Ontogenic Expression of Somatostatin-Messenger RNA in the Intestinal Tract of Neonatal Rats

Ontogenic expression of somatostatin (SRIF)—messenger RNA (mRNA) in the gastrointestinal tract was examined in neonatal rats aged from 1 day preterm to 60 days postpartum in comparison with that in the hypothalamus. SRIF-mRNA in the hypothalamus was already expressed in prenatal rats and its developmental change was relatively small. In contrast, a unique pattern of SRIF-mRNA expression was seen in the different intestinal regions, gastric antrum, duodenum, jejunum and colon. In the duodenum, SRIF-mRNA level was low at birth, markedly increased during the postnatal 3 days and declined to the previous level by day 21. Jejunal SRIF-mRNA was found in neonates but progressively decreased in a similar way to duodenum. On the contrary, gastric SRIF-mRNA level, which was low during early development, rose rapidly to a peak on day 21 and gradually declined to an adult level. In the colon age-related change was not conspicuous, remaining at a low level. These results indicate that (1) expression of SRIF gene in the intestinal tract is regulated by local factor(s) as well as developmental stage, and (2) shift of SRIF-mRNA pattern occurs during weaning from the duodenum-dominant infantile pattern to the gastric-dominant adult pattern.     

Differential responses of scirrhous and well-differentiated gastric cancer cells to orthotopic fibroblasts.

Scirrhous gastric cancer cells proliferate rapidly with fibrosis, when the cancer cells invade into the submucosa of the stomach. To investigate the mechanisms responsible for the rapid proliferation, the growth interaction between gastric cancer cells and fibroblasts was examined. Human gastric cancer cell lines established from scirrhous carcinoma or well-differentiated adenocarcinoma were used. Human fibroblast cell lines were obtained from various organs. The growth interaction between gastric cancer cells and fibroblasts was examined by calculating the number of cancer cells or by measuring [3H]thymidine incorporation of cancer cells. Gastric fibroblasts specifically stimulated the growth of scirrhous gastric cancer cells, but not that of well-differentiated adenocarcinoma cells. The growth factor(s) produced from gastric fibroblasts were then partially purified and characterised. The growth-promoting factor(s) had apparent molecular weights of 10000 dalton and was sensitive both to heat and proteinase treatment. No inhibition for the factor(s) was achieved with defined anti-growth factor antibodies. In this study, differential responses of scirrhous and well-differentiated gastric cancer cells to orthotopic fibroblasts were shown. Rapid proliferation of scirrhous gastric carcinoma should be partly controlled by orthotopic fibroblasts. The growth factor(s) from gastric fibroblasts, which was distinct from various defined growth factors such as epidermal growth factor (EGF), basic fibroblast growth factor (b-FGF), transforming growth factor-alpha (TGF-alpha), keratinocyte growth factor (KGF), vascular endothelial growth factor (VEGF), insulin-like growth factor I (IGF-I), hepatocyte growth factor (HGF), platelet-derived growth factor (PDGF) and transforming growth factor beta 1 (TGF-beta 1) may play an important role in the progression of scirrhous gastric cancer cells.     

CD44H Plays an Important Role in Peritoneal Dissemination of Scirrhous Gastric Cancer Cells

The role of the adhesion molecule CD44H in the peritoneal adhesion and invasion of cancer cells was assessed using cell lines with low and high peritoneal seeding ability, OCUM-2M (2M) and OCUM-2MD3 (2MD3), respectively. The in vitro binding ability to peritoneal components (mesothelial cells, fibroricetin and type I collagen) and invasive ability of 2MD3 cells were higher than those of 2M cells. The expression level of CD44H on 2MD3 cells was higher than that on 2M cells as determined by western blot analysis and flow cytometry. The adhesiveness of 2MD3 cells to hyaluronic acid, which is expressed on the surfaces of mesothelial cells, was greater than that of 2M cells. The binding ability of 2MD3 cells to mesothelial cells was inhibited in the presence of anti-CD44H monoclonal antibody, but that of 2M cells was not. These results suggested that the 2MD3 cell binding to mesothelial cells is regulated by the CD44-hyaluronic acid dependent system. The in vitro binding to submesothelial components and the invasiveness of 2MD3 cells were also inhibited in the presence of anti-CD44H antibody. The in vivo inoculation of 2MD3 cells treated with an anti-CD44H antibody resulted in a significant prolongation of survival time as compared with control mice that were inoculated with 2MD3 cells alone. In conclusion, CD44H was associated with attachment not only to hyaluronic acid on mesothelial cells, but also to peritoneal stromal components. Thus, CD44H may play an important role in cancer cell binding and invasion in the peritoneal dissemination of scirrhous gastric cancer cells.     

