bluuebunny/crossref_metadata_2025 huggingface dataset:
Differential Effects of Cigarette Smoking and Alcohol Consumption on the Plasma Levels of Carotenoids in Middle‐aged Japanese Men
Authors: Tsubono Yoshitaka , Tsugane Shoichiro , Gey K. Fred
Published: August 2005

Although several studies have reported that cigarette smoking and alcohol consumption are inversely associated with the plasma level of beta‐carotene even after adjustment for dietary carotene intake, their effects on other carotenoids have not been examined extensively. The authors examined the associations of smoking and alcohol with plasma levels of the five principal carotenoids (beta‐carotene, alpha‐carotene, lutein, lycopene, and zeaxanthin). The subjects were 634 apparently healthy men aged 40–49 years who were sampled randomly from five areas in Japan with varying rates of mortality from gastric cancer. Multiple linear regression analysis controlling for age, serum total cholesterol, serum triglycerides, body mass index, and consumption frequencies of green vegetables, yellow vegetables and fruit, showed that both smoking and alcohol had a significant inverse association with the plasma levels of beta‐carotene and alpha‐carotene; only smoking reduced the level of lutein, and neither smoking nor alcohol significantly affected the level of lycopene or zeaxanthin. With regard to the reduction of beta‐carotene and alpha‐carotene, the effect of smoking was smaller in drinkers than in nondrinkers, and the effect of alcohol was smaller in smokers than in nonsmokers, and significant interactions between smoking and alcohol were observed (P=0.034 for beta‐carotene and 0.026 for alpha‐carotene). The results indicate that the differential effects of smoking and alcohol should be considered when the health effects of plasma carotenoids are examined.
Rare Activation of the Human c‐Ha‐ras Transgene of Mice in Hemangioendothelial Sarcomas and Liver Tumors Induced by Glu‐P‐1
Authors: Inoue Rie , Ushijima Toshikazu , Katami Makoto , Kushida Hiromi , Wakabayashi Keiji , Sato Hidetaka , Asamoto Makoto , Katsuki Motoya , Sugimura Takashi , Nagao Minako
Published: August 2005

A transgenic mouse (Tg), having the human c‐Ha‐ras proto‐oncogene, has been demonstrated to develop hemangioendothelial sarcomas (HESs) which are associated with the transgene mutation, but not to develop liver tumors. In this study, we examined the effects of 2‐amino‐6‐methyldipyrido [1,2‐a:3′,2′‐d] imidazole (Glu‐P‐1), a food‐borne carcinogen, which has been demonstrated to induce HESs and liver tumors in CDF1 mice, on Tg mice. Chronic administration of 0.05% Glu‐P‐1 in the diet induced HESs in Tg (7/17), but not in 18 non‐transgenic mice (N‐Tg). With basal diet, two out of 17 Tg but none of 22 N‐Tg, developed HESs. In contrast, Glu‐P‐1 administration induced liver tumors, both in Tg and in N‐Tg; 16/17 in Tg and 13/18 in N‐Tg. The incidence of hepatocellular carcinomas in Tg was higher than that in N‐Tg (8/17 versus 3/18). With basal diet, only one out of 17 Tg and none of 22 N‐Tg developed liver tumors. The Ha‐ras mutation in tumors developed by the groups administered Glu‐P‐1, was examined. No mutations were detected in the transgene and mouse c‐Ha‐ras genes in all three HESs examined. In contrast, when 29 liver tumors taken from Tg were examined, two mutations of the transgene were detected in two HCCs. No mouse c‐Ha‐ras gene mutations were detected in any of the 47 liver tumors examined, which had developed in Tg and N‐Tg mice. These results suggest that the transgene plays a role in the development of HESs induced by Glu‐P‐1, but not as a result of its mutation. Futher, the transgene plays no significant role in the development of liver tumors induced by Glu‐P‐1, but does play a role in the malignant conversion of some liver tumors, as a result of its mutation.
Detection of ras Gene Mutations in Perioperative Peripheral Blood with Pancreatic Adenocarcinoma
Authors: Nomoto Shuji , Nakao Akimasa , Kasai Yasushi , Harada Akio , Nonami Toshiaki , Takagi Hiroshi
Published: August 2005

Surgeons wish to know of any correlation between an operation and the incidence of metastasis. In perioperative periods, pancreatic cancer cells were identified by detecting mutant K‐ras gene by two‐step PCR and RFLP analysis in blood samples taken from peripheral blood. In no case was K‐ras point mutation detected in blood before operation, although the mutant band was observed in all cases at the time the lesion was resected. Surprisingly, in five of ten cases, positive bands were identified just after laparotomy, before we had reached the primary lesion. In almost all cases, mutant K‐ras was detected until the fourteenth postoperative day. These findings suggest that cancer cells exist in the circulation, and have a potential for hematogenous metastasis during the perioperative period. In conclusion, surgical stress causes hematogenous dissemination of pancreatic cancer cells, and surgeons should employ the appropriate anti‐metastasis therapy in the perioperative period.
Simultaneous Measurement of Unscheduled and Replicating DNA Synthesis by Means of a New Cell Culture Insert DNA Retention Method: Rapid Induction of Replicating DNA Synthesis in Response to Genotoxic Carcinogens
Authors: Okumura Ataru , Tanaka Takuji , Mori Hideki
Published: August 2005

