis did not change during the entire course. HAI and administration of 5-FU suppository, 5'-DFUR, and cimetidine were continued. As of 18 months after the onset of the combination therapy, NC has been maintained, and the general condition of the patient is favorable.

Geranylgeranylacetone stimulates mucin synthesis in cultured guinea pig gastric pit cells by inducing a neuronal nitric oxide synthase.

Nitric oxide (NO) has been considered to play an important role in the regulation of blood flow, mucosal integrity, and mucus production in the stomach. We investigated the stimulatory actions of epidermal growth factor (EGF) and a cytoprotective compound, geranylgeranylacetone (GGA), on mucin synthesis in guinea pig gastric pre-pit cells, maintained in a serum-free culture system. GGA increased [3H]glucosamine uptake and the accumulation of mucus granules positive for galactose oxidase-Schiff reaction in the cells. This stimulatory action of GGA was equivalent to that of EGF, but GGA did not stimulate the cell growth. Both EGF and GGA increased the release of NO degeneration products, NO2- and NO3-. The [3H]glucosamine uptake was completely inhibited by the non-selective NO synthase (NOS) inhibitors, N(G)-nitro-L-arginine and N(G)-monomethyl-L-arginine, and it was only partially inhibited by a more selective inhibitor for inducible NOS isoform (iNOS), aminoguanidine. Northern blotting with a cDNA probe for rat iNOS, and Western blotting with a polyclonal antibody against iNOS, demonstrated that GGA did not up-regulate the iNOS mRNA expression nor induce its protein. In contrast, GGA and EGF induced neuronal NOS, but not endothelial NOS, which was confirmed by immunoblot analyses with antibodies against these constitutive NOS isoforms. Thus, the present experiments suggests that GGA, as well as EGF, stimulates mucin synthesis at least in part through an NO-dependent pathway, leading to an increase in the integrity of the gastric mucosa.

Phenylarsine oxide inhibits heat shock protein 70 induction in cultured guinea pig gastric mucosal cells.

Phenylarsine oxide (PAO) forms a stable ring complex with vicinal dithiols that can be reversed with 2,3-dimercaptopropanol (DMP) but not by dithiothreitol (DTT) or 2-mercaptoethanol (2-ME). PAO at 2 microM or higher inhibited heat shock protein 70 (HSP70) induction within minutes in cultured guinea pig gastric mucosal cells exposed to heat (43 degrees C) for 30 min. PAO did not affect the nuclear translocation and phosphorylation of heat shock factor 1 (HSF1) induced by heat stress, but it completely blocked the binding activity of HSF1 to the heat shock element (HSE), leading to the block of expression of HSP70 mRNA and accumulation of HSP70 in the cells. These inhibitions were completely reversed with 2 microM DMP but not with 0.1 mM DTT or 1 mM 2-ME, suggesting specific interactions between PAO and vicinal dithiol-containing molecules. Thioredoxin (Trx) reversed the inhibition of the binding activity of HSF1 in whole cell extracts prepared from PAO-treated, heat-stressed cells. Our results suggest that PAO may react with vicinal-containing molecules including Trx and specifically block the interaction between HSF1 and HSE.

A non-toxic heat shock protein 70 inducer, geranyl-geranyl-acetone, restores the heat shock response in gastric mucosa of protein-malnourished rats.

Acute gastric mucosal lesions caused by stress or noxious stimuli are important to consider in the management of critically or chronically ill patients. Protein malnutrition has been implicated as a risk factor for stress ulcer and subsequent complications in those patients. When male Wistar rats fed a 5% or 20% casein diet for 3 weeks were exposed to restraint and water-immersion stress, the low-protein diet significantly increased the ulcer index. The low-protein diet did not change the level of heat shock factor 1 (HSF1) in gastric mucosa but it did attenuate the HSF1 activation after exposure to the stress, resulting in the inhibition of HSP70 mRNA expression and HSP70 induction in gastric mucosa. HSP70 is crucial for the maintenance of cell integrity during pathophysiologic conditions; therefore the impaired HSP70 induction appeared to at least in part aggravate stress ulcer. We also tested whether a non-toxic HSP70 inducer, geranyl-geranyl-acetone (GGA), effectively improved the mucosal integrity by stimulating HSP70 induction under protein malnutrition. Intragastric administration of GGA (200 mg/kg twice a day) to the protein-malnourished rats for up to 1 week failed to stimulate the HSP70 induction. However, the administration of GGA (200 mg/kg twice a day) for 3 weeks restored HSP70 induction and induced higher resistance against stress ulcer as compared with results in vehicle-treated, normally nourished rats. Our results suggest that GGA may have a potential benefit for the prevention of stress ulcer in chronically or critically ill patients with protein malnutrition.

Significant prognostic factors in patients with early gastric cancer.

