Vascular density of superficial esophageal squamous cell carcinoma determined by direct observation of resected specimen using narrow band imaging with magnifying endoscopy.

Observation of the microvasculature using narrow band imaging (NBI) with magnifying endoscopy is useful for diagnosing superficial squamous cell carcinoma. Increased vascular density is indicative of cancer, but not many studies have reported differences between cancerous and noncancerous areas based on an objective comparison. We observed specimens of endoscopic submucosal dissection (ESD) using NBI magnification, and determined the vascular density of cancerous and noncancerous areas. A total of 25 lesions of esophageal squamous cell carcinoma that were dissected en bloc by ESD between July 2013 and December 2013 were subjected to NBI magnification. We constructed a device that holds an endoscope and precisely controls the movement along the vertical axis in order to observe submerged specimens by NBI magnification. NBI image files of both cancerous (pathologically determined invasion depth, m1/2) and surrounding noncancerous areas were created and subjected to vascular density assessment by two endoscopists who were blinded to clinical information. The invasion depth was m1/2 in 20, m3/sm1 in four and sm2 in one esophageal cancer lesion. Mean vascular density was significantly increased in cancerous areas (37.6 ± 16.3 vessels/mm2) compared with noncancerous areas (17.6 ± 10.0 vessels/mm2) (P < 0.05). The correlation coefficients between vascular density determined by two endoscopists were 0.86 and 0.81 in cancerous and noncancerous areas, respectively. Receiver operating curve (ROC) analysis revealed that the area under the curve (AUC) of vascular density was 0.895 (95% CI, 0.804-0.986). For this ROC curve, sensitivity was 78.3% and specificity was 87.0% when the cutoff value of vascular density was 26 vessels/mm2. NBI magnification confirmed significant increases in vascular density in cancerous areas compared with noncancerous areas in esophageal squamous cell carcinoma. The rates of agreement between vascular density values determined by two independent operators were high.

Phase II study of S-1 plus leucovorin in patients with metastatic colorectal cancer.

BACKGROUND: S-1, a novel oral fluoropyrimidine, is well tolerated in patients with metastatic colorectal cancer (mCRC). The response rate of S-1 for colorectal cancer is high, ranging from 35% to 40%. This study aimed to evaluate the safety and efficacy of S-1 combined with oral leucovorin (LV) to enhance antitumor activity in chemotherapy-naive patients with mCRC. PATIENTS AND METHODS: S-1 was given orally twice daily for two consecutive weeks at a daily dose of 80-120 mg, followed by a 2-week rest period, within a 4-week cycle. LV was given orally twice a day at a daily dose of 50 mg, simultaneously with S-1. RESULTS: Of the 56 patients with previously untreated mCRC, 32 (57%) had partial responses. The median follow-up period was 27.2 months. The median time to progression was 6.7 months (95% confidence interval 5.4-7.9). The median survival time was 24.3 months. There was no treatment-related death or grade 4 toxicity. The most common grade 3 toxic effects were diarrhea (32%), anorexia (21%), stomatitis (20%), and neutropenia (14%). CONCLUSION: S-1 combined with LV therapy demonstrated promising efficacy and acceptable safety in chemotherapy-naive patients with mCRC without the concurrent use of irinotecan, oxaliplatin, or molecular-targeted drugs.

Plasma level of a 5-fluorouracil metabolite, fluoro-beta-alanine correlates with dihydropyrimidine dehydrogenase activity of peripheral blood mononuclear cells in 5-fluorouracil treated patients.

5-fluorouracil (5-FU) is mostly metabolized after administration, and the metabolizing enzyme, dihydropyrimidine dehydrogenase (DPD), seems to be the rate-limiting factor. However, there are few reports on the final metabolite, fluoro-beta-alanine (FBAL). We report here the results of determination of the FBAL level in 5-FU treated patients and the correlation between the FBAL level and the DPD activity in peripheral blood mononuclear cells (PBMCs). Blood samples were collected from 20 patients, who had received continuous intravenous infusion (CIV) of 5-FU (320 mg/m2/24 hr) after resection of colorectal cancer, and the FBAL level was determined by high performance liquid chromatography (HPLC), after derivatizing into o-phthalaldehyde (OPA) and detecting fluorescence. DPD activity was measured in cytosol prepared from PBMCs using HPLC radioassay. The average FBAL plasma level during CIV of 5-FU was 911.0 ng/ml (521.0 to approximately 1834.6 ng/ml) and that of DPD activity in PBMCs was 282.6 pmol/min/mg-protein (145.0 to approximately 568.0 pmol/min/mg-protein). There was a significant correlation between the FBAL level and the DPD activity (r=0.805, p<0.0001). FBAL level in plasma may be useful in predicting the DPD activity in PBMCs, however, further studies are required considering the small number of cases in this study.