Human factor VIII inhibitor alloantibodies with a C2 epitope inhibit factor Xa-catalyzed factor VIII activation: a new anti-factor VIII inhibitory mechanism

Factor VIII (FVIII) inhibitor alloantibodies react with the A2, C2, or A3-CI domains of FVIII and inactivate FVIII activity. We recently demonstrated that an anti-C2 monoclonal antibody with a Val2248-Gly2285 epitope, inhibited factor Xa (FXa)-catalyzed FVIII activation, and that a FXa binding site for FVIII was located within residues Thr2253-Gln2270. In this study, we investigated whether anti-C2 alloantibodies inhibit FXa-catalyzed FVIII activation. Anti-C2 alloantibodies from four patients inhibited FVIII activation by FXa in one-stage clotting assay. Furthermore, analysis by SDS-PAGE showed that all alloantibodies inhibited FVIII proteolytic cleavage by FXa independently of phospholipid. To confirm direct inhibition of FVIII and FXa interaction, we examined the effect of alloantibodies on FVIII binding to anhydro-FXa, a catalytically inactive FXa, in ELISA. All alloantibodies and C2-affinity purified F(ab)'2 preparations inhibited FVIII binding to anhydro-FXa dose-dependently. Our results revealed a new inhibitory mechanism of FVIII, mediated by inhibition of FXa in the presence of anti-C2 alloantibodies.     

Genetic alterations in adenoma-carcinoma sequencing of intraductal papillary-mucinous neoplasm of the pancreas

We investigated the adenoma-carcinoma sequence in intraductal papillary-mucinous neoplasm from the aspect of genetic changes. The formalin-fixed paraffin-embedded tumors and surrounding normal pancreatic tissues from patients with 16 intraductal papillary-mucinous adenoma of the pancreas (IPMA) and 10 intraductal papillary-mucinous carcinoma of the pancreas (IPMC) were provided for DNA extraction after microdissection. SSCP-DNA sequencing analysis demonstrated K-ras mutations at codon 12 in 75% of IPMA and 70% of IPMC, while those at codon 13 were observed neither in IPMA nor IPMC. There were no characteristic K-ras mutation types in IPMA and IPMC and no significant differences in incidence of K-ras mutations between the two categories. The frequencies of p53 mutations analyzed by SSCP-DNA sequencing were not high in IPMA (18.8%) and IPMC (30%), showing no significant difference between them. LOHs of APC in IPMA and IPMC were infrequent (6.3 and 20%, respectively) and showed no significant difference in incidence between the two categories. The LOH frequencies of DCC in IPMA and IPMC were 31.3 and 40%, respectively, and were not statistically different from each other. Taken together, genetic changes such as K-ras, p53, APC and DCC mutations may not be associated with adenoma-carcinoma sequence in intraductal papillary-mucinous neoplasm of pancreas.     

Very low incidence of microsatellite instability in intraductal papillary-mucinous neoplasm of the pancreas

Intraductal papillary-mucinous carcinoma (IPMC) of the pancreas, a new entity of pancreatic cancer with a favorable prognosis, has shown a gradual increase in the number of reported cases. Patients with high-frequency microsatellite instability (MSI-H) tumors have been shown to survive longer than those with low-frequency MSI (MSI-L) or microsatellite stable (MSS) tumors in colorectal and gastric cancer. We investigated whether MSI-H in patients with IPMC can contribute to a good prognosis. The formalin-fixed paraffin-embedded tumors and surrounding normal pancreatic tissues from 10 patients with IPMCs and 16 with intraductal papillary-mucinous adenomas (IPMAs) were provided for DNA extraction after microdissection. Polymerase chain reaction (PCR) was carried out using 8 microsatellite primer marker sets. The mixed PCR samples were analyzed using a genetic analyzer. MSI-H was determined by assessment of microsatellite variations in 3 or more of the 8 tested markers. Immunohistochemical staining of the MSI-responsible proteins hMLH1 and hMSH2 was conducted for both the IPMC and IPMA samples. Ten percent of IPMC harbored MSI-H tumors, whereas no MSI-H tumors were detected in the IPMAs. Thirty percent of IPMC tumors and 25% of IPMA tumors showed MSI-L. All IPMCs and IPMAs showed normal expression of both hMLH1 and hMSH2. MSI-H and loss of hMLH1 and hMSH2 are very rare events in both IPMCs and IPMAs. We conclude that a good prognosis for patients with IPMC is not associated with MSI-H.     

