BACKGROUND AND AIM: To study the effects of Bisphenol A (BPA) and/or diazinon on proliferation of MCF-7 cells. MATERIALS AND METHODS: Estrogen-sensitive and insensitive MCF-7 cells were used to evaluate proliferating effects of BPA and diazinon with concentrations of 1.0×10-12～1.0×10-5mol/L (10 times difference between two consecutive groups). Then the change of proliferating effects after using estrogen receptor inhibitor ICI182780 in each group was assessed. solrent control was establishect in the experiments Combined effects of BPA and diazinon on proliferation of MCF-7 at concentrations of 1.0×10-8, 5.0×10-9 and 2.5×10-9mol/L were evaluated by 2×2 factorial analysis. RESULTS: The highest relative proliferation rate of BPA (2.473) and diazinon (2.167) (compared with solvent control,P<0.05) were all at 1.0×10-8mol/L. After adding ICI182780, the proliferating effects of the drugs on MCF-7 declined. The drugs had no obvious proliferating effects on estrogen-insensitive MCF-7 cells. The combined effects of BPA and diazinon at 1.0×10-8, 5.0×10-9 and 2.5×10-9 mol/L were all antagonistic. CONCLUSION: Bisphenon A and diazinon had obvious estrogen-like activity. They exerted proliferating effects by stimulating the estrogen receptor. The combined effect of BPA and diazinon was antagonistic.

BACKGROUND AND AIM: To explore mutations of PIG-A gene exon2, exon4, exon5 and expression of CD55 and CD59 in granulocytes of patients with aplastic anemia. MATERIALS AND METHODS: Genomic DNA from peripheral blood of 30 aplastic anemia patients and 20 normal controls were extracted, and PIG-A gene exon2, exon4, exon5 were then examined with polymerase chain reaction(PCR), nucleotide sequences were analyzed by bidirectional sequencing after PCR products were purified. The expressions of CD55 and CD59 in granulocytes from peripheral blood of the two groups above were detected by flow cytometry. RESULTS: The mutations of PIG-A gene exon2 including base substitution, deletion, insertion occurred in 11 of 30 aplastic anemia patients , the mutations of exon4, exon5 were also found in five aplastic anemia patients, with only base substitution. Two or more exon mutations were found in 5 aplastic anemia patients, but the normal controls had no in mutations. The expression of CD55 and CD59 were (86.57±5.90)% and (88.17±5.90)%,respectively, in aplastic anemia patients with PIG-A gene mutation;and (91.87±4.79)% and (94.24±3.76)%, respectively, in aplastic anemia patients with no PIG-A gene mutation. Both were significantly decreased(P<0.05)compared with (97.86±1.52)% and (98.82±1.42)%, respectively, in granulocytes of normal control group. CONCLUSION: The PIG-A gene mutations and decreased expressions of CD55 and CD59 in granulocytes may suggest clonal hematopoietic in patients with aplastic anemia.

BACKGROUND AND AIM: ATF3 was down-regulated in esophageal squamous cell carcinoma (ESCC), but the roles of ATF3 in ESCC cells still remained unclear. The purpose of this study was to explore the effect of ATF3 on the proliferation of ESCC cells. MATERIALS AND METHODS: The recombinant expressing plasmid was constructed by inserting the full coding sequence of ATF3 gene into the eukaryotic expressing vector pcDNA3. Then, the expressing plasmid was stably transfected into EC109 cells, an ESCC cell line, and the over-expressing ATF3 cell clones were obtained. Colony formation assay and tumor formation assay in nude mice were used to explore the effect of ATF3 over-expression on the proliferation of ESCC cells. RESULTS: With the over-expression of ATF3, the colony formation ability of EC109 cells was decreased and the tumor formation of EC109 cells in female mice was inhibited. CONCLUSION: Over-expression of ATF3 could inhibit the proliferation of ESCC cells and ATF3 may play important roles in the progression of ESCC.

