In vitro Antigenotoxicity of Ulva rigida C. Agardh (Chlorophyceae) Extract against Induction of Chromosome Aberration, Sister Chromatid Exchange and Micronuclei by Mutagenic Agent MMC

Objective To determine the in vitro possible clastogenic and cytotoxic activities of Ulva rigida crude extracts （URE）, and identify their antigenotoxic and protective effects on chemotherapeutic agent mitomycine-C （MMC）. Methods Anti-clastogenic and anti-genotoxic activities of Ulva rigida crude extracts （URE） were studied using chromosome aberration （CA）, sister chromatid exchange （SCE）, and micronuclei （MN） tests in human lymphocytes cultured in vitro. Results The chromosome aberration, sister chromatid exchange or micronuclei tests showed that URE at concentrations of 10, 20, and 40 lag/mL had no clastogenic activity in human lymphocyte cell culture. Three doses of URE significantly decreased the number of chromosomal aberrations and the frequencies of SCE and MN when compared with the culture treated with MMC （P〈0.0001）. Conclusion Although URE itself is not a clastogenic or cytotoxic substance, it possesses strong antigenotoxic, anti-clastogenic, and protective effects on MMC in vitro.

Antimutagenic potential of curcumin on chromosomal aberrations in Allium cepa

Turmeric has long been used as a spice and food colouring agent in Asia. In the present investigation, the antimutagenic potential of curcumin was evaluated in Allium cepa root meristem cells. So far there is no report on the biological properties of curcumin in plant test systems. The root tip cells were treated with sodium azide at 200 and 300 μg/ml for 3 h and curcumin was given at 5, 10 and 20 μg/ml for 16 h, prior to sodium azide treatment. The tips were squashed after colchicine treatment and the cells were analyzed for chromosome aberration and mitotic index. Curcumin induces chromosomal aberration in Allium cepa root tip cells in an insignificant manner, when compared with untreated control. Sodium azide alone induces chromosomal aberrations significantly with increasing concentrations. The total number of aberrations was significantly reduced in root tip cells pretreated with curcumin. The study reveals that curcumin has antimutagenic potential against sodium azide induced chromosomal aberrations in Allium cepa root meristem cells. In addition, it showed mild cytotoxicity by reducing the percentage of mitotic index in all curcumin treated groups, but the mechanism of action remains unknown. The antimutagenic potential of curcumin is effective at 5 μg/ml in Allium cepa root meristem cells.

Mathematical Modeling of Carcinogenesis Based on Chromosome Aberration Data

Objective： The progression of human cancer is characterized by the accumulation of genetic instability. An increasing number of experimental genetic molecular techniques have been used to detect chromosome aberrations. Previous studies on chromosome abnormalities often focused on identifying the frequent loci of chromosome alterations, but rarely addressed the issue of interrelationship of chromosomal abnormalities. In the last few years, several mathematical models have been employed to construct models of carcinogenesis, in an attempt to identify the time order and cause-and-effect relationship of chromosome aberrations. The principles and applications of these models are reviewed and compared in this paper. Mathematical modeling of carcinogenesis can contribute to our understanding of the molecular genetics of tumor development, and identification of cancer related genes, thus leading to improved clinical practice of cancer.

Induction of Chromosomal Aberrations by Propoxur in Mouse Bone Marrow Cells

Propoxur is a widely used dithiocarbamate insecticide. In this study, the clastogenic effect of propoxur has been evaluated using chromosomal aberration assay in mouse bone marrow cells. Single i. p. administration of propoxur, at 25 mg/kg b.wt., a maximum tolerated dose (MTD) and 12 .5mg/kg b.wt (50% of MTD) have significantly induced different types of aberrations after 24 h of treatment. The aberrations were dose and time dependent and reached a maximum after 24 h of exposure. The sresult suggest a genotoxic potential of propoxur.

