Ken Haruma

Clinicopathological significance of vascular endothelial growth factor (VEGF)‐C in human esophageal squamous cell carcinomas

The purpose of this study was to investigate the expression of vascular endothelial growth factor (VEGF) ‐C in human esophageal squamous cell carcinomas to elucidate its role in lymph node metastasis and tumor progression. The expression of VEGF‐C and flt‐4 genes was examined in 5 esophageal carcinoma cell lines, 12 fresh biopsy specimens and 48 archival surgical specimens of human esophageal carcinoma tissues by RT‐PCR and immunohistochemistry. Immunohistochemistry using antibodies against CD34 (endothelial cell specific) was also carried out and microvessels were quantified by counting vessels in a 200× field in the most vascular area of the tumor. Of the 5 human esophageal carcinoma cell lines, 4 constitutively expressed VEGF‐C mRNA. In 8 (66.7%) of 12 cases, VEGF‐C mRNA was detected in only tumor tissues but not in normal mucosa by RT‐PCR. There was a significant relationship between VEGF‐C and flt‐4 mRNA expression. Out of the 48 surgical specimens of esophageal carcinomas, 19 (39.6%) and 10 (20.8%) exhibited intense VEGF‐C immunoreactivity in the cytoplasm of many cancer cells and the stromal cells, respectively. In contrast, Flt‐4 was mainly expressed on the lymphatic endothelial cells. Normal and dysplastic esophageal squamous epithelium exhibited no or faint cytoplasmic staining of VEGF‐C. VEGF‐C expression correlated with depth of tumor invasion, tumor stage, venous invasion, lymphatic invasion and lymph node metastasis. Vessel count was significantly higher in the VEGF‐C positive tumors than in the negative tumors. These results overall suggest that VEGF‐C may play a role in tumor progression via lymphangiogenesis and angiogenesis in human esophageal carcinoma.

Endoscopic treatment of submucosal invasive colorectal carcinoma with special reference to risk factors for lymph node metastasis

A clinicopathological analysis of the risk factors for lymph node metastasis was performed in 177 patients with submucosal invasive colorectal carcinoma (CRC). The submucosal deepest invasive portion was histologically subclassified as well (W), moderately (M), or poorly (Por) differentiated. M type was further subdivided into moderately-well (Mw) and moderatelypoorly (Mp) differentiated. The pattern of tumor growth was classified as polypoid growth (PG) and non-polypoid growth (NPG). Lymph node metastasis was detected in 21 (12%) of the 177 patients. Macroscopically, type IIc and IIa+IIc lesions showed a significantly higher incidence of lymph node metastasis (44% and 30%) than type IIa and I (4% and 8%). Regarding the histologic subclassification, Por and Mp lesions showed a significantly higher incidence of lymph node metastasis (67% and 37%) than W and Mw lesions (4% and 14%). NPG tumors showed a significantly higher incidence of lymph node metastasis (29%) than PG tumors (7%). The depth of submucosal invastion and lymphatic invasion (ly) were also significantly correlated with incidence of lymph node metastasis (submucosal scanty (sm-s) invasion 4%, massive invasion 20%; ly(+) 23%, ly(−) 5%). None of the lesions with both sm-s invasion and of W or Mw type showed lymph node metastasis. These results indicate that submucosal invasive CRC with both sm-s invasion and of W or Mw type, which shows no ly, is the appropriate indication for endoscopic curative treatment.

Transfection of interleukin-8 increases angiogenesis and tumorigenesis of human gastric carcinoma cells in nude mice

The growth and spread of tumour cells depends on adequate vasculature. We have previously reported that the expression of interleukin-8 (IL-8) directly correlates with the vascularity of human gastric carcinomas. To provide evidence for a causal role of IL-8 in angiogenesis and tumorigenicity of human gastric cancer, we used the lipofectin method to stably transfect the human TMK-1 gastric carcinoma cells (low endogenous IL-8) with an IL-8 expression vector or control vector. Transfection with IL-8 did not affect the proliferation of cultured cells, yet the culture supernatants of the transfected (but not control) cells stimulated proliferation of human umbilical vein endothelial cells. The IL-8-transfected and control cells were injected into the gastric wall of nude mice. IL-8-transfected cells produced rapidly growing, highly vascular neoplasms as compared to control cells. These results provide direct evidence for the role of IL-8 in the angiogenesis and tumorigenicity of human gastric carcinomas.