BACKGROUND AND AIM: To construct the vector pcDNA3.1/SET-TAP carrying tandem affinity purification tag for the differentially expressed protein SET which can be used to further study protein-protein interactions of SET in L-02 liver cells. MATERIALS AND METHODS: Total RNA was extracted from L-02 liver cells: the open reading frame of SET was isolated by using RT-PCR and the TAP gene was amplified from plasmid.Adopting overlap PCR to construct the fusion gene(SET-TAP) through a chimeric primer,then the fusion gene and pcDNA3.1/zeo(+) were digested by EcoRⅠand XhoⅠ.The recombinant vector pcDNA3.1/SET-TAP was constructed through T4 ligase for 16 h at 16 ℃:and then transformed into E.coli DH 5α.After the sequence was confirmed by using double enzyme digestion and sequence analysis:L-02 liver cells were transiently transfected with recombinant vector via Lipofectamine 2000.The expression of fusion protein was preliminary detected by using real-time PCR and western blotting. RESULTS: Results from double enzyme digestion and sequencing showed that the fusion gene was correctly inserted into the vector pcDNA3.1/zeo(+).The pcDNA3.1/SET-TAP could transcribe and express the fusion protein effciently in L-02 liver cells verified by real-time PCR and western blotting. CONCLUSION: The recombinant vector pcDNA3.1/SET-TAP was successfully constructed,laying the foundation to further study the protein-protein interactions of oncoprotein SET in L-02 liver cells treated with trichloroethylene.

BACKGROUND AND AIM: To evaluate DNA damage induced by radiotherapy in nasopharyngeal cancer (NPC) patients. MATERIALS AND METHODS: Comet assay was employed to detect comet frequency , tail length and Olive moment of peripheral blood lymphocytes from 19 NPC patients undergoing radiotherapy at the accumulated dose of 0,2,4,8,10 and 16 Gy. RESULTS: After radiotherapy , the comet frequency and tail length in peripheral lymphocytes rose significantly at the dose of 2 Gy(P<0.01 and P<0.05) whilst Olive moment rose significantly at the dose of 8 Gy(p<0.01). The comet frequency , tail length and Olive moment all increased following radiation dose in this study(0～16 Gy) ,with a clear dose-response relationship. CONCLUSION: Radiotherapy caused DNA damage in NPC patients at low dose, demonstrating dose-effect relationship between radiation dose and comet frequency , tail length and Olive moment.

BACKGROUND AND AIM: The associations between the changes of p63 and p73 in mRNA levels and the BaP-induced DNA damage in H1299 cells and in 16HBE cells were investigated. MATERIALS AND METHODS: Both H1299 cells and 16HBE cells were treated with BaP at various concentrations (8,16,32,64 and 128 μmol/L) for 4 h and 12 h.At the two time points, the levels of MDA、SOD and GSH-Px were measured using the test kits. The mRNA levels of p53,p63,p73,mdm2 and mdm4 genes were detected by RT-PCR assay. DNA damage in the cells were evaluated by Comet assay. RESULTS: At 4 h,enhanced MDA level was observed (P<0.05). However, levels of SOD and GSH-Px were significantly decreased(P<0.05 for both) in both kinds of cells. After 16HBE and H1299 cells were treated for 4 h and 12 h, DNA damage and mRNA expression levels of mdm2 and mdm4 genes increased in a dose-dependent manner(P<0.01 for all). But enhanced mRNA expression level of p53 was observed only at 12 h(P<0.01). In addition,significant up-regulation of p63 and p73 genes in mRNA levels at the two time points were observed in H1299 cells(P<0.05). CONCLUSION: The BaP-induced DNA damage may be associated with enhanced mRNA levels of p63 and p73 genes in the p53-null H1299 cells.

