BACKGROUND ＆ AIM: To study whether heat shock could induce the formation of γH2AX foci in CHL cells, and the effect of heat shock on N-methyl-N＇-nitro-N-nitrosoguanidine(MNNG)-induced γH2AX foci formation. MATERIALS AND METHODS: The cytotoxic effect of heat shock was evaluated by MTT test.The phosphorylation of γH2AX was assessed using immunofluorescent microscopy and flow cytometry. RESULTS: MTT test showed that heat shock decreased cell viability.There was no significant difference in the number of γH2AX foci between heat-shock treated group and control group as shown by immunofluorescent microscopy. Flow cytometry analysis revealed that pre-heat shock followed by MNNG treatment decreased the mean γH2AX fluorescent intensity from 0.316±0.042 in the MNNG-treatment alone group to 0.194±0.011. CONCLUSION: Heat shock did not induce γH2AX foci formation in CHL cells.In addition, heat shock could inhibit the phosphorylation of H2AX induced by MNNG.

BACKGROUND ＆ AIM: To study effects of polychlorinated biphenyl PCB126 on B(a)P_induced genotoxicity. MATERIALS AND METHODS: HepG2 cells were treated either with different concentrations of PCB126 (0.01, 0.10,1.00 nmol/L) alone,or with different concentrations of PCB126 for 48 h then with B(a)P (50 μmol/L) and PCB126 together for another 24 h. DMSO and 3_MC were used as solvent control and positive control, respectively. EROD activity and micronuclei(MN) formation were analyzed through fluorescence spectrophotometry and a cytokinesis_block micronucleus(CBMN) assay, respectively. The frequencies of MN(‰) and nuclei division index (NDI) were calculated. RESULTS: Comparing to solvent control, EROD levels increased markedly in HepG2 cells treated with PCB126 (0.01, 0.10, 1.00 nmol/L) and B(a)P (50 μmol/L) alone or together, and MN frequencies increased significantly only in HepG2 cells treated with B(a)P alone(P<0.05,P<0.01). Comparing to B(a)P treatment alone, the EROD levels and MN frequencies increased significantly in HepG2 cells treated with B(a)P and PCB126 (0.10,1.00 nmol/L) together(P<0.05,P<0.01). CONCLUSION: PCB126 at certain concentrations showed no genotoxic effect in HepG2 cells, but it might enhance the genotoxic properties of B(a)P.

BACKGROUND ＆ AIM: To study the acute toxicity and genotoxicity of selenium-enriched probiotics. MATERIALS AND METHODS: Acute toxicity test of mice,Ames test,micronucleus test of bone marrow PCE cell in mice, sperm abnormality test of mice were used. RESULTS: The oral acute toxicity study revealed that the LD50 of selenium-enriched probiotics was 18.49 g/kg in mice. The results of genotoxicity tests were all negative, including Ames test, micronucleus test and sperm shape abnormality test. CONCLUSION: Selenium-enriched probiotics showed certain toxicity but no genotoxicity in this study. Thus,selenium-enriched probiotics was safe at the daily-recommended dose(50 mg/person/day) and has the potential for development into functional foods.

BACKGROUND ＆ AIM: To study the possible effect of Bax in vitamin E succinate-induced apoptosis of human gastric carcinoma. MATERIALS AND METHODS: Fluorescent staining was used to detect apoptosis and the mitochondrial transmembrane potential(ΔΨm) of human gastric carcinoma SGC-7901 cells. The expression of Bax was measured by Western Blot analysis after the cells were treated with VES at 5, 10, 20 μg/ml. The translocation of Bax and cytochrome c were determined by Western Blot analysis in mitochondrial and cytosolic-enriched cellular fractions. RESULTS: VES caused apoptosis of SGC-7901 cells. The expression of Bax was increased in whole cell lysate with a dose-dependent relationship. Bax was translocated from the cytosol to the mitochondria. The permeabilization of mitochondrial membranes was increased as determined by the loss of ΔΨm. And cytochrome c was released from the mitochondria to the cytosol. CONCLUSION: VES could induce apoptosis of human gastric carcinoma SGC-7901 cells through mitochondrial pathway by Bax.