In order to measure simultaneously replicating DNA synthesis (RDS) and unscheduled DNA synthesis (UDS) in rat hepatocytes responding to exposure to carcinogens, a new method, namely the “cell culture insert DNA retention (CDR)” method, was developed. All CDR procedures for cell culture, digestion of cytoplasm and retention of DNA were performed on membranes attached to cell culture containers. Four subgroups of primary cultures of hepatocytes prepared from rats were exposed to a genotoxic or non‐genotoxic carcinogen with or without 10 mM hydroxyurea and incubated for 4 h with 10 μCi/ml [3H]thymidine. The membranes were then processed for both liquid scintillation and autoradiography. Among seven tested chemicals, three genotoxic agents, 3,2′‐dimethyl‐4‐aminobiphenyl, 2‐acetylaminofluorene and diethylnitrosamine, and two non‐genotoxic carcinogens, nafenopin and phenobarbital, induced RDS within 4 h after the exposure, indicating that these carcinogenic agents induce cell proliferation in non‐proliferating rat hepatocytes prior to the emergence of genotoxic changes. Several indices were devised to characterize the genotoxicity of the tested chemicals. The induction patterns obtained showed a wide variation in the individual characteristics of carcinogen‐induced genotoxicity and mitogenicity in the early phase of initiation. This is the first report of simultaneous measurement, by using a combination of autoradiography and liquid scintillation, of UDS and RDS induced in rat hepatocytes. The described CDR approach will be useful for risk assessment and characterization of carcinogenic and tumor‐promoting agents.
Anti‐tumor Promoting Activity of Canventol and Its Synthetic Analogs through Inhibition of Protein Isoprenylation
Authors: Komori Atsumasa , Okabe Sachiko , Suganuma Masami , Kerr Michael A. , Busch‐Petersen Jakob , Oh Lynette M. , Zhuo Jincong , Kamali Kannangara G. S. , Zou Xianglong , Tius Marcus A. , Fujiki Hirota
Published: August 2005

Canventol, a synthetic compound, is a new inhibitor of tumor promotion on mouse skin by okadaic acid. We previously reported that canventol acts by inhibiting both protein isoprenylation and tumor necrosis factor‐α (TNF‐α) release. In this study we examined the potencies of 10 newly synthesized canventol analogs through their effect on mevalonate metabolism, and then examined 3 representative analogs for inhibition of protein isoprenylation. Since canventol in vitro did not directly inhibit farnesyl protein transferase or geranylgeranyl protein transferase‐I, the effects of canventol and its synthetic analogs on the fate of [3H]mevalonate in cells were studied. Canventol at 500 μM changed the ratio of newly synthesized sterols (cholesterol and lathosterol) to ubiquinones from 0.7 to 8.2 in NIH/3T3 cells which had previously been labeled with [3H]mevalonate, suggesting that the altered pattern of mevalonate metabolism is associated with inhibition of protein isoprenylation in the cells. We named this ratio the inhibition of protein isoprenylation index (IPI index). The 10 analogs showed a wide range of IPI indices. Two analogs, S3 and S9 had effects similar to, or stronger than, canventol. Three analogs, C44, C46 and C47, with lower IPI indices, inhibited tumor promotion on mouse skin slightly less than canventol itself did. This study shows that inhibition of protein isoprenylation in the cells, indicated by an increase in the IPI index, is a new biomarker for estimating inhibition of tumor promotion.
Expression and Roles of Heat Shock Proteins in Human Breast Cancer
Authors: Yano Masao , Naito Zenya , Tanaka Shigeo , Asano Goro
Published: August 2005

Heat shock proteins (hsps) are thought to play important roles in the cell cycle and various processes of carcinogenesis. Therefore, we evaluated the expression of hsps, mainly hsp90 and hsp70, in human breast cancer tissues. Hsp90α mRNA was expressed at much higher levels in the cancerous tissue than in the non‐cancerous tissue. In addition, a close correlation between hsp90α mRNA expression and the proliferating‐cell‐nuclear‐antigen labeling index (PCNA LI) was observed for the cancerous tissue. These findings suggest that increased expression of the hsp90α isoform may play a role in cell proliferation. On the other hand, hsp90β mRNA expression was significantly higher in poorly differentiated carcinomas than in well differentiated carcinomas of the breast. The intracellular localization of hsp70 was consistent with that of ubiquitin. In specimens showing hsp70 in the nucleus, the PCNA LI was significantly high, Hsc73 mRNA, a member of the hsp70 family, was also expressed at higher levels in cancerous tissues associated with a high PCNA LI than in non‐cancerous tissues. These results suggest that hsp90α may play a role in cancer cell proliferation and that hsp90β may contribute to cell differentiation and structural constitution. In addition, hsp70, especially hsc73, is related to ubiquitin and seems to be a marker for cancer proliferation.
Inhibitory Effects of Tea Catechins, Black Tea Extract and Oolong Tea Extract on Hepatocarcinogenesis in Rat
Authors: Matsumoto Natsuki , Kohri Toshiyuki , Okushio Kazuo , Hara Yukihiko
Published: August 2005

Inhibitory effects of individual tea catechins ((‐)‐epicatechin, (‐)‐epigallocatechm, (‐)‐epicatechin gallate, (‐)‐epigallocatechin gallate), black tea extract and oolong tea extract on hepatocarcinogenesis were investigated. Male F344 rats received a single dose of diethylnitrosamine (200 mg/kg, i.p.), and thereafter phenobarbital (0.05%) was administered in the drinking water for a period of 6 weeks. Tea catechins, black tea extract or oolong tea extract were given during the entire experimental period, during only the initiation period or during only the promotion period. All four tea catechins, black tea extract and oolong tea extract (0.05 or 0.1%) significantly decreased the number and area of preneoplastic glutathione S‐transferase placental form‐positive foci in the liver. These results suggest that tea catechins, black tea extract and oolong tea extract have a chemopreventive action against hepatocarcinogenesis.
Mutational Analysis of the p53 and K‐ras Genes and Allelotype Study of the Rb‐1 Gene for Investigating the Pathogenesis of Combined Hepatocellular‐Cholangiocellular Carcinomas
Authors: Imai Yasuo , Oda Hideaki , Arai Masami , Shimizu Seiichiro , Nakatsuru Yoko , Inoue Tohru , Ishikawa Takatoshi
Published: August 2005