BACKGROUND AND AIM: This study was designed to investigate antioxidative activities and DNA damage to determine toxicological effects of benzo(a)pyrene (BaP) on liver of Sebastiscus marmoratus. MATERIALS AND METHODS: Sebastiscus marmoratus were exposed through a water column to Bap (10; 100; 1 000 ng/L) and sampled at 0 d; 7 d; 25 d; 50 d after exposure and 7 d; 20 d after recovery. Activities of superoxide dismutase(SOD); glutathione S-transferase(GST); reduced glutathione(GSH) content and DNA single strand breaks were detected. RESULTS: Total SOD activity was inhibited after 7 d of exposure; while it was induced after 25 d and 50 d of exposure. GSH content and GST activity were mainly induced. DNA damage was accentuated by longer exposure period and higher exposure concentration. CONCLUSION: Combination of SOD; GST activities and GSH content or detection of DNA single strand breaks in liver of Sebastiscus marmoratus can serve as potential biomarkers to monitor marine pollution of polycyclic aromatic hydrocarbons(PAHs).

BACKGROUND AND AIM:To explore the polymorphisms of δ-aminolevulinic acid dehydratase(ALAD); vitamin D receptor(VDR) and genetic susceptibility of lead posioning in Han; Uygur and Kazak children. MATERIALS AND METHODS: The ALAD and VDR genotypings were determined by PCR-RFLP in 489 Han; 499 Uygur and 525 Kazak individuals from Urumqi city of Xinjiang province. RESULTS: The genotype frequencies of ALAD and VDR showed significant differences in Han; Uygur and Kazak subjects (P<0.01). According to VDR- BsmI、Taq I and Apa I haplotype analysis; haplotype Atb and AtB in Han were considerately decreased in lead poisoning group(P<0.05) while haplotype aTb and ATb significantly increased in lead poisoning group(P<0.01).However; such results were not found in Uygur and Kazak (P>0.05). CONCLUSION: A significant difference was seen in the frequency distribution of ALAD and VDR genotype among the different races. Haplotype Atb and AtB might be protective factors while haplotype ATb and aTb might be risk factors in Han.

BACKGROUND AND AIM: To construct and analyze the gene-expression profiles in the sperms of asthenospermic patients. MATERIALS AND METHODS: 30 semen samples from asthenospermia patients and 20 semen samples from healthy normal adult men were collected; and total RNAs extracted to produce cDNAs probes. Hybridization with Phalanx OneArrayTM contained 30 968 probes was carried out after the labeled cDNAs were purified by PCR Product Purification Kit. RESULTS: Among the 30 968 probes; 394 genes were up-regulated; 437 genes were down-regulated; the others showed no different expressions. Functional cluster displayed that response to external stimulus; defense; stress and inflammation were the most important reasons leading to asthenospermia; and protease inhibitor could also lead to asthenospermia. Furthermore; genes associated with spermatogenesis and negative regulation of apoptosis were very important to maintain high sperm motility. CONCLUSION: Construction of gene-expression profiles in the sperms of asthenospermic patients could help to analyze the molecular mechanism of sperm motility and to study the etiopathogenisis of asthenospermia.

BACKGROUND AND AIM: To illustrate the relationship between the methylation of FHIT and p16 genes and the development of lung cancer. MATERIALS AND METHODS: Methylation of the promoters of FHIT gene and p16 gene was evaluated by methylation-specific PCR in 59 lung cancer tissues; 32 adjacent non-carcinoma tissues and 11 bronchal epithelial squamous tissues. RESULTS: FHIT methylation in lung cancer tissue samples; adjacent non-carcinoma tissue samples were 37.3%(22/59) and 0%(0/32)respectively; with significant difference(P<0.01). Methylation was found in 2 of 11 (18.1%) bronchal epithelial squamous samples. p16 methylation of lung cancer tissue samples; adjacent non-carcinoma tissue samples were 50.8%(30/59)and 0.0%(0/32)respectively; showing a significant difference(P<0.01); Methylation was found in 2(18.1%)of 11 bronchal epithelial squamous samples. FHIT/p16 combined detection of methylation found many more positive tissues in smoking patients than the single gene. There was a significant difference between smokers and non-smokers(P<0.05). CONCLUSION: The 5＇- CpG island methylation of FHIT and p16 was frequent in lung cancer and may be an early event in lung carcinogenesis.