BACKGROUND ＆ AIM: To study the cytotoxic activity of CTL and cytokines production of lymphocytes induced by 4-1BBL-transfected colon26 in vitro. MATERIALS AND METHODS: The eukaryotic expression vector pMKITneo/4-1BBL was transfected into colon26 by lipofectamine2000, then the cells with high expression of 4-1BBL were selected by G418. The tumor cell vaccines (TCV) were obtained by treatment with mitomycin(MMC).The TCV were co-cultivated with syngeneic murine cytotoxic T lymphocytes (CTL).Then the cytotoxic activity of CTL and the cytokines production of spleen cells were tested. RESULTS: The colon26 cells transfected with 4-1BBL showed strong expression of 4-1BBL protein on cell surface. Compared with TCV-colon26,the TCV-colon26/4-1BBL cells could induce a more efficient cytotoxic activity of CTL against its parental tumor cell colon26(P<0.01),but not against non-parental tumor cell S180.The TCV-colon26/4-1BBL could significantly enhance the ability of murine spleen cells to produce cytokines(P<0.01). CONCLUSION: The murine colon26 tumor cells transfected with 4-1BBL could induce an efficient anti-tumor immune response.

BACKGROUND ＆ AIM: To study the correlation between EGFR gene fragments and nuclear matrix proteins (NMPs) in human bladder transitional cell carcinoma (hBTCC). MATERIALS AND METHODS: The nuclear matrix proteins were prepared from fresh tissues and T24 culture cells, and total cell proteins was also prepared from fresh tissues of hBTCC. Genomic DNA and nuclear matrix DNA (high concentration sail and gradient DNaseⅠtreated) were separated from hBTCC tissues. Two pair primers were synthesized according to the cDNA sequence of EGFR gene.Two EGFR fragments were amplified by PCR using the two pair primers in genomic DNA and different nuclear matrix DNA.The positive fragments with primersⅠwere also sequenced. The binding capacity of EGFR gene to nuclear matrix in bladder cancer was also measured by Southwestern Blot assay using single stranded end labeled probes prepared by PCR product Ⅱ. RESULTS: Both genomic DNA and nuclear matrix DNA (NM DNA0, NM DNA25 ,NM DNA50 ,NM DNA100) of human bladder carcinoma could amplify 110 bp positive fragment with primersⅡ. However both genomic DNA and nuclear matrix DNA showed a 940 bp positive fragment with primersⅠexcept NM DNA100. The sequence of 940 bp was identical to the DNA sequence of EGFR gene. Southwestern blot analysis demonstrated that EGFR gene fragment (3901-4010nt) was bound to an about 60 kD nuclear matrix protein in both NMPs (tissues and T24 cells) and total cell protein of bladder carcinoma tissues, but not in the cytoplasm. CONCLUSION: Active EGFR gene is strongly bound to the nuclear matrix and nuclear matrix proteins.Nuclear matrix proteins may regulate the high expression of EGFR gene including the course of transcription and post-transcription and/or translation in the human bladder transitional cell carcinoma.

BACKGROUND ＆ AIM： To explore the predictive values of MMP-7 and MMP-9 in malignant transformation of hydatidiform mole. MATERIALS AND METHODS： The protein expressions of MMP-7 and MMP-9 were evaluated by immunohistochemistry in 18 normal villi,49 hydatidiform moles,in which 8 were malignantly transformed moles. The chemiluminescent immunoass was used to measure the level of HCG in 49 hydatidiform moles. RESULTS： The protein expressions of MMP-7 and MMP-9 were significantly higher in malignantly transformed moles than that in normal villi and nonmalignantly transformed moles(P<0.05). There was no statistical difference between normal villi and nonmalignant transformed mole(P>0.05). 8 hydatidiform mole cases which 3 cases persist positive and 5 cases up and down malignant into invasive mole. CONCLUSION： MMP-7 and MMP-9 may be involed in the invasion of the cytotrophoblast cells.Examining the expressions of MMP-7 and MMP-9 could be useful indexes for prediction of gestational trophoblastic disease and offer basis for clinical therapy.

BACKGROUND ＆ AIM: To explore the relationship between expressions of NF-κB and COX-2 in tissue of cervical cancer and HPV16 infection. MATERIALS AND METHODS: The expressions of NF-κB and COX-2 were assessed by immuohistochemical staining in 46 specimens of cervical cancer and 31 specimens of normal cervical tissue. The infection of HPV16 DNA were determined by PCR. RESULTS: The expression rates of NF-κB and COX-2 in cervical cancer appeared significantly higher than that in normal cervical tissue(P<0.01).The expressions of NF-κB and COX-2 in HPV DNA positive group were significantly higher than that in negative group(P<0.05) of cervical cancer. CONCLUSION: There were higher expressions of NF-κB and COX-2 in cervical carcer tissues. The activation of NF-κB and overexpression of COX-2 may be related to HPV 16 infection.