Because combined hepatocellular‐cholangiocellular carcinoma is rare and its biological features and pathogenesis have not been well established, we investigated alterations of the p53, K‐ras and Rb‐1 genes, as well as expression patterns of carcinoembryonic antigen and keratin, in seven combined hepatocellular‐cholangiocarcinoinas out of 557 hepatocellular carcinomas autopsied at Tokyo University during 30 years. Mutations of the p53 gene were found in two cases, at codon 244 (GGC to TGC) in the cholangiocellular carcinoma component of case 1 (mixed type, showing an intimate intermingling of both elements) and at codon 234 (TAC to AAC) in both components of case 5 (combined type, consisting of contiguous but independent masses of both elements). Mutation of the K‐ras gene (codon 12, GGT to GAT) was seen only in the cholangiocellular carcinoma component of clinically apparent double cancer, case 6. Allelic alteration of the Rb‐1 gene was observed in two cases, deletion of both alleles in the hepatocellular carcinoma component of case 3 (combined type) and replication error of the same pattern in both components of case 4 (mixed type). Immunohistochemical analysis showed that the hepatocellular carcinoma components of five cases (cases 2, 3, 5, 6, 7) were immunoreactive for keratin, suggesting biliary epithelial transformation. In four of the five cases (cases 3 and 5 combined, case 7 mixed and case 6 double cancer), cholangiocellular carcinoma components were also positive for keratin. These results suggest that both components of combined hepatocellular‐cholangio‐carcinoma have the same genetic and phenotypic character and might have arisen from the same origin in some cases.
Promoting Effect of Unilateral Nephrectomy on Urinary Bladder Carcinogenesis in Rats
Authors: Saikawa Shigeki , Kanamaru Hiroshi , Li Benyi , Matsukawa Shigeru , Okada Kenichiro
Published: August 2005

The effect of unilateral nephrectomy on the growth of bladder tumors was investigated in male F344 rats treated with 0.05%N‐butyl‐N‐(4‐hydroxybutyl)nitrosamine(BBN) for 6 weeks by means of macroscopical and microscopical examinations. Unilateral nephrectomy 4 weeks after the initiation of BBN administration significantly increased both the number and the volume of visible tumors per rat as compared with the sham‐operated group. Histological examination also revealed that the numbers of papillonodular hyperplasia and cancer per basement membrane were significantly increased by unilateral nephrectomy. The results imply a growth‐enhancing effect of unilateral nephrectomy on carcinogenesis of the rat bladder.
Tumor Necrosis Factor Production and Colon Cancer
Authors: Ito Hisashi , Yagita Akikuni , Fujitsuka Mitsuharu , Atomi Yutaka , Tatekawa Isao
Published: August 2005

Tumor necrosis factor (TNF) production in B10 mice exhibiting H‐2 gene heterogeneity and in C3H/ He mice differing in lipopolysaccharide (LPS) responsiveness was investigated following stimulation with OK‐432. TNF‐producing capacity in these mice was H‐2‐restricted, while their LPS responsiveness was independent of the gene. TNF production in humans was found to be HLA‐B antigen‐restricted. An investigation was then made of the effect of endogenous TNF induction with OK‐432 on the survival rate of colorectal cancer patients. Patients in the TNF‐producing group showed a trend toward having a better prognosis as compared to those in the TNF‐nonproducing group. Cancer therapy formulated with consideration of host responsiveness to OK‐432 may afford greater therapeutic benefit and may prolong survival.
Lentinan Enhances Sensitivity of Mouse Colon 26 Tumor to cis‐Diamminedichloroplatinum(II) and Decreases Glutathione Transferase Expression
Authors: Murata Tetsuo , Hatayama Ichiro , Kakizaki Ikuko , Satoh Kimihiko , Sato Kiyomi , Tsuchida Shigeki
Published: August 2005

We investigated the influence of a combination of lentinan, a biological response modifier, and cis‐diamminedichloroplatinum(II) (CDDP) on the growth and glutathione S‐transferase (GST) content of colon 26 tumor to examine whether lentinan represses GST expression and enhances the therapeutic effects of CDDP. Female CDF1 mice inoculated subcutaneously with transplantable colon 26 adenocarcinoma cells (1X106/mouse) received intraperitoneal administrations of lentinan, CDDP, or the two drugs in combination, on days 10, 14, 17 and 21 after the inoculation. On day 24, tumor weights (estimated from their length and width) were significantly lower in the CDDP+lentinan group (2.7±1.3g) than in the CDDP alone group (4.3±0.7g, p<0.05), both values being less than in the nontreated control group (7.2±1.5g). The major GST form of colon 26 tumor was identified as GST‐II, the Pi class form, and a minor form as GST‐III belonging to the Mu class. Both GST‐II and GST‐III values on day 24 were significantly decreased in the lentinan alone (0.90±0.29 and 0.26±0.11 μg/mg protein, respectively) and lentinan+CDDP groups (0.98±0.22 and 0.29±0.07 μg/mg protein), as compared with the control levels (1.39±0.20 and 0.52±0.11 μg/mg protein). However, these values were not different between the CDDP alone and lentinan+CDDP groups. Neither tissue interleukin (IL)‐6, glutathione nor platinum values were different between the two groups. IL‐6 values were elevated in about half of the samples treated with lentinan or CDDP and exhibited a modest inverse correlation with GST‐II levels (r= ‐ 0.46). A GST inhibitor, ethacrynic acid, enhanced the sensitivity of cultured colon 26 cells to CDDP, suggesting the possible involvement of GST in modulating the cytotoxicity of CDDP to this cell line. These results indicated that lentinan administration decreases tissue GST‐II and GST‐III contents and enhances the sensitivity of colon 26 tumor to CDDP.
Intracellular Hyperthermia for Cancer Using Magnetite Cationic Liposomes: In vitro Study
Authors: Shinkai Masashige , Yanase Mitsugu , Honda Hiroyuki , Wakabayashi Toshihiko , Yoshida Jun , Kobayashi Takeshi
Published: August 2005