BACKGROUND AND AIM: We studied the mutation effects of sodium sulfite and sodium nitrite in root tip cells of Vicia faba. MATERIALS AND METHODS: The root tips of Vicia faba were treated with different concentrations of sodium sulfite and sodium nitrite (in the concentrations of 25;50;100;200;400 mg/L) with distilled water as control for four hours.The root tips of Vicia faba were stained with carbol fuchsin and examined. The micronucleus rate and the rate of chromosome aberration were determined within root tip cells of Vicia faba. RESULTS: Different concentrations of sodium sulfite and sodium nitrite increased the micronucleus rate and the rate of chromosome aberration in root tip cells of Vicia faba. Both these rates increased with treatment concentration.The micronucleus rate of sodium nitrite was higher than that of sodium sulfite at the some concentration. Sodium sulfite and sodium nitrite induced chromosome to produce various types of chromosomal aberrations: free chromosome;chromosome fragments;chromosome adhesion;lagging chromosome;chromosome bridge;polarized chromosome and ring chromosome. CONCLUSION: Sodium sulfite and sodium nitrite revealed obvious mutation effects and caused certain genetic toxicity in root tip cells of Vicia faba. The effects of sodium nitrite were stronger than that of sodium nitritet.

BACKGROUND AND AIM: To study the effects of polychlorinated biphenyl(PCB) on hippocampal neuron cells in rats. MATERIALS AND METHODS: Bilateral hippocampi were isolated from neonatal Wistar rats; cellular suspensions were prepared under sterile conditions. Rats were divided into control group and three experimental groups; the dosage of PCB were low (1×10-8 mol/L ); middle(1×10-7 mol/L) and high (1×10-6 mol/L);respectively. Light microscope and immunohistochemical technology were used to examine the effects of PCB on structure and function of hippocampal neuron cells in rats. RESULTS: The structural damage of hippocampal neuron cells was related to the dose of PCB; neuron cells were most seriously affected in the high dose group. Axons and dendrites disappeared to remnants; with atrophy of cell bodies. PCB could induce bcl-2 and TGF-β1 expressions in hippocampal neuron cells; bcl-2 expression was detected in control; low dose and middle dose groups; strongest in the low dose group; while bcl-2 expression was not found in the high dose group. TGF-β1 expression was positive in control group. PCB could inhibit TGF-β1 expression in low dose group ; but the expression was up-regulated by PCB in middle dose group. CONCLUSION: PCB could affect the development of hippocampal neuron cells.

BACKGROUND AND AIM: To investigate the effects of thioglycolic acid (TGA) on the nuclear status during progesterone-induced in vitro Xenopus oocyte maturation. MATERIALS AND METHODS: Xenopus oocytes were treated with TGA in vitro at doses of 0, 5, 25 and 125 μg/ml. Germinal vesicle breakdown (GVBD) was checked during culture. Oocytes collected at 8 h were stained with hoechst 33258 to evaluate the chromosome status. RESULTS: Obvious early occurrence of GVBD was found in TGA-treated Xenopus oocytes, as indicated by GVBD50 (time to which 50％ of the oocytes underwent GVBD) and comparison was significant(P<0.05). In immunofluorescence-stained oocytes, no condensed chromosome was seen at GV phase, but metaphase plate was formed at MI phase and metaphase plate and the first polar body(PB1)were all observed at MII phase. However, there were lower MI-MII transition rates and PB1 extrusion rates in TGA-treated groups as compared with control(P<0.05). CONCLUSION: TGA treatment could inhibit Xenopus oocyte maturation as indicated by MI-MII transition, despite GVBD was accelerated by TGA.

BACKGROUND AND AIM To study the safety of bioactive peptide powders. MATERIALS AND METHODS Acute toxicity test of mice; Ames test; micronucleus test of bone marrow PCE cell in mice; sperm shape abnormality test of mice were used. RESULTS Bioactive peptides revealed a LD50>10 g/kg in mice. The results of genetic toxicity test were all negative; including Ames test; micronucleus test and sperm shape abnormality test. CONCLUSION The bioactive peptide powders was a substance with no toxicity and no genotoxicity under our experimental conditions.