BACKGROUND ＆ AIM: To study the inhibitory effect of extracts of digestive gland on tumor cells using the mice tumor model. MATERIALS AND METHODS: Tumor cells of H22, S180 and MGC-803 were transplanted into mice，the extract was given through the gastrointestinal tract continuously,and the volume of the tumor was measured at different stages. RESULTS: After treatment with the digestive gland extract, the tumor development velocity in tumor bearing mice decreased significantly, and the tumor volume was markedly reduced. The tumour proliferation ratio was less than 60％,the tumor volume reduction ratio was more than 43.7％, and the tumor weight was decreased significantly after different doses of the extracts. CONCLUSION: Extract of digestive gland has obvious suppressive effect on tumor development.

BACKGROUND ＆ AIM: Osthole is a natural coumarin compound, this paper reports on the anti-tumor activities of osthole in vitro and in vivo. MATERIALS AND METHODS: In vitro, anti-tumor assay was performed with human pulmonary adenoma cells A549 and liver cancer cells Bel-7402, MTT method was employed, and half-inhibitory concentration (IC50) were recorded. In vivo, mouse liver cancer H22 was selected and conventional assay method was employed. Three different doses of osthole were administered orally, and tumor inhibitory percentage,thymus gland, spleen indexes and liver weight were measured. RESULTS: IC50 of osthole in A549 and Bel-7402 cells were 67.83 μg/ml and 123.9 μg/ml, respectively. In vivo tumor inhibitory rates in H22 sarcoma was 62％～73％, All osthole-treated groups compared with control animals revealed statistical significance(P<0.01). But osthole groups showed small differences in spleen index, thymus index, and liver weight compared with control mice(P>0.05). Nevertheless differences, when compared with the positive control drug cis-platinum (5 mg/kg dosage), thymus index, spleen index and liver weight were statistically different(P< 0.01). CONCLUSION: Osthole had obvious anti-tumor activities in vitro and in vivo, and did not reveal any toxic effects in experimental animals with the administered doses.

BACKGROUND ＆ AIM： To study the oxidative effect of H2O2 on HepG2 cells in different phases of the cell cycle to explore the apoptosis mechanism of cancer cells under oxidative stress. MATERIALS AND METHODS：Synchronized cells were collected by arresting with TdR, then cells at different phases of cell cycle were treated by 800 μmol/L H2O2 and content of MDA, activities of XOD, 5'NT , anti- O2 and anti-•OH were measured. RESULTS： The content of MDA and activities of XOD, anti-•OH,anti- O2 increased and activities of 5'NT decreased significatly in cells treated with H2O2 . Compared with non-synchronized cells, the content of MDA and activities of XOD, 5'NT，anti-•OH and anti- O2 increased markedly in synchronized S-phase cells and G2/M-phase cells .The content of MDA and activities of XOD, 5'NT， anti- O2 were significantly lower in synchronized G1-phase cells. CONCLUSION： When synchronized cells were treated with H2O2, the degree of lipid peroxidation, activities of enzyme that catalyse production of free radicals， and level of anti-oxidation were all associated with the cell cycle. This may be due to different sensitivities to oxidative damage induced by H2O2 at the different phases of the cell cycle.

BACKGROUND ＆ AIM: To study the genetic damage of acridine on insect cells. MATERIALS AND METHODS: Using different concentrations of acridine to treat spodoptera frugiperda 9 cells(sf9),autographa californica multiple nucleopolyhedrovirus(AcMNPV), to study the effect of the acridine on cell growth and development, the percentage of micronucleus formation and the virulence of AcMNPV. RESULTS: The injurious effects of acridine on Sf9 cells in cell activity and micronucleus percentage revealed that cell division and growth became slow,the surface of the cell became rough, and the micronucleus percentage was10.4‰ after treatment with 5 μg/ml acridine. Some cells had broken membranes or died when treated with 10 μg/ml, and with the micronucleus percentage of 22‰, tri-nucleus, multi-nucleus and nuclear cleavage were also found. When acridine-treated AcMNPV infected the sf9 cells, AcMNPV could proliferate within the cell and form polyhedra, but some abnormal polyhedra like triangle or quadrangle were formed at the same time. CONCLUSION: Using certain dosage of acridine to treat sf9 cells and AcMNPV could cause damage to the cell and the formation of abnormal polyhera of AcMNPV.