The interplay between gastric cancer cell lines and PBMCs mediated by the CC chemokine RANTES plays an important role in tumor progression.

In the present study, we investigated the effect of the RANTES-mediated interaction between gastric carcinoma cell lines and peripheral blood mononuclear cells (PBMCs) in tumor progression. RANTES production in PBMCs stimulated by highly metastatic cancer cell line-conditioned supernatants was higher than in those stimulated by a less metastatic gastric cancer cell line-conditioned supernatant. RANTES receptors were expressed in PBMCs, but not in those cancer cell lines; therefore it was suggested that RANTES might affect PBMCs but not cancer cells. Matrix metalloproteinase (MMP)-9 expression in PBMCs was examined. Similar to RANTES production, MMP-9 expression in PBMCs stimulated by highly metastatic cell line-conditioned supernatants was higher than in that stimulated by a less metastatic cell line-conditioned supernatant. Invasion assays of gastric cancer cell lines were performed. Cancer cells cultured with PBMCs invaded into Matrigel more frequently than those without PBMCs. This invasive activity was highly inhibited by an anti-RANTES antibody. These results suggest that tumor cells can acquire the potential for invasion by cooperating with PBMCs and RANTES plays an important role in the interplay between tumor cells and PBMCs. It is thus thought that RANTES might be a candidate molecular target in the therapeutic strategy for gastric cancers.

Synthetic sulfonolipids deduced from sulfonoquinovosyl diacylglycerols of sea urchin reduces hepatic ischemia-reperfusion injury in rats.

BACKGROUND: In hepatic surgery and liver transplantation, ischemia-reperfusion (I/R) is an unavoidable process, and protection against hepatic I/R injury is a major unresolved problem. In this study, we investigated whether 3-O-(6-deoxy-6-sulfono-beta-D-glucopyranosyl)-1,2-di-O-acylglycerol bound to saturated C18 fatty acids (beta-SQAG9), which was derived from sea urchin intestines, could reduce this injury. This agent was recently reported to have immunosuppressive effects in allogeneic rat skin grafts. MATERIALS & METHODS: Male Lewis rats were divided into two experimental groups. Group 1 rats were injected with SQAG9 (50 mg/kg) into the penile vein 15 minutes before the induction of ischemia and into the portal vein just reperfusion. The same amounts of normal saline were injected into rats in the control group (group 2). Each experimental groups included six rats. Seventy percent hepatic ischemia (20 minutes) was induced by occluding the blood vessels and bile duct with a vascular clamp. For examination of hepatic function, serum levels of aspartate aminotransferase, (AST) alanine transaminase (ALT), and lactic dehydrogenase (LDH) were measured. In addition, histological examination was also assessed. RESULTS: Three hours after reperfusion, the mean plasma concentration of AST, ALT, LDH in group 1 was suppressed compared with group 2. Six hours after reperfusion, the hepatic damage in group 1 was mild in comparison with that in group 2. CONCLUSIONS: Our data demonstrated that SQAG-9 reduced the warm hepatic I/R injury.

Expression analysis of vascular endothelial growth factors and their relationships to lymph node metastasis in human colorectal cancer.