‘Magnetite cationic liposomes (MCL)’ were developed as a means to generate intracellular hyperthermia. Affinity of the MCL to glioma cells was ten times higher than that of magnetite‘neutral’ liposomes due to the electrostatic interaction based on the positive charge of the MCL. Heat generation of the MCL was studied using agar phantoms and small pellets of rat glioma cells. When a high‐frequency magnetic field, 118 kHz, 384 Oe was applied to glioma cells in the presence of MCL, the glioma cell pellet of 80 μl (5.4 mm in diameter) was heated to over 43°C and all the cells died after 40 min irradiation owing to the hyperthermic effect. The terminal temperature of the cell pellet was proportional to the pellet volume when other parameters were constant. It thus appears that the MCL can heat a tumor of more than 80 μl in volume to above 42°C.
Serological Evaluation of Soluble CD44 in Renal Cancer
Authors: Kan Masaharu , Kanayama Hiro‐omi , Naruo Seiichi , Tsuji Masahito , Kojima Keiji , Kurokawa Yasushi , Kagawa Susumu
Published: August 2005

In this study, we examined the feasibility of using elevated serum CD44 concentration as an indicator in renal cancer. We performed enzyme‐linked immunosorbent assays using 63 sera obtained from 47 patients with renal cancer and 16 healthy controls and evaluated the clinico‐pathological parameters. The concentration of soluble CD44 standard (sCD44std), indicating the concentration of all circulating CD44 isoforms, was significantly higher in renal cancer patients than in normal individuals (745±170 ng/ml vs. 563±159 ng/ml, P=0.001). The concentration of soluble CD44 splice isoforms sharing exon v6 (sCD44v6) was also higher in the same patients (287±121 vs. 220±59, P=0.056). However, there were no correlations between the concentrations of sCD44std or sCD44v6 and clinico‐pathological parameters such as grade, stage, histological type, tumor size and growth type. The ratio of sCD44std/sCD44v6 was higher in the rapid growth‐type cancers than in the slow growth‐type cancers (3.95±2.12 vs. 2.63±0.82, P=0.014). These findings suggested that the serum concentration of unknown soluble CD44 isoforms not sharing exon v6, which are present in sCD44std, increases in patients with rapid growth‐type cancers. These findings indicated that sCD44std and sCD44v6 are not useful indicators of tumor burden and metastasis in patients with renal cancer, but that an unknown sCD44 isoform(s) plays a role in the biological behavior of the rapid growth‐type cancers.
Suppression of Human Pancreatic Cancer Growth in BALB/c Nude Mice by Manumycin, a Farnesyl:protein Transferase Inhibitor
Authors: Ito Toshio , Kawata Sumio , Tamura Shinji , Igura Takumi , Nagase Toshihiko , Miyagawa Jun‐ichiro , Yamazaki Eiji , Ishiguro Hiroshi , Matsuzawa Yuji
Published: August 2005

Activating mutations of Ki‐ras have been detected in most human pancreatic adenocarcinomas. Since Ras protein requires farnesylation to function, we investigated the effects of manumycin, a potent farnesyl:protein transferase inhibitor, on the growth in nude mice of a human pancreatic cancer cell line, MIA PaCa‐2, with a point mutation in the Ki‐ras gene. Tumor‐bearing mice received intraperitoneal injection of 1 or 5 mg/kg manumycin daily for 5 days, or 2 mg/kg manumycin daily for 2 weeks. Growth of inoculated tumors was significantly inhibited by the treatment. The treatment significantly (P<0.05) lowered the numbers of bromodeoxyuridine‐incorporating tumor cells. Manumycin did not have apparent hepatotoxicity in vivo. Farnesyl:protein transferase inhibitors could offer a new approach for cancer chemotherapy.
Chemopreventive Effects of 3‐Phenylpropyl Isothiocyanate on Hamster Lung Tumorigenesis Initiated with N‐Nitrosobis(2‐oxopropyl)amine
Authors: Nishikawa Akiyoshi , Furukawa Fumio , Ikezaki Shinichiro , Tanakamaru Zen‐yo , Chung Fung‐Lung , Takahashi Michihito , Hayashi Yuzo
Published: August 2005

The chemopreventive effects of 3‐phenylpropyl isothiocyanate (PPITC) were investigated in N‐nitrosobis(2‐oxopropyl)amine (BOP)‐initiated hamsters. A total of 120 female 5‐week‐old hamsters were divided into 6 groups. Animals in groups 1–3, each consisting of 30 hamsters, were twice sc injected 7 days apart as an initiation treatment. Hamsters in groups 1 and 2 were respectively given 100 μmol and 10 μmol of PPITC by gavage 2 h prior to each BOP treatment. Animals in group 3 were treated with BOP alone, serving as an initiation‐positive control. Animals in groups 4–6, each consisting of 10 hamsters, were given 100 μmol or 10 μmol of PPITC alone, or non‐treated, thus being available as matched negative controls to groups 1–3. At termination (experimental week 51 after the first BOP injection), the incidences of lung adenomas and/or adenocarcinomas were significantly decreased in groups 1 and 2 as compared to the group 3 value (P<0.01). The combined lung tumor incidences were inhibited by 94% and 59% at 100 and 10 μmol doses, respectively. The inhibitory effects of PPITC were thus dose‐dependent. The data for multiplicity of lung tumors dramatically illustrated the inhibitory effects of PPITC, and there were also statistically significant differences in the chemopreventive effect between 100 μmol and 10 μmol PPITC treatments. On the other hand, the PPITC treatments did not significantly modulate the development of neoplastic lesions in the pancreas, liver and kidney, although the treatments did show inhibitory tendencies, except on the liver lesions. Under the present experimental conditions, PPITC itself did not exhibit tumorigenicity or apparent toxicity. The results in the present study thus clearly indicate that PPITC has an effective chemopreventive action on BOP‐induced lung tumorigenesis in hamsters.
Features of DNA Oligonucleosomal Fragmentation in Human Tumor Cell Lines and Its Detection by Flow Cytometry: Utility and Limitations
Authors: Kikuchi Hiroaki , Ujiie Shigeki , Wakui Akira , Yokoyama Akiko , Kanamaru Ryunosuke
Published: August 2005