Correlation of ratio of serum pepsinogen I and II with prevalence of gastric cancer and adenoma in Japanese subjects

Objective:Gastric cancer (GC) and adenoma (GA) are reported to be related to atrophic gastritis, in which the serum pepsinogen (PG) I level and the PGI/PGII ratio (I/II ratio) are reduced. To verify that the finding of a low PG level increases the risk for GC and GA, we investigated the correlation between low PG levels and the prevalence of GC and GA in individuals.Methods:The 2,039 subjects (734 Japanese men, mean age 68.5 yr, and 1,305 women, mean age 66.7 yr), selected from among 10,996 local residents who underwent health check-ups based on reductions in their serum PG levels, underwent upper gastrointestinal endoscopy.Results:Gastrointestinal endoscopy detected 21 GCs and 15 GAs. The prevalence of GC was higher than that in the residents without low serum PG. The percentage of early stage of GC (90%) was significantly higher than that of GC detected in unscreened residents (56.9%). The prevalence of GC in men was closely and significantly correlated with the I/II ratio (r = 0.935, p= 0.0063), whereas there was less correlation with age (r = 0.842, p= 0.0734). The prevalence of GA was also closely and significantly correlated with the I/II ratio in men (r = 0.881, p= 0.0203), but not with age (r = 0.163, p= 0.7928). In women the prevalence of GC (r 5 0.744, p= 0.090) and GA (r = 0.678, p= 0.1392) did not correlate as strongly with the I/II ratio, although the highest prevalence was seen in the group with the lowest I/II ratio.Conclusion:Our study verified that a low I/II ratio signifies a high risk for GC and GA and that measuring serum PG levels can be used as a screening method for GC and GA.

Interleukin 8 and vascular endothelial growth factor — prognostic factors in human gastric carcinomas?

Gastric carcinoma cells express potent angiogenic factors including vascular endothelial growth factor (VEGF). We previously reported that interleukin-8 (IL-8) acts as an angiogenic factor for human gastric carcinomas. More recently, we found that IL-8 upregulates matrix metalloproteinase-9 (MMP-9) expression and increases invasive activity of gastric carcinoma cells. The purpose of this study was to determine whether the expression of IL-8 and VEGF correlates with clinicopathological parameters in human gastric carcinomas. IL-8 and VEGF expression levels were measured by an enzyme-linked immunosorbent assay (ELISA) in 56 gastric carcinomas and the surrounding normal mucosa. Macroscopic and histopathological tumour findings, presence of metastasis and prognosis were obtained from the patient records and endoscopic, surgical and pathological reports. IL-8 protein levels were higher in most neoplasms than in the corresponding normal mucosal tissue. In contrast, VEGF expression in the tumours was similar to that in normal mucosa. The IL-8 level in the neoplasms correlated significantly with the depth of invasion, venous invasion and lymphatic invasion. VEGF expression in the tumours correlated well with the depth of invasion and lymph node metastasis. No correlation between IL-8 and VEGF expression in the tumours was observed. The survival rates of patients with tumours displaying high IL-8 and VEGF expression levels were significantly lower (P<0.05) than those of patients with tumours displaying low IL-8 and VEGF expression. The results suggest that IL-8 and VEGF may be independent and important prognostic factors in human gastric carcinomas.

Immunoreactive MUC1 expression at the deepest invasive portion correlates with prognosis of colorectal cancer

This study sought to examine the relationship between MUC1 expression at the deepest invasive portion, invasive/metastatic potential, and prognosis of colorectal cancer in relation to cellular proliferation. MUC1 expression was detected immunohistochemically using KL-6 antibody (anti-MUC1 monoclonal antibody) in 100 surgically resected specimens of advanced colorectal cancer. Distinct staining of the luminal surfaces, defined as positive immunoreactive (IR)-MUC1 expression, was seen in more than 30% of the tumor cells at the deepest invasive portion. The proliferating cell nuclear antigen labeling index (PCNA-LI) was also examined in the same areas. IR-MUC1 expression was detected in 71 (71%) of 100 lesions. Lesions with lymphatic or venous invasion showed a significantly higher incidence of IR-MUC1 expression than those without lymphatic or venous invasion (80 vs. 42% and 82 vs. 61%, respectively). Lesions with lymph node metastasis showed a significantly higher incidence of IR-MUC1 expression than those without lymph node metastasis (88 vs. 53%). Lesions with liver metastasis showed a significantly higher incidence of IR-MUC1 expression than those without liver metastasis (92 vs. 59%). Dukes’ stage was also significantly correlated with IR-MUC1 expression. The incidence of IR-MUC1 expression did not significantly differ with regard to histologic subclassification and depth of invasion. There was no significant correlation between IR-MUC1 expression and the PCNA-LI. IR-MUC1 expression at the deepest invasive portion revealed a significant correlation with prognosis; furthermore, in patients with better differentiated lesions, in those with lesions confined to muscularis propria or subserosa (subadventitial) invasion, in those with Dukes’ B and C, or in those undergoing curative resection, IR-MUC1 expression significantly correlated with prognosis. Patients with high PCNA-LI lesions showed a significantly poorer prognosis than those with low PCNA-LI lesions. Only in patients undergoing curative resection, patients with IR-MUC1-positive and high PCNA-LI lesions showed a significantly poorer prognosis than those with IR-MUC1-negative and low PCNA-LI lesions. The significant risk factors in the order of poorer prognosis in patients undergoing curative resection by the multivariate analysis were the histologic grade (moderately–poorly, poorly or mucinous adenocarcinomas), IR-MUC1 expression, and lymph node metastasis. These results indicate that IR-MUC1 expression is an important predictor of the metastatic potential and the prognosis of colorectal cancer, independent of histologic grade, depth of invasion or cellular proliferative activity. Combined analysis of IR-MUC1 and histologic grade, and combined expression of IR-MUC1 and PCNA at the deepest invasive portion are especially useful in predicting colorectal cancer prognosis.