This study was undertaken to determine whether expressions of the vascular endothelial growth factor (VEGF) family (VEGF-A, VEGF-B, VEGF-C, and VEGF-D) are correlated with clinicopathological parameters, with particular reference to lymph node metastasis in colorectal cancer. Total RNA was isolated from 82 surgical specimens of colorectal cancer and matched to normal mucosa with (n = 41) or without (n = 41) lymph node metastasis. The mRNA expression of each VEGF family member was quantified by real-time quantitative (RTQ) RT-PCR assay. VEGF-B and VEGF-C mRNA were significantly higher both in the tumors with lymph node metastasis (p = 0.027 and p = 0.024, respectively) and in tumors with lymphatic invasion (p = 0.042 and p = 0.005, respectively). In contrast, VEGF-D mRNA was down-regulated in tumors with lymphatic involvement (p = 0.047). Among the other clinicopathological factors, we noted that VEGF-A mRNA was higher in tumors with liver metastasis than in those without (p = 0.018) and was higher in tumors with venous invasion than in those without (p = 0.007). The results of this study demonstrate that high levels of VEGF-B, C and low levels of VEGF-D mRNA expression are associated with lymph node metastasis and lymphatic involvement. These results suggest that a balance among VEGF-B, VEGF-C, and VEGF-D might contribute to the lymphangiogenic process and metastasis in colorectal cancer.

Frequent beta-catenin alteration in gallbladder carcinomas.

To investigate the contribution of beta-catenin to the development of gallbladder carcinoma, genetic alteration in beta-catenin gene, ctnnb-1 and subcellular localization of beta-catenin protein were searched. Mutational analysis of exon 3 in ctnnb-1, which encodes the serine/threonine residues for GSK3beta phosphorylation sites, was performed for 21 gallbladder carcinomas affected with/without the pancreaticobiliary malunion, PBM, and 6 non-cancerous tissues affected with PBM. We also analyzed subcellular localization of beta-catenin protein in all cases immunohistochemically. Nucleotide sequencing analysis revealed that none of them carried mutations that altered amino acid residues in the potential GSK3beta phophorylation sites, but one nucleotide substitution was found. We also analyzed subcellular localization of beta-catenin protein in all cases immunohistochemically, and confirmed its accumulation in both the nucleus and cytoplasm in 10 out of 21 cancer tissues, while the non-cancerous tissues which were affected with PBM and histologically diagnosed as hyperplasia or dysplasia displayed intense membranous staining. A significant correlation between cytoplasmic or nuclear beta-catenin immunoreactivity and clinicopathological status of gallbladder carcinomas was found, especially in the poorer histological differentiation grade(p < 0.05). In conclusion our results suggested that beta-catenin alteration might be a minor contributor to the development of gallbladder carcinomas through abnormal Wnt-wingless signalling, however, decreased membranous expression of beta-catenin might be correlated to carcinoma progression through loss of cell adhesive function in E-cadherin-catenin fashion.

Twenty-year changes in the prevalence of overweight in Japanese adults: the National Nutrition Survey 1976-95.

The aim of this study was to describe the 20-year changes in body mass index (BMI; kgm(-2)) and the prevalence of overweight in Japanese adults. Cross-sectional annual nationwide surveys (National Nutrition Survey, Japan) were carried out with a large probability sample of the Japanese population. Data sets of the 1976-95 surveys, comprising 91983 men and 120822 women (> or =20 years of age), were used. The analyses were carried out in age and gender groups, and by residential area according to the size of the municipality (metropolitan areas, cities and small towns). The mean BMI increaseed in men with an increment of +0.44 kg m(-2) 10 years(-1) and slightly decreased in women, by -0.09 kg m(-2) 10 years(-1), after adjustment for age. A decreasing trend of mean BMI was most significant in the female 20-29 years age-group (-0.38 kg m(-2) 10 years(-1)), in contrast to an increasing trend observed in elderly women (60-69 and 70+ years age-groups). The prevalence of preobese (BMI: 25-29.9kg m(-2)) and obese (BMI > or = 30 kg m(-2)) men increased from 14.5% and 0.8%, respectively, in the time-period 1976-80 to 20.5% and 2.01% during 1991-95. The increasing trend was most evident in the youngest age-group (20-29 years) and in those from small towns. The overall prevalence of preobese and obese women did not change during the 20-year study period. The prevalence in younger women decreased, this trend being more prominent in metropolitan areas. Although the prevalence of overweight (BMI > or =25 kg m(-2)) in Japanese men and elderly women has increased in the last 20 years, the mean BMI in younger women, especially those in metropolitan areas, has decreased. A population approach to control obesity and prevent obesity-associated diseases should be mainly focused on men and women older than 40 years of age.

beta-Catenin alteration in cancer of the ampulla of Vater.