Cultured HL‐60, HeLa S3 and WiDr cells were treated with various doses of ethanol, then subjected to flow cytometry and gel electrophoresis of cellular DNA. On electrophoresis of DNA from HL‐60 cells treated with 0.5 or 1.0 mM ethanol, a ladder pattern was recognized after 3 h. At higher doses of ethanol (2.0 and 5.0 mM), a smear pattern resulted. On flow cytometry, however, A0 cells (lower fluorescence level than G0+ G1 cells) were noted from 0.5 to 5,0 mM ethanol. The observation of A0 cells at higher doses indicated loss of DNA after random DNA degradation. HeLa S3 and WiDr cells were partially detached from flasks after administration of ethanol and separated into adherent and non‐adherent categories. In DNA from non‐adherent HeLa S3 cells treated with 0.5 mM ethanol, a ladder pattern was observed after 24 h. On flow cytometry, prior to the appearance of A0 cells, an accumulation in the G2+M‐phase became obvious after 3 h. Increased mitotic indices indicated that this phenomenon was due to M‐phase arrest. Adherent HeLa S3 cells showed no DNA Oligonucleosomal fragmentation or A0 cells. These findings indicate that detection of A0 cells by flow cytometry is not proof of cell death by DNA Oligonucleosomal fragmentation.
A Defect in Cell–to–cell Adhesion via Integrin–Fibronectin Interactions in a Highly Metastatic Tumor Cell Line
Authors: Abe Yoshiko , Tsutsui Tateki , Mu Jie , Kosugi Atsushi , Yagita Hideo , Sobue Kenji , Niwa Ohtsura , Fujiwara Hiromi , Hamaoka Toshiyuki
Published: August 2005

We investigated the role of integrin–fibronectin (FN) interactions in tumor cell adhesion. Two cloned tumor cell lines designated OV–LM (low–metastatic) and OV–HM (high–metastatic) were isolated from a murine ovarian carcinoma, OV2944. OV–LM and OV–HM cells exhibited high and low RGDS–sequence–dependent adhesiveness to FN, respectively. Both lines expressed comparable levels of α5 and αv integrins, which are capable of reacting with RGDS on FN. To compare the functions of these integrins between the two tumor lines, the signaling mechanism following FN stimulation was examined. Significant levels of phosphorylation of focal adhesion kinase(FAK)were detected in bothOV–LM and OV–HM cells before FN stimulation. Whereas the level of FAK phosphorylation was appreciably enhanced in OV–LM cells stimulated with FN, stimulation of OV–HM cells with FN induced a reduction in the FAK phosphorylation in association with a significant decrease in theamount of FAK protein in the soluble compartment of cell lysates. A difference in the deposition of FN on the cell surfacewas also observed between the two types of tumor lines; OV–HM cells had an appreciably smaller amount of FN than OV–LM. Consistent with the functional abnormality of the integrin–FAK system and the smaller amount of FN on OV–HM, this clone exhibited a reduced cell–cell adhesion in the in vitro cell aggregation assay. Namely, OV–LM cells displayed a time–dependent increase in the formation of cell aggregates, whereas most OV–HM cells remained single. The formation of aggregates by OV–LM cells was inhibited by addition of RGDS peptide. These results indicate that the highly metastatic clone, OV–HM, exhibits a decreased capacity of cell–cell adhesion mediated by integrin–FN interactions and suggest that this defect is mainly due to the dysfunction of integrins/FAK rather than a decrease in the amount of integrins expressed on tumor cells.
All‐trans‐retinoic Acid‐dependent Inhibition of E‐Cadherin‐based Cell Adhesion with Concomitant Dephosphorylation of β‐Catenin in Metastatic Human Renal Carcinoma Cells
Authors: Ryuto Masahiro , Jimi Sei‐ichiro , Ono Mayumi , Naito Seiji , Nakayama Yoshifumi , Yamada Yuji , Komiyama Sohtaro , Kuwano Michihiko
Published: August 2005

We previously described an in vitro invasion assay model, using a monolayer of vascular endothelial cells grown on collagen gel, that mimics the metastatic abilities of the highly metastatic human renal carcinoma cell lines, MM‐1,3 and 8 and their poorly metastatic counterparts, SN12C and Q‐8. MM‐1, 3 and 8 cells were observed to penetrate the monolayer of vascular endothelial cells and grew in a spreading or scattering manner with loose cell‐cell contact on collagen gel or on vascular endothelial cells. SN12C and Cl‐8 cells failed to penetrate and grew in a clustering manner with tight cell‐cell contact. Treatment with all‐trans‐retinoic acid (ATRA) at non‐toxic concentrations induced clustering or growth of MM‐1, 3 and 8 cells on collagen gel or on vascular endothelial cells with tight cell‐cell contact, and inhibited penetration. The clustering induced by ATRA was virtually blocked in the presence of anti‐E cadherin antibody. E‐Cadherin and β‐catenin were each localized mainly at the cell‐cell adherent junctions of colonizing cell populations that had been treated with ATRA. While the cellular levels of E‐cadherin and β‐catenin did not change significantly following ATRA treatment, the tyrosine residue of β‐catenin was rapidly dephosphorylated. The concomitant administration of Na vanadate, an inhibitor of tyrosine dephosphorylase, inhibited both the ATRA‐induced clustering and the dephosphorylation of β‐catenin tyrosine. ATRA‐induced clustering of MM‐3 cells may be linked to the state of tyrosine phosphorylation of β‐catenin.
Growth‐inhibitory Effects of N, N‐Diethyl‐2‐[4‐(phenylmethyl)phenoxy]‐ethanamine‐HCl Combined with Cisplatin on Human Ovarian Cancer Cells Inoculated into Nude Mice
Authors: Hiramatsu Hisakazu , Kikuchi Yoshihiro , Kudoh Kazuya , Kita Tsunekazu , Tode Takehiko , Nagata Ichiro
Published: August 2005