Up-regulated TGF-beta mRNA expression in splenic T cells of high IgA-prone mice: a murine model of IgA nephropathy with glomerulosclerosis

BACKGROUND/AIMS: Recently, we established a high serum IgA-prone inbred (HIGA) mouse strain as a murine model of spontaneous IgA nephropathy by selective mating of high serum IgA ddY mice, and found that they showed enhanced production of glomerular extracellular matrix components with increased expression of TGF-beta mRNA and protein in the kidneys. In this study, we examined the roles of lymphocytes in the development of high serum IgA in this strain. METHODS: We performed flow cytometric analyses of T and B cells in splenic mononuclear cells (SMNCs) from these mice using BALB/c mice as normal controls. We also compared serum TGF-beta1 concentrations and TGF-beta mRNA expression levels in the B-cell-depleted (T-cell-rich) fraction of SMNCs in these mice. RESULTS: HIGA mice showed significantly fewer CD3-positive cells compared with BALB/c mice when young, but not when aged. The CD4/CD8 ratio of HIGA mice was lower than that of BALB/c mice, but this difference was not significant. Although the number of B220-positive cells did not vary significantly, the ratio of surface IgA-positive B cells was significantly increased in both young and adult HIGA mice. The B-cell-depleted SMNCs from HIGA mice exhibited higher levels of expression of TGF-beta and TGF-beta1 mRNA than controls from a young age, which were maintained throughout life, but there were no differences in PDGF, MCP-1 or bFGF expression between these two strains. In contrast to local mRNA expression, serum TGF-beta1 concentration was decreased in HIGA mice compared with BALB/c controls. CONCLUSION: These findings suggest that the mating procedure performed to establish HIGA mice selected for a unique phenotype of local up-regulation of TGF-beta production in the kidneys, as well as T cells that may contribute to both the early and consistently high serum IgA expression and enhanced production of renal extracellular matrix components in HIGA mice. Additionally, TGF-beta1 may act locally, not systemically, in a paracrine or autocrine manner.     

The relationship between stages and biochemical markers of smoking. Workplace-based cross-sectional and longitudinal studies

In order to clarify smokers' characteristics by "Stages of Change" based on Prochaska's transtheoretical model, we conducted cross-sectional and logitudinal studies with biochemical markers of smoking and smoking habits. In a workplace-based sample of 277 male smokers, we examined cross-sectionally the relationships between stages and biochemical markers of smoking which include expired carbon monoxide concentrations and urinary nicotine metabolite concentrations, and smoking habits which include the number of cigarettes smoked per day, yields of cigarettes, inhalation patterns, time to first morning cigarette, and quit attempts in the past. Additionally we examined longitudinally the relationship between stages and expired carbon monoxide concentrations, the number of cigarettes, and yields of cigarettes. In the cross-sectional study there were significant differences among stages on expired carbon monoxide concentrations (P = 0.006), urinary nicotine metabolite concentrations (P = 0.049), the number of cigarettes smoked per day (P = 0.001), and yields of cigarettes (P = 0.042) using analyses of variance. There were also significant differences among stages on time to first morning cigarette (P = 0.018) and quit attempts in the past (P < 0.001) using chi-square tests. In the longitudinal study for each level of elevation in stage during a one-year period, expired carbon monoxide concentrations decreased on an average of 2.3 ppm (P = 0.125) and the number of cigarettes smoked per day decreased on an average of 2.8 cigarettes per day (P = 0.07). However, the yields of cigarettes did not change during the one-year period.     

Expandable metallic stent for the treatment of left-sided colonic obstruction with malignant disease

Recently, it was reported that an EMS (expandable metallic stent) was useful for treatment of colorectal obstruction. In our department, EMSs were used for seven patients with left-sided colonic obstruction with unresectable malignant disease. After these treatments with EMS, their symptoms were improved and they were able to intake food. Stomal formation was avoided except in one patient with severe soiling. In conclusion, EMS is thought to be useful for the improvement of quality of life in the patients with unresectable malignant colonic obstruction.