BACKGROUND: Early gastric cancer is defined as a gastric carcinoma confined to the mucosa or submucosa regardless of lymph node status, and it has an excellent prognosis with a 5-year survival rate of more than 90%. From 1985 to 1995, we encountered 266 cases of early gastric cancer in our hospital. METHODS: A retrospective analysis of the 266 cases of early gastric cancer was performed to evaluate the prognostic significance of clinicopathological features (age, gender, tumor size, tumor location, depth of invasion, lymph node metastasis, histological type, lymphatic invasion, vascular invasion, histological growth pattern, cancer-stromal relationship and type of operation). RESULTS: The overall survival rate of all the patients with early gastric cancer was 95.7%. In univariate analysis, the statistical significant prognostic factors were regional lymph node metastasis (P = 0.0004), lymphatic invasion (P = 0.0053) and cancer-stromal relationship (P = 0.0016). Absence of lymph node metastasis and lymphatic invasion, and a medullary-type histopathology were associated with improved survival. In multivariate analysis, the statistically significant prognostic factors were lymph node metastasis and cancer-stromal relationship. CONCLUSIONS: Presence of lymph node involvement and a scirrhous type of gastric cancer are associated with poor prognosis. Lymph node dissection with gastric resection is necessary for patients with early gastric cancer who have a high risk of lymph node metastasis. Postoperative chemotherapy is recommended for a scirrhous type of early gastric cancer.

Clinicopathologic prognostic factors in patients with Borrmann type 4 gastric cancer: univariate and multivariate analyses.

BACKGROUND: Advanced gastric cancer is classified into four Borrmann types, types 1 to 4. Type 4 is a relatively undifferentiated carcinoma with little or no gland-forming capability. Despite recent advances in the diagnosis and surgical management of gastric cancer, most tumors of Borrmann type 4 are not detected at an early stage and the prognosis remains poor; the five-year survival rate after gastric resection ranges from 10 to 20 percent. We evaluated the affects of several clinicopathologic variables on the 5-year survival rate after resection of Borrmann type 4 gastric cancer. METHODS: Data on clinical characteristics were obtained from the records of patients who underwent gastric resection between 1985 and 1995 at the Department of Surgery, Sendai National Hospital, and follow-up data were obtained from our tumor registry. Pathologic characteristics were determined from a detailed review of all available histopathologic slides. The relationship between clinicopathologic variables and 5-year survival rate was estimated by the Kaplan-Meier survival curve and the logrank test. Multivariate Cox's proportional hazards regression analysis was then performed to determine which variables were independent prognostic factors. RESULTS: Eighty-seven patients with Borrmann type 4 gastric cancer underwent a resection during the study period at our hospital. The overall 5-year survival rate was 14.8%. The relationship between clinicopathologic variables and 5-year survival rate was determined by constructing a Kaplan-Meier survival curve. Tumor location (upper, middle and distal vs whole stomach, p=0.0214), lymph node metastasis, capillary microinvasion, and peritonitis carcinomatosa (absent vs present, p<0.05) significantly influenced survival. When multivariate analysis using Cox's proportional hazards regression of 5-year survival was performed, capillary microinvasion, peritonitis carcinomatosa (absent vs present) and tumor location (distal vs whole stomach) emerged as the statistically significant independent prognostic factors associated with long-term survival. CONCLUSION: Capillary microinvasion and the presence or absence of peritonitis carcinomatosa are more powerful predictors of 5-year survival than is lymph node metastasis. Patients with gastric cancer of the whole stomach have a poorer prognosis than do those with carcinoma in the antrum of the stomach.

Borrmann's type IV gastric cancer: clinicopathologic analysis.

OBJECTIVE: To determine whether there is a specific pattern of clinicopathological features that could distinguish Borrmann's type IV gastric cancer from other types of gastric cancer. DESIGN: A retrospective study of patients with advanced gastric cancer treated between 1985 and 1995. SETTING: The Department of Surgery, Sendai National Hospital, a 716-bed teaching hospital. PATIENTS: The clinicopathologic features of 88 patients with Borrmann's type IV carcinoma of the stomach were reviewed from the database of gastric cancer. The results were compared with those of 309 patients with other types of gastric carcinoma. MAIN OUTCOME MEASURES: Gender, age, tumour size, depth of invasion, histologic type, cancer-stromal relationship, histologic growth pattern, nodal involvement, lymphatic and vascular invasion, type of operation, cause of death and 5-year survival. RESULTS: Women were afflicted as commonly as men in the Borrmann's type IV group. These patients tended to be younger and to have larger tumours involving the entire stomach than patients with other types of cancer. Histologic type was commonly diffuse and scirrhous, and serosal invasion was prominent with infiltrative growth. Nodal involvement and lymphatic invasion were more common in patients with Borrmann's type IV than in those with other types of gastric cancer. The disease was advanced in most instances and a total gastrectomy was performed in 55% of the patients. The survival rate of patients with Borrmann's type IV tumour was lower than for patients with other types of gastric cancer (p < 0.005, log-rank test). CONCLUSIONS: In Borrmann's type IV gastric cancer, early detection and curative resection are crucial to extend the patient's survival. Aggressive postoperative chemotherapy is recommended when a noncurative resection is performed.

Early gastric cancer in the young: clinicopathological study.

BACKGROUND: Thirty-four cases of early gastric carcinoma in patients under 50 years of age treated in the period from 1985 to 1995 were reviewed. METHODS: These constituted 3.7% of 923 cases of gastric cancer in patients of all ages that were treated at Sendai National Hospital during the same 10-year period. Data were compared with those of 194 patients 50 years of age or older. RESULTS: The incidence of gastric cancer in men and women was almost the same in both groups. Tumours tended to be located distally in the stomach. Macroscopically, depressed lesions were more common in younger patients. Significant differences were observed in depth of invasion, histological type and histological growth patterns. CONCLUSIONS: The distinctive histological features of early gastric cancer in younger patients were a diffuse type of cancer with infiltrative tumours in the mucosal layer. The prognosis of younger patients was similar to that of older patients.