Frequent p53 gene mutations in serrated adenomas of the colorectum.

Serrated adenoma has been recently proposed as a distinct histological lesion of the colorectum. This study examined p53 immunoreactivity, mutations of exons 5–8 of the p53 gene, codon 12 of the Ki‐ras gene by PCR–SSCP analyses, and microsatellite instability in 19 serrated adenomas, ten adenocarcinomas in/with serrated adenomas, 23 hyperplastic nodules, four hyperplastic polyps and 29 tubular adenomas of the colorectum. Eleven of 11 (100 per cent) serrated adenomas had p53 immunoreactivity and all six (100 per cent) adenocacinomas in/with serrated adenomas exhibited moderate to severe p53 immunoreactivity. It was confirmed that 9 of 19 (47 per cent) serrated adenomas and 5 of 10 (50 per cent) adenocarcinomas in/with serrated adenomas harboured p53 gene mutations. On the other hand, no p53 gene mutation was detected in the other colorectal lesions. Meanwhile, 11 (58 per cent) serrated adenomas and six (60 per cent) adenocarcinomas in/with serrated adenomas had Ki‐ras gene mutations, as also did 9 of 23 (39 per cent) hyperplastic nodules, 3 of 4 (75 per cent) hyperplastic polyps, and 12 of 29 (41 per cent) tubular adenomas. Microsatellite instability was detected in one (5 per cent) serrated adenoma and one (10 per cent) adenocarcinoma in a serrated adenoma. The other lesions did not show microsatellite instability. Serrated adenomas had significantly frequent p53 gene mutations compared with hyperplastic lesions or tubular adenomas (p<0·005). On the other hand, they did not exhibit significant differences in mutations of the Ki‐ras gene or in microsatellite instability. Genetic changes were then examined in small parts of serrated adenomas, such as the upper or lower parts of crypts, to determine the extent of gene mutations by using a microdissection technique. Exon 15 of the APC gene and the DCC gene, in addition to the p53 and Ki‐ras genes and microsatellite instability, were analysed. Identical mutations of the p53 gene were found in both invasive adenocarcinomas and adjacent serrated adenomas by direct sequencing, suggesting single clonal origins for those lesions. Mutations of the APC gene and microsatellite instability were heterogeneous in some lesions. No loss of heterozygosity (LOH) of the DCC gene was found. These findings suggest that mutations of the p53 gene are the most characteristic genetic alterations in serrated adenomas, as a relatively early event in a multistep carcinogenic pathway of this type of colorectal lesion, that might be distinct from the ordinary adenoma–carcinoma sequence or from carcinogenesis via mutations of mismatch repair genes. Copyright

Negative association between Helicobacter pylori infection and reflux esophagitis in older patients: case-control study in Japan.

Background. Recent studies have clarified a close association between H. pylori infection and gastritis, peptic ulcer disease, and gastric cancer, but there is little information concerning the relationship between H. pylori infection and reflux esophagitis (RE). We investigated the relationship between H. pylori, RE, and corpus gastritis.

The Role of Endoscopic Findings for the Diagnosis of Helicobacter pylori Infection: Evaluation in a Country with High Prevalence of Atrophic Gastritis

This study examines endoscopic findings in the diagnosis of Helicobacter pylori (H. pylori) in the Japanese population.

Serum pepsinogen in screening for gastric cancer

To establish a sensitive and efficient screening method for gastric cancer using serum pepsinogen, we investigated the characteristics of serum pepsinogen I and II levels and the I/II ratio and their cut-off points. We found that the pepsinogen I level and the I/II ratio were significantly lower in patients with gastric cancer than in control subjects, especially in patients with cancers of the differentiated type, the elevated type, and the depressed type without ulceration. However, sex, depth of invasion, and location of tumor did not correlate with the pepsinogen levels. A suitable cut-off point in screening for gastric cancer was a pepsinogen I level of less than 50 ng/ml and a I/II ratio of less than 3.0, as determined by receiver operator characteristics curves. The sensitivity, the specificity, and the accuracy of detection for all types of gastric cancer were approximately 55%, 75%, and 72%, respectively. If restricted to cancers of the elevated and the depressed type without ulceration, the sensitivity was approximately 85%, and the specificity and accuracy were approximately 76% and 77%, respectively. These results suggest that, in screening for gastric cancer when using pepsinogen levels and morphological examinations, the suitable cut-off point in regard to specificity is as stated above. However, regarding sensitivity, when the pepsinogen method is used alone, a pepsinogen I level of less than 70 ng/ml and a I/II ratio of less than 3.0 is acceptable.