To investigate the contribution of beta-catenin to the development of carcinoma of the ampulla of Vater, genetic alterations of beta-catenin gene, CTNNB-1 were searched. Mutational analysis of exon3 in CTNNB-1, which encodes the serine/threonine residues for GSK-3beta phosphorylation sites, was performed on 21 cases of carcinoma of the ampulla of Vater, by means of polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) followed by nucleotide sequencing. We found one deleted mutation at codon 32 to approximately 65 in one case of carcinoma of the ampulla of Vater. We also analyzed subcellular localization of beta-catenin protein in all cases immunohistochemically, and confirmed its accumulation in the nucleus in four cases including in a CTNNB-1 mutated one. This is the first study to show CTNNB-1 mutation and beta-catenin expression in carcinoma of the ampulla of Vater. These results suggested that abnormal Wnt-wingless signaling and in particular beta-catenin alteration caused accumulation of beta-catenin, which might partially contribute to the development of carcinoma of the ampulla of Vater.

Vascular endothelial growth factor C promotes human gastric carcinoma lymph node metastasis in mice.

Vascular endothelial growth factor (VEGF)-C, one of several members of the VEGF family, is a relatively specific lymphangiogenic growth factor. VEGF receptor (VEGFR)-3 (or Flt4) is a VEGF-C receptor with expression restricted to lymphatic endothelial cells. Since the mechanisms by which carcinoma cells metastasize to lymph nodes remain unclear, we constructed a VEGF-C transfectant (AZ-VEGF-C) from the AZ521 human gastric carcinoma cell line, which ordinarily shows little nodal metastatic potential and little VEGF-C expression. We orthotopically implanted transfected tumor cells into the stomachs of nude mice. The number of mice developing lymph node metastases and the number of lymph node metastases per mouse with nodal metastases were higher than with implants of mock-transfected control cells. Specifically, percentages of mice with lymph node metastases were 95.5% (21/22) for AZ-VEGF-C and 29.4% (5/17) for controls (P<0.01), while mean numbers of involved lymph nodes were 3.76 for AZ-VEGF-C and 1.00 for controls (P<0.01). No difference was found between AZ-VEGF-C and controls regarding cell growth and chemotactic responses in vitro, or in volumes of tumors arising from implanted cells. When we performed immunohistochemical staining for VEGFR-3 in these tumors to investigate lymphangiogenesis by VEGF-C, the number of vessels stained for VEGFR-3 in tumors and surrounding tissues was higher for AZ-VEGF-C than for controls. VEGFR-3-positive vessels occupied 14.9/1000 of microscopically examined areas for AZ-VEGF-C, but only 1.30/1000 for controls (P<0.001). Our results suggest that VEGF-C is a specific lymphangiogenic growth factor with an important role in lymph node metastasis.

Adenosquamous carcinoma of the colorectum: report of two cases.

Adenosquamous carcinomas of the colorectum are rare neoplasms. Our experience with two cases is presented in this paper. One patient, who complained of bloody stool, was found to have adenocarcinoma in the sigmoid colon. He received a laparoscopy-assisted sigmoidectomy. The histological examination revealed that the tumor was adenosquamous carcinoma. To date, he has survived six months post operatively without evidence of recurrence. The other patient, who complained of anal bleeding, was found to have rectal adenocarcinoma and received a low anterior resection. Histological examination revealed that the tumor was an adenosquamous carcinoma. He remains alive, with no evidence of recurrence, nine years post operatively. Both cases showed paracolic lymph node metastasis. Because of its very low incidence, the histogenesis, malignancy and prognosis of this disease remain unclear. Thus, further clinical and histological study of this disease entity is required.

Sepsis and cholestasis: basic findings in the sinusoid and bile canaliculus.