BACKGROUND ＆ AIM: The morphological and pathological changes of the esophageal and gastric epithelium in mice treated with different concentrations of Lugol's iodine solution were investigated. MATERIALS AND METHODS: 1％, 3％, 5％, 7％, 9％ of Lugol's iodine solutions were perfused into the esophagus of 120 male Kunming mice. Five mice each were killed at 2 hours, 1 day, 3 days and 7 days after Lugol's iodine treatment. The pathological changes of the esophagus and the stomach were examined. RESULTS: Two hours after esophageal perfusion with 7％ and 9％ Lugol's iodine solution, oedema of different degree was found in the lower part of esophageal mucosa of the mice. Three days after perfusion with 3％，5％，7％ and 9％ Lugol's iodine solution, the esophageal epithelia of the mice were covered by a coating. The coating was a mixture of inflammatory cells and squamous epithelium. It became more obvious with the increasing concentration of Lugol's iodine solution. The esophageal epithelium was red and oedematous after the coating was taken away. Seven days after perfusion, the esophageal epithelium recovered in all the five groups. Two hours after esophageal perfusion with 3％，5％，7％ and 9％ Lugol's solution, oedema, ulceration and haemorrhage of different degrees were found in the gastric mucosa. One day later, the inflammation became more serious.Three days after perfusion, the inflammation subsided gradually. The inflammatory reaction of the gastric epithelium in different groups and at different periods were all concentrated on the anterior wall and the greater curvature, with which the Lugol's iodine solution made contact during perfusion. The inflammation in the fore stomach which was covered with squamous epithelium was obviously less severe than that of gastric mucosa with columnar epithelium. The pathological changes of the esophagus and fore stomach consisted of exfoliation of the squamous epithelium and infiltration of inflammatory cells. The changes of the gastric mucosa showed necrosis, haemorrhage, and infiltration of inflammatory cells on the anterior wall. CONCLUSION: Lugol's iodine solution perfused into the esophagus could damage the mucosa of the esophagus and stomach even at very low concentration. The damage became worse with increase of the concentration.

BACKGROUND ＆ AIM: To observe the changes of heme oxygenase-1 (HO-1) expressed in renal cortex of rats with acute cadmium exposure. MATERIALS AND METHODS: 24 male SD rats were divided randomly into 4 groups. There were a control group and the three experimental groups received daily intraperitoneal injections of CdCl2 (0.4,0.8,1.6 mg/kg) for five days. 0.9％ NaCl substituted for CdCl2 in control. The specimens of kidney were prepared to measure the expression of HO-1 by RT-PCR and immunohistochemistry technique on the fifth day after injection. RESULTS: The physical expression of HO-1 was detected in control group. The levels of expressed mRNA in 0.8 and 1.6 mg/kg groups increased dramatically (P<0.05) compared with the control while the mRNA concentration of 0.4 mg/kg group was equal to the control. A dose-effect relationship between the cadmium exposure and expression of HO-1 mRNA(r=0.97,P<0.05)was also found. Analysis of immunohistochemistry showed that changes of protein expression was more sensitive than mRNA in all three groups compared with the control(P<0.05). CONCLUSION: The acute exposure to cadmium could induce HO-1 expression in renal cortex of rats and this might provide potential protection.

BACKGROUND ＆ AIM: To study the effect of LaCl3 on the expression of CyclinD1 and CDK4 in hepatocellular carcinoma cells. MATERIALS AND METHODS: The in vitro growth of CBRH-7919 cells was observed following treatment with 0.01, 0.10, 1.00 mmol/L LaCl3 for 1,3,5 days, the changes of cell cycle were assessed by flow cytometry. At the same time,changes of CyclinD1 and CDK4 were studied by immunocytochemical analysis. CBRH-7919 cells without LaCl3 were used as control groups. RESULTS: The growth of CBRH-7919 cells was markedly inhibited by 0.10 and 1.00 mmol/L LaCl3 treatment for 3, 5 days,with a time-depentent effect in 1.00 mmol/L group, as compared with control group, the difference was statistically significant(P<0.01). The percentage of CBRH-7919 cells in G0/ G1 phase was significantly increased by 0.10 and 1.00 mmol/L LaCl3 groups for 3, 5 days, as compared with control group (P<0.01). However the expression of CyclinD1 and CDK4 was significantly decreased in 0.10 and 1.00 mmol/L LaCl3 groups for 3, 5 days as compared with control group(P<0.01). CONCLUSION: LaCl3 could obviously inhibit the growth of CBRH-7919 cells by down-regulating CyclinD1 and CDK4 and blocking the cell cycle progression from G1 to S.