In 5‐day incubation of an estrogen receptor‐negative human ovarian cancer cell line (KF) with diethyl‐2‐[4‐(phenylmethyl)phenoxy]ethanamine‐HCl (DPPE), the concentration of DPPE required for 50% inhibition of KF cell proliferation (IC50) was 1.7 μM. The IC50 of DPPE for inhibition of protein kinase C (PKC) activity was 3.0 μM, a similar value to those of other antiestrogens such as tamoxifen and clomiphene. DPPE also inhibited phosphorylation of mitogen‐activated protein kinase in KF cells. When treatment with DPPE was started 7 days after inoculation of KF cells into nude mice, 50 mg/kg DPPE alone resulted in a significant growth retardation in the early stage of tumor growth. Although 25 mg/kg DPPE showed a similar effect to 2 mg/kg cisplatin (CDDP), the combination had the most marked tumor growth‐inhibitory effect. Nude mice treated with combinations of CDDP and DPPE survived significantly longer than not only untreated, but also CDDP‐alone‐treated mice, while 50 mg/kg but not 25 mg/kg DPPE alone had an effect comparable to that of 2 mg/kg CDDP alone. If treatment with DPPE was begun from the day after tumor inoculation, the inhibitory effect of DPPE was further enhanced, especially when combined with CDDP. If treatment with DPPE was started in nude mice with a lower tumor burden, 25 mg/kg as well as 50 mg/kg DPPE had a similar effect to 2 mg/kg CDDP, in terms of survival. When DPPE was combined with CDDP, the effect was significantly enhanced, compared to that of either alone. These treatments could be done without any adverse side effect. Thus, we conclude that DPPE has an antiestrogen action and its tumor growth‐inhibiting activity is enhanced on administration in combination with CDDP.
Somatic Hypermutations in the VH Segment of Immunoglobulin Genes of CDS‐positive Diffuse Large B‐Cell Lymphomas
Authors: Kume Masaaki , Suzuki Ritsuro , Yatabe Yasushi , Kagami Yoshitoyo , Miura Ikuo , Miura Akira B. , Morishima Yasuo , Nakamura Shigeo , Seto Masao
Published: August 2005

De now CDS‐positive (CD5+) diffuse large B‐cell lymphoma (DLBL) has recently been identified as constituting a homogeneous subgroup with distinct clinicopathologic and genotypic characteristics, but its origin remains to he elucidated. Previous studies by sequence analysis of the variable region of the immunoglobulin heavy chain (VH) have shown that CDS5 B‐cell malignancies such as mantle cell lymphoma (MCL) and B‐cell chronic lymphocytic leukemia (B‐CLL) cells represent pre‐geminal center (pre‐GC) stage B cells in contrast with the post‐GC stage of most DLBLs, which show somatic hyper mutations in VH genes. In the present study, we investigated the VH sequence of de novo CD5+ DLBL to clarify whether CD5+ DLBL represents the pre‐GC stage, as do other CD5+ B‐cell malignancies, or the post‐GC stage, as is typical of DLBL. All eight cases (four CDS4 DLBL and four CDS‐negative (CD5−) DLBL) examined by us showed somatic hypermutatiohs in the VH segment and two of the CD5− DLBL cases showed intra‐clonal diversity, suggesting that CD5+ DLBLs were derived from the same maturation stage as CD5− DLBL, but were distinct from the other indolent CD5+ B‐cell lymphomas of B‐CLL and MCL. These data suggest that de novo CD5+ DLBLs do not merely lie within a continuous spectrum with B‐CLL and MCL, but represent a biologically distinct variant within the diagnostic framework of diffuse large B‐cell lymphoma.
Photodynamic Effect of Polyethylene Glycol–modified Fullerene on Tumor
Authors: Tabata Yasuhiko , Murakami Yoshiyuki , Ikada Yoshito
Published: August 2005

Fullerene (Cm) efficiently generates singlet oxygen when irradiated with light, and thus should have a photodynamic effect on tumors, if it is accumulated in the tumor tissue. To explore tumor targeting of CU, we chemically modified the water–insoluble C60 with polyethylene glycol (PEG), not only to make it soluble in water, but also to enlarge its molecular size. When injected intravenously into mice carrying a tumor mass in the hack sulicutis, the C60–PEG conjugate exhibited higher accumulation and more prolonged retention in the tumor tissue than in normal tissue. The conjugate was excreted without being accumulated in any specific organ. Following intravenous injection of CorPEG conjugate or Photofrin® to tumor–bearing mice, coupled with exposure of the tumor site to visible light, the volume increase of the tumor mass was suppressed and the C60 conjugate exhibited a stronger suppressive effect than Photofrin. Histological examination revealed that conjugate injection plus light irradiation strongly induced tumor necrosis without any damage to the overlying normal skin. The antitumor effect of the conjugate increased with increasing irradiation power and Cm dose, and cures were achieved by treatment with a dose of 424 μg/kg at an irradiation power of 107 J/cm2. These findings indicate that PEG–modified cm is a candidate agent for photodynamic tumor therapy.
RAPID COMMUNICATION
Authors: Ohata Takeji , Fukuda Kazunori , Takahashi Mami , Sugimura Takashi , Wakabayashi Keiji
Published: August 2005

Although nitric oxide (NO) is an important biological mediator, its excessive production in inflammation is thought to be a causative factor for cellular injury and, over the long term, cancer. In the present study, the effects of several fatty acids on NO production in murine macrophage cell line RAW264 cells stimulated with lipopolysaccharide were examined. Suppression of NO production was observed with the ω3 polyunsaturated fatty acids (PUFAs), docosahexaenoic acid, eicosapentaenoic acid and α‐linolenic acid, in a dose‐dependent fashion. In contrast, no inhibition was observed with ω6 PUFA (linoleic acid),ω9 PUFA (oleic acid) or a saturated fatty acid (stearic acid). Western and northern blot analyses suggested that suppression of the induction of inducible NO synthase gene expression is responsible for the inhibition of NO production by ω3 PUFAs. The inhibitory effect of ω3 PUFA on NO production in activated macrophages could contribute to their cancer chemopreven‐tive influence.
Induction of Specific Cytotoxic T Lymphocytes against Autologous Brain Tumor by Crossreactive Allo‐tumor Cell Stimulation
Authors: Saitoh Seiji , Kurisaka Masahiro , Mori Koreaki , Maeda Nagamasa , Fujimoto Shigeyoshi
Published: August 2005