It is well known that the liver plays a major role in the clearance of systemic toxemia and is postulated as a regulational organ in the host-defense system. The well-controlled interaction between hepatic parenchymal cells and sinusoidal lining cells including macrophages and Kupffer cells can systematically regulate even critical infections. However, when patients are under the overload condition caused by severe infection, rejection of a transplanted liver and other hapatic dysfunction often are experienced following surgery. Among various signs and symptoms of hepatic dysfunction, progressive cholestasis is recognized as a polarized representation of the irreversible changes in hepatic constitutional cellular functions, especially in hepatic parenchymal cells. Bile canaliculi, the smallest components of the biliary tree, lie between the apical surfaces of adjacent hepatocytes. Septic cholestasis might be a result of disturbance of the total bile canalicular system, i.e., bile secretion, canalicular contraction, and so on. Recently, the molecular biology of the hepatocellular transport system has become better understood, and the pathophysiological condition of cholestasis can be explained as a representation of the intracellular molecular transcriptional system. Cellular changes in surgical cholestasis and molecular findings concerning the bile canaliculus are introduced in this article.

[Weekly low-dose CPT-11 and HCFU for advanced colorectal cancer on an outpatient treatment basis].

We attempted a new outpatient treatment using weekly low-dose CPT-11 for advanced colorectal cancer patients. A 73-year-old female with para-aortic lymph node metastases from advanced rectal cancer was given outpatient treatment for more than 5 months with weekly low-dose CPT-11 and HCFU. CPT-11 was given intravenously at a dose of 20 mg/m2 on day 1 every week. On days 2-7, she was treated by oral administration of HCFU (600 mg). Her serum CEA level decreased and continued to do so for more than 5 months. The size of the para-aortic lymph node was reduced by approximately 40%. There were no adverse effects except leukopenia (grade 2). These results suggest that weekly low-dose CPT-11 and oral HCFU may be an effective therapy on an outpatient basis in cases of advanced colorectal cancer.

[Combination chemotherapy of continuous infusion 5-FU and daily low-dose CDDP in colorectal carcinoma].

Continuous intravenous infusion (CVI) of 5-fluorouracil (5-FU) plus daily low-dose cisplatin (CDDP) was evaluated in 59 patients with advanced and unresectable colorectal carcinoma. The procedure consisted of 5-FU 320 mg/m2/day, CVI and CDDP 2.5-3.5 mg/m2/day, infused for one hour five times a week (low-dose FP therapy) was performed for at least four weeks. The rates of complete and partial response, along with side effects were studied. Also, the effect of low-dose FP therapy on the prognosis of patients with colorectal carcinoma was investigated. The rate of complete and partial response was 57.1%. The frequency of severe side effects (grade 3 and 4) was 10.5%, and no death from side effects was noted. The effect of low-dose FP therapy on the prognosis of advanced and unresectable colorectal carcinoma was studied. It is suggested that this chemotherapy might contribute to the survival of patients with colorectal carcinoma. We designed the chemotherapy of intermittent administration (day by day) of 5-FU 600 mg/m2/day and daily CDDP 2.5-3.5 mg/m2/day (intermittent FP) in order to decrease the rate of side effects and their severity. No cases with grade 3 or 4 side effects were experienced, but the response rate was mostly similar to that of low-dose FP therapy. We concluded that low-dose FP and intermittent FP therapy might be fairly effective for advanced and unresectable colorectal carcinoma.

[Estimation of 99mTc-GSA receptor amount by non-linear 3-compartment model: ligand-receptor binding model without blood sampling].

99mTc-GSA (galactosyl serum albumin) receptor amount (Ro) was estimated by a non-linear 3-compartment model of the ligand-receptor binding without blood sampling. Forward/reverse rate constants and receptor amount were assumed to be variable. Relationship between this parameter and other conventional parameters including ICG R15 (15-minutes retention rate of indocyanine green) was evaluated for 43 surgical candidates with liver tumors. Linear relationships between Ro and HH15 and LHL15 were R2 = 0.73 and 0.72, respectively. Linear relationship between Ro and Rmax, the maximum removal rate, is also good (R2 = 0.84), and the regression line (y = 0.038x + 0.066) was slightly over 0 at y-interception. Linear relationship between Ro and ICG R15, was poor (R2 = 0.39) and relationship was rather a concave shape. Linear relationship of reduction rate between Ro and non-tumor tissue volume of the liver, which assessed at the same day of two weeks after the operation, was y = 1.09x - 0.01 (R2 = 0.82). GSA receptor amount, Ro, seems to change proportional to non-tumor liver tissue volume changing before and after hepatectomy. Complementary to ICG R15, it may be an useful and intuitive parameter for hepatectomy.