BACKGROUND AND AIM: To investigate the genotoxicity of Fructus Schisandrae Chinensis concoction using mouse lymphoma assay(MLA) and mouse bone marrow micronucleus test (MNT). MATERIALS AND METHODS: In MLA; four doses 0.97;1.94;3.88;7.77 mg/ml (crude drug)were used to treat L5178Y cells for 3 hours with and without metabolic activation; then expressed for 2 days. The mutation frequency plates were prepared and incubated for 12 days. Colony size in each plate was scored; and total mutation frequency and percentage of small colony mutants were calculated and assayed. In MNT; three dose levels of 3.13;6.25;12.50 g/kg (crude drug)were used.10 ICR mice (5 males/5 females) in each group were gavaged every 24 h twice and sacrificed after 24 hours of the last dosing. Both femur bones were collected to prepare bone marrow smear. For each mouse; the number of micronucleated polychromatic erythrocytes (MNPCE) in 2000 polychromatic erythrocytes (PCE) was counted; and the mean of rate of MNPCE of each group was calculated. RESULTS: MLA indicated that the total mutation frequency showed a dose-dependent increase with metabolic activation; and was statistically significant (P<0.01) compared with negative control with and without metabolic activation. Percentage of small colony mutants was similar with positive control at high concentrations. MNT indicated the concoction did not show inhibitory effects on bone marrow; and the rate of MNPCE of each group induced did not show significant increase compared with negative control. CONCLUSION: Fructus Schisandrae Chinensis concoction induced tk gene mutation and chromosomal damage in L5178Y cells with and without metabolic activation. It indicated that the test article may have potential genotoxicity for humans. However Fructus Schisandrae Chinensis concoction did not show chromosomal damage of bone marrow in ICR mice; demonstrating no genotoxicity in vivo with metabolic activation under this study condition.

BACKGROUND AND AIM: To evaluate the safety of the compound of lycopene and soybean isoflavone in order to provide some toxicological data for its application. MATERIALS AND METHODS: Acute toxicity test、bone marrow cell micronucleus test、mice sperm abnormality test、Ames test were conducted in accordance with the ＂Technical Standards for Testing & Assessment of Health Food＂. RESULTS: Acute toxicity test of oral dosing showed LD50>10.0 g/kg in mice. Salmonella typhimurium/mammals microsomal enzyme test and micronucleus test of bone marrow cells showed that the compound was safe without mutagenesis. No toxicity was observed in sperm shape abnormality test in mice. CONCLUSION: The compound of lycopene and soybean isoflavone is a non-toxic、 non-mutagenic preparation.

BACKGROUND AND AIM To investigate the toxicity in SD rats fed continuously with monascus yellow pigment. MATERIALS AND METHODS SD rats were divided into four groups (0.23、0.47、0.93 g/kg and a blank control) and fed continually for 90 days. During the experimental period， rats were weighed once every week， general conditions were assessed， and feed consumptions were recorded. On the 45th day， blood was collected from tails to perform complete blood count and biochemical tests. After 90 days， eyeballs were extracted and blood was used for blood count and biochemical tests， livers， kidneys， spleens and testicles were weighed， and pathological examinations of livers， kidneys， spleens， stomach intestines， testicles and ovaries were done. RESULTS The body weight increase and feed efficiency of all dose groups were not significantly different from those of control group(P>0.05).Complete blood count the hemoglobin content， erythrocyte count， leukocyte count and differential in all dose groups were normal.Blood chemical test:serum ALT， AST， urea， creatinine， total cholesterol， triglyceride， glucose， total protein and albumin in all dose groups were normal.Organ/body weight ratios the ratios of kidney/body， spleen/body， liver/body， testicle/body of all dose groups were not significantly different from those of control group(P>0.05).Pathological examination:no special pathological changes were found in the livers， kidneys， stomachs and spleens of rats in all dose groups. CONCLUSION No toxic effects were observed in rats continuously fed with monascus yellow pigment for 90 days， suggesting it could be used as a natural pigment in food production.