Cytotoxic T lymphocytes (CTL) against autologous malignant brain tumor were generated in peripheral blood lymphoid cells (PBL) prepared from a patient with a malignant brain tumor by stimulation of the cultured PBL for 7 days with attenuated Crossreactive malignant melanoma (MM2) cells pretreated with mitomycin C. The Crossreactive MM2 cells were effective for antigen stimulation for CTL induction in place of autologous glioblastoma cells, which are difficult to expand in culture. The optimal ratio between nylon wool‐passed T lymphocytes and nylon wool‐adherent accessory cells to induce CTL in the patient's PBL was found to be 25 to 1. In vitro ‐activated CTLs induced by MM2 were cytotoxic not only to MM2, but also to the autologous tumor cells in an HLA class I‐restricted manner, and their surface phenotype was found to be CD3+ and CD8+. CTL therapy using cross‐reactive allogeneic tumor cells as the stimulator could be clinically valuable to treat malignant brain tumors.
Antibodies to Human Papillomavirus 16, 18, 58, and 6b Major Capsid Proteins among Japanese Females
Authors: Matsumoto Koji , Yoshikawa Hiroyuki , Taketani Yuji , Yoshiike Kunito , Kanda Tadahito
Published: August 2005

Among genital human papillomaviruses (HPVs), the so‐called high‐risk (HPV 16, 18, etc.) and intermediate‐risk (HPV 58, etc.) viruses are believed to be etiologically associated with cervical cancer. To estimate the extent of infection with common HPVs among Japanese females, we examined 328 sera from healthy donors (201) and patients with cervical intraepithelial neoplasia (GIN) (22), cervical cancer (67), and condyloma acuminatum (CA) (38) for IgG antibodies against L1 capsid protein by enzyme‐linked immunosorbent assay using virus‐like particles of HPVs 16, 18, 58 and 6b (low‐risk) as antigens. Antibodies recognizing conformational epitopes were found in the sera from both the patients and the healthy donors. The prevalences of anti‐HPV 16,18, and 58 antibodies in the sera from the patients with CIN (45%) and cervical cancer (49%), and that of anti‐HPV 6b in the sera from the patients with CA (55%), were significantly higher than those in the sera from the age‐matched healthy donors (12%, 14%, and 23%, respectively). Anti‐HPV 16 was not found in some of the sera from patients with HPV 16‐DNA positive CIN or cervical cancer, suggesting that HPV infection may not always induce production of anti‐capsid antibodies or that the level of antibodies may not always be maintained until development of CIN or cancer. Some of the sera contained antibodies against more than one type of HPV, suggesting that the donors had been infected with different HPVs. The type‐specific antibodies against capsid L1 protein of one type of HPV may not be able to prevent infections with other types of HPVs.
S‐Methylcysteine and Cysteine Are Inhibitors of Induction of Glutathione S‐Transferase Placental Form‐positive Foci during Initiation and Promotion Phases of Rat Hepatocarcinogenesis
Authors: Takada Nobuyasu , Yano Yoshihisa , Wanibuchi Hideki , Otani Shuzo , Fukushima Shoji
Published: August 2005

S‐Methylcysteine (SMC) occurs in a variety of plants, including Allium sativum, Phaseolus vulgaris, and Cruciferae. In this study, we synthesized five organosulfur compounds (OSCs), SMC and four analogs, and examined their modifying effects on diethylnitrosamine‐induced neoplasia of the liver in male F344 rats, using the medium‐term bioassay system of Ito (Ito test) based on the two‐step model of hepatocarcinogenesis. In addition, we investigated the modifying effects of SMC and cysteine on the initiation stage of rat hepatocarcinogenesis. Carcinogenic potential was scored by comparing the numbers and areas of induced glutathione 5‐transferase placental form (GST‐P)‐positive hepatocellular foci. All OSCs examined had a tendency to decrease the number of GST‐P‐positive foci when given in the promotion stage of the Ito test, and in particular SMC and cysteine exerted significant inhibitory effects. When given during the initiation stage, these two OSCs also significantly inhibited focus formation. Regarding the mechanism underlying the inhibitory effects of SMC and cysteine, measurement of ornithine decarboxylase in SMC‐ and cysteine‐treated liver tissues after partial hepatectomy (PH) revealed a significantly reduced activity, and the proportion of hepatocytes positive for proliferating cell nuclear antigen was significantly decreased by SMC or cysteine administration. Moreover, examination of the expression of the early response proto‐oncogenes, c‐fos, c‐jun, and c‐myc, after PH demonstrated down‐regulated induction of c‐jun mRNA transcripts by SMC, sustained for an eight‐hour period. Our results support the view that SMC and cysteine are chemopreven‐tive agents for rat hepatocarcinogenesis and that their intake may he of importance for cancer prevention.
Protection by αG‐Rutin, a Water‐soluble Antioxidant Flavonoid, against Renal Damage in Mice Treated with Ferric Nitrilotriacetate
Authors: Shimoi Kayoko , Shen Bingrong , Toyokuni Shinya , Mochizuki Rika , Furugori Michiyo , Kinae Naohide
Published: August 2005