Trends of average intake of macronutrients in 47 prefectures of Japan from 1975 to 1994--possible factors that may bias the trend data.

To describe the geographical patterns and trends of macronutrient intake in Japan for the recent 20 years, we analyzed the data sets of the National Nutrition Surveys (J-NNS). First, we calculated regression coefficients for survey year by prefecture for energy, protein, fat, and carbohydrate intake. These results, however, could be affected by the changes in age and sex distributions of the survey samples. Secondly, as the food consumption data were based on household as a unit, we used proportions of the subjects who belong to age-specific groups and female subjects in a sub-sample for each prefecture in a given year for adjustment by general lineal model. As a result, 1% increase in the subjects aged 1-4, 10-19 and 65- years and female subjects was equivalent to the changes in average energy intake by -5.88, +2.27, -1.45 and -1.62 kcal, respectively. After the adjustment for age and sex, number of significant negative coefficients among 47 prefectures decreased for energy and carbohydrate intake, and that of positive coefficients increased for fat intake. This suggested that unadjusted trend data might lead to an overestimation of decreasing trends of energy and carbohydrate intake, and an underestimation of increasing trend of fat intake in the recent 20 years.

GML sensitizes cancer cells to Taxol by induction of apoptosis.

Recently we identified a novel gene, gml, whose expression is regulated in a p53-dependent manner and found that gml expression was correlated with the sensitivity of esophageal cancer cells to anticancer drugs. To further investigate the biological mechanism of gml in determining the chemosensitivity of cancer cells to clinically useful agents, we introduced gml cDNA into TE10, an esophageal cancer cell line that lacks endogenous gml expression. In two resulting stable cell lines which expressed gml cDNA in the absence of wildtype p53, cell death occurred within 6 h after treatment with Taxol. TE10 parent cells or TE10 cells transfected with vector alone displayed relative resistance for 36 h. Induction of gml did not by itself affect viability. Morphological analysis confirmed that the increased chemosensitivity to Taxol conferred by gml was due to apoptosis. These data suggest that reduced expression of gml is likely to be associated with poor response rates to chemotherapy, and that an assay for gml expression might serve a clinical purpose as a predictor of chemotherapeutic sensitivity.

Cloning of P2XM, a novel human P2X receptor gene regulated by p53.

Through cloning of functional p53-binding sites (p53-tagged sites) from the human genome, we isolated a novel gene inducible by wild-type p53. Its cDNA sequence contained an open reading frame encoding a 431-amino acid peptide that showed a significant homology with members of the P2X family. This protein also revealed a similarity to RP-2, a gene activated in thymocytes undergoing programmed cell death. Northern blot analysis showed that it was expressed predominantly in skeletal muscle. Hence, we designated the gene P2XM (P2X specifically expressed in skeletal muscle). P2XM was localized to chromosomal band 22q11, where frequent loss of heterozygosity has been observed in rhabdoid tumors. Although we detected no genetic alteration in the coding sequences, one of four rhabdomyosarcoma cell lines examined had completely lost expression of this gene. Furthermore, a minor splice variant lacking a part of exon 1 that would encode residues corresponding to transmembrane domain M1 was relatively more abundant in two of seven sarcoma cell lines, one of which was derived from a rhabdomyosarcoma, and the other was derived from an osteosarcoma. The results suggest that P2XM may play a significant role in the proliferation and/or differentiation of skeletal muscle cells and that its altered expression may be involved in the development of some sarcomas.

Eighteen cases exposed to sarin in Matsumoto, Japan.

Forty-six patients who were exposed to sarin consulted our hospital because of darkness of vision, and ocular pain, vomiting, dyspnea and headaches on June 27 and 28, 1994. Eighteen patients were admitted and 4 of them were in the critical state. There were 6 features: 1) depression of plasma cholinesterase activity (17 of 18 patients, 94%), 2) hypokalemia (4/18, 22%), 3) depression of triglyceride (12/18, 67%), 4) hypocapnia (5/17, 29%), 5) partial pressure of oxygen (PaO2) <80 mmHg, or requirement of O2 inhalation (15/18, 83%), 6) white blood cells (WBC) >9,000 per mm3 (13/18, 72%). Seventeen patients were discharged from hospital, but one patient is still suffering from akinetic mutism after two years.