BACKGROUND ＆ AIM: To study Bcl-2 and Bax expressions in spermatogenic cells, ultrastructural changes in spermatogenic cells and sperms of mice after exposure to mercury. MATERIALS AND METHODS: 32 male mice were divided into 4 groups,3 groups treated with intraperitoneal mercuric chloride of 0.5,1.0,5.0 μmol/(kg•d) for 5 days,and one normal saline control group.The positive expression rates of Bcl-2 and Bax protein and average optical density in spermatogenic cells by the immunohistochemical method were assayed, and the ultrastructural changes of spermatogenic cells and sperms were assessed by the transmission electron microscope. RESULTS: Bcl-2 protein expression positive rates and average optical density were lower than of normal group(P<0.05).Bax protein expression positive rates and average optical density in 1.0,5.0 μmol/kg groups were higher than those in normal group (P<0.05).The ultrastructure of nuclear membrane,chromatin,mitochondria and sperm mitochondria in three treated groups showed different degrees of injury and the degree of ultrastructural change was more severe with increasing concentration of mercury. CONCLUSION: Mercury exposure caused abnormal Bcl-2 and Bax protein expression, average optical density change and ultrastructural changes in spermatogenic cells and sperms.

BACKGROUND ＆ AIM: To investigate concentrations and mutagenic activities of ambient particulate matter (PM) and polycyclic aromatic hydrocarbons(PAHs) in an industrial area of northeastern China. MATERIALS AND METHODS: The concentrations of PM, PAHs and benzo(a)pyrene [B(a)P] were determined by high liquid chromatography (HPLC) and mutagenicity of the samples were examined by the Ames Salmonella mutagenic assay over two years from September 2000 to August 2002 in a northeast industrial city of China. RESULTS: Concentrations of PM, PAHs and BaA were significantly higher in the industrial area than those in the other sampling area(P<0.05 or P<0.01). Mutagenic activities of PM and its extracted organic matters(EOM) were significantly higher in the industrial area than those in the other sampling area(P<0.05 or P<0.01). CONCLUSION: Concentrations of PM and PAHs were high in the industrial area, and there were strong indirect mutagenic activities.

BACKGROUND ＆ AIM: To explore the influence of different concentrations and different proportions of Wright-Giemsa dye on its staining effect, through application to micronucleus test of HepG2 cells induced by hexavalent chromium[Cr(Ⅵ)]. MATERIALS AND METHODS: HepG2 cells were stained with different proportions of Wright-Giemsa dye and for different stain time, and their influence on staining effects were assessed. RESULTS: Better results could be obtained when cells were stained with a concentration of Wright-Giemsa at 3∶1 for 3-5 minutes. CONCLUSION: Wright-Giemsa stain could be applied to micronucleus test with favorable effects.

BACKGROUND ＆ AIM: To study the mutagenesis of Rubitecan (9NC). MATERIALS AND METHODS: The micronucleus test in human lymphocytes (in vitro) and in bone marrow PCE of mice (in vivo) were used. Induction of transplantated tumor in mice with 9NC was observed. RESULTS: 9NC had no mutagenicity in human lymphocytes within 0～0.187μg/ml and no mutagenicity to murine bone marrow PCE within 0～0.375 mg/kg body weight (P>0.05), but could inhibit the growth of transplantated tumor Hpes (heptome) and S180 (sarcoma) in mice. Mutagenic effects of 9NC was found with dosages of ≥0.25 μg/ml (in vitro) and ≥0.75 mg/kg body weight (in vivo). CONCLUSION: 9NC demonstrated mutatgenic effects at different dose ranges.

BACKGROUND ＆ AIM：To study the genotoxicity of N-Nicotinyl-aspartic acid. MATERIALS AND METHODS:The mutagenicity of N-Nicotinyl-aspartic acid was examined using Ames test, CHO cell chromosome aberration test and micronucleus test. RESULTS： No increase in the number of revertant colonies was found in the Ames test with and without S9 mixture. The micronucleus rates and chromosome aberration rates at all doses were not significantly different from the control group . CONCLUSION： Mutagenicity of N-Nicotinyl-aspartic acid was not observed in this study.

BACKGROUND ＆ AIM: To explore the mutagenicity of Liranaftate. MATERIALS AND METHODS: The mutagenicity of liranaftate was tested by the micronucleus assay, chromosomal aberration test of CHL and the Ames assay. RESULTS: It was found that all of the tests that mentioned above were negative. CONCLUSION: Liranaftate had no mutagenicity in this test.