The protective effect of aG‐Rutin against ferric nitrilotriacetate (Fe‐NTA)‐induced renal damage was studied in male ICR mice. Fe‐NTA induces renal lipid peroxidation, leading to a high incidence of renal cell carcinoma in rodents. Administration of αG‐Rutin (50 μmol as rut in/leg) by gastric intubation 30 min after i.p. injection of Fe‐NTA (7 mg Fe/kg) most effectively suppressed renal lipid peroxidation. Repeated i.p. injection of Fe‐NTA (2 mg FeAg/day for the first 3 days and 3 mg Fe/ kg/day for 12 days, 5 days a week) causes subacute nephrotoxicity as revealed by induction of karyomegalic cells in renal proximal tubules. A protective effect was observed in mice given αG‐Rutin 30 min after each Fe‐NTA treatment. To elucidate the mechanism of protection by αG‐Rutin, the pharmacokinetics and hydroxyl radical‐scavenging effect of αG‐Rutin were investigated by HPLC analysis and by electron spin resonance (ESR) spin trapping with 5,5‐dimethyl‐l‐pyrroline‐N‐oxide (DMPO), respectively. When mice were given αG‐Rutin (50 μmol as rutin Ag) by gastric intubation, rapid absorption into the circulation was observed. The plasma concentration of äG‐Rutin reached the highest level 30 min after oral administration and then decreased to the control level within 60 min. äG‐Rutin inhibited the formation of DMPO‐OH in a concentration‐dependent manner. Further, chelating activity of äG‐Rutin to ferric ions was shown by spectrophotometric analysis. These results suggest that absorbed äG‐Rutin works as an antioxidant in vivo either by scavenging reactive oxygen species or by chelating ferric ions and this serves to prevent oxidative renal damage in mice treated with Fe‐NTA.
Interferon‐γ‐inducing Factor Gene Transfection into Lewis Lung Carcinoma Cells Reduces Tumorigenicity in vivo
Authors: Fukumoto Hisao , Nishio Makoto , Nishio Kazuto , Heike Yuji , Arioka Hitoshi , Kurokawa Hirokazu , Ishida Tomoyuki , Fukuoka Kazuya , Nomoto Taisuke , Ohe Yuichiro , Saijo Nagahiro
Published: August 2005

To investigate the immunoregulatory effect of murine mterferon‐γ‐inducing factor (mIGIF), we transfected Lewis lung carcinoma (IXC) cells with a mammalian expression vector containing the mIGIF complementary DNA. The culture medium of the transfectant cells stimulated interferon‐γ (IFN‐γ) production by spleen cells in vitro in the presence of anti‐CD3 antibody and markedly potentiated the effect of interleukin‐12 (IL‐12) on IFN‐γ production by spleen cells. mIGIF transfectant cells showed reduction of tumorigenicity and induction of an in vivo immimo‐protective effect against the parental LLC cells. To examine the combined effect of systemic administration of recomhinant IL‐12 (rIL‐12) and local mIGIF on the tumorigenicity, mice were challenged with LLC or transfectant cells on day 0, and the tumor‐bearing mice were injected with 50 ng of rIL‐12 intraperitoneally from day 7 to 11. Systemic rIL‐12 showed an anti‐tumor effect. However, mIGIF gene expression did not potentiate this effect of systemic rIL‐12 in vivo.
Cytotoxin Genes of Helicobacter pylori in Chronic Gastritis, Gastroduodenal Ulcer and Gastric Cancer: An Age and Gender Matched Case‐Control Study
Authors: Matsukura Norio , Onda Masahiko , Kato Shunji , Hasegawa Hirokazu , Okawa Kei‐ichi , Shirakawa Takasi , Tokunaga Akira , Yamashita Kiyohiko , Hayashi Akio
Published: August 2005

Helicobacter pylori (H. pylori) infection is involved in many gastrointestinal diseases, such as chronic gastritis (CAG), peptic ulcer and gastric cancer (GCA). Both host factors and H. pylori strain differences may contribute to differences in the diseases. Thus, we conducted an age and gender matched case‐control study of 35 patients each with CAG, gastric ulcer (GUL), duodenal ulcer (DUL) and gastric cancer (GCA) to examine the role of strain differences of the H, pylori cytotoxin genes cagA and vacA in these diseases. We employed polymerase chain reaction to examine the gastric juice for H. pylori DNA. The test was positive for 26 (74.3%) CAG, 29 (82.9%) GUL, 28 (80.0%) DUL and 27 (77.1%) GCA patients, showing no statistically significant difference among the diseases (P= 0.84). cagA and vacA genes (picked up by using a vacAl+vacA2 primer pair which detected nonvariable regions of the vacA gene) were detected by PCR in the H. pylori DNA‐positive cases as follows: CAG, 92.3% and 76.9%; GUL, 100% and 86.2%; DUL, 89.3% and 89.3%; GCA, 92.6% and 85.2%, respectively. No statistically significant differences were found in the frequencies of these cytotoxin genes in H. pylori ‐positive cases among the various gastric diseases (P=0.39 for cagA and P=0.64 for vacA).
Expression of Globo‐series Gangliosides in Human Renal Cell Carcinoma
Authors: Saito Seiichi , Orikasa Seiichi , Satoh Makoto , Ohyama Chikara , Ito Akihiro , Takahashi Toshiko
Published: August 2005

Gangliosides have been shown to be involved in development, differentiation, oncogenesis, and cancer progression. We investigated immunohistochemical expression of globo‐series gangliosides in human renal cell carcinoma (RCC) and whether their expression is related to the clinical course. The expression of globo‐series gangliosides was evaluated in fresh‐frozen sections of 55 primary renal tumors and 8 metastatic deposits using monoclonal antibodies RM1 and RM2, which define mono‐sialosyl and disialosyl galactosylgloboside, respectively. The immunorcaetivity of primary tumors to RM1 and/or RM2 was correlated with the clinicopathological data. Cumulative incidence of metastasis detected at initial diagnosis and during the follow‐up period was significantly higher in the cases whose primary tumors were RMl/RM2‐positive (RM1 and/or RM2‐positive) than in the RM1/ RM2‐negative (neither RM1 nor RM2‐positive) cases (P<0.05). During the follow‐up period, metastasis developed in none of the RMl/RM2‐negative cases which had not shown metastasis at initial diagnosis. High nuclear grade was observed only in the RMl/RM2‐positive cases. The RM1/RM2‐positive rate of the metastatic deposits was higher than that of the primary tumors. Furthermore, a metastatic deposit obtained from one of the cases whose primary tumors were equivocal for RM1/ RM2 was extensively stained by RM1 and RM2. These results indicate that globo‐series gangliosides may be one of the biochemical indicators related to the metastatic potential of human RCC.