MODULATION OF THE FREQUENCY OF HUMANCYTOMEGALOViRUS-INDUCED CHROMOSOMEABERRATIONS BY CAMPTOTHECIN

The effects of selected DNA repair inhibitors on the frequency of human cytomegalovirus (HCMV)-induced chromosome aberrations in human peripheral blood lymphocytes (PBLs) were evaluated. Trealment of HCMV-infected PBLs with camptothecin (0.05 to 0.3μg/ml), an inhibitor of topoisomerase I, for 30 hr resulted in a significant (P<0.01) synergistic enhancement of the frequency of HCMV-induced chromosome damage, on the other hand, a significant increase in the frequency of chromosome damage was not noted for infected PBLs treated with either 3-aminobenzamide (3-AB) (3 to 30μg/ml), an inhibitor of poly (ADP-ribose) poiymerase, ur novobiocin 3 to 30 |ig/ml) an inhibitor of topoiso-merase I or excision repair processes for 30 hr. chromatid-type breaks including chromosome exchanges were the predominant type of chromosome aberrations observed in the HCMV-infected cells treated with camptothecin suggesting that HCMV infection is associated with the induction of single-strand DNA breaks. Furthermore, these findings suggest that HCMV infection does rol inflict dircc: DNA damage which is repaired through 3-AB- or novobiocinsensitive pathways.Human cytomegalovirus (HCMV) is a common pathogen which irferfs about 80% of the world's population causing, for the most part, persistent subcliniclil infeclions (Wellcr, 1971). A relatively small percentage of otherwise healthy immunologically competent people experience clinical HCMV disease (Cohen et al., 1985). Generalized HCMV infection, however, is the bane of individuals whose immune system is compromised (Schooley, 1990) Rubin, 1990). Molecular epidemiological studies strongly suggest that HCMV is one of the most frequent cause of congenital infections and that these infections result in a high incidence of birth defects and developmental abnormalities (Alford et al., 1990). Several studies (Nachtigal et al., 1978; Luleci et al., 1980; AbuBakar et al., 1988) have shown that HCMV infectior can result in an increase in the frequency of chromosome aberrations. Since the ability to cause chromosome damage may be significant in the induction of birth defects and possibly in HCMV-induced malignancy, we undertook the present investigation to evaluate the mechanisms by which HCMV may cause chromosome damage. In ths study, the effects of selected DNA repair inhibitors on the frequency of HCMV-induced chromosome aberrations were evaluated. The results indicae that the presence of camptothecin, but nut 3-aminobenzamidc (3-AB) ro novobiocin, results in a concentration-dependent increase in the frequent) of chromosome aberrations in HCMV-infected peripheral blood lymphocytes (PBLs). Accordingly, it is proposed that HCMV-induced chromosome damage in PBLs does not substantially involve DNA repair activities sensitive to inhibition of poly ADP-ribosylation or excision repair processes, but is related to camp of thccin-sersitive DNA repair, conceivably involving the activity of topoisomrrasc I .

ANALYSES OF CHROMOSOME ABERRATIONS IN LYMPHOCYTES AND BONE MARROW CELLS INDUCED BY RADIATION OR BENZENE

The chromosome and chromatid type aberration can be induced by benzene and the dicentric and ring ones were not observed in vitro experiment but observed in vivo one. In vitro experiment a good linear regression can be given between benzene concentrations and total aberration cells while power regression for radiation dose.The chromosome aberrations induced by benzene combined with radiation in rabbit blood lymphocytes are higher than in bone marrow cells.

PRE-EXPOSURE OF MICE TO LOW DOSE OR LOW DOSE RATE IONIZING RADIATION REDUCES CHROMOSOME ABERRATIONS INDUCED BY SUBSEQUENT EXPOSURE TO HIGH DOSE OF RADIATION OR MITOMYCIN C

The phenomenon of cytogenetic adaptive and cross-adaptive response induced by low dose irradiation and chemical mutagen in mice is described. We found, firstly, that adaptation can be induced by acute low dose Xirradiation (0-100 mGy). Secondly, a cross-adaptation can occur between X-irradiation and mitomycin C (MMC). And finally, mice pre-exposed to chronic low dose rate￣(60)Co-Gamma irradiation (0-226.0 mGy/day) are less susceptible to chromosome aberration induced by subsequent acute higher Xirradiation. Therefore, our data suggest that radioadaptive response depends on dose, dose rate and time interval. Possible mechanisms are also discussed.

Acute toxicity and chromosomal aberration toxicity of insect wax and its policosanol

An acute toxicity test in Sprague Dawley(SD)rats and a chromosomal aberration toxicity test in Chinese hamster lung(CHL)fibroblasts were conducted to promote the application of insect wax and its policosanol.Results of oral acute toxicity test showed that the LD;values of insect wax and its policosanol were higher than 5000 mg/kg.The rats showed no obvious toxic symptoms and survived,suggesting that these substances were not toxic.The chromosomal aberration rates of the CHL cells incubated with insect wax for 3-6 h under metabolic activation conditions and for 3-6 h and 24 h under non-metabolic activation conditions were lower than 5%.No significant difference was found compared with the negative control group.Also,no significant difference in chromosomal aberration rates was found between each dose group of policosanol under metabolic or non-metabolic activation conditions and the negative control group.Therefore,the chromosomal aberration rates of CHL cells treated with insect wax and its policosanol were negative regardless of metabolic activation conditions.These results indicate the absence of acute toxicity and potential chromosomal aberration in the tested dose range of insect wax and its policosanol.

The Level and Distribution of Chromosomal Aberration of Tomato Seeds at Different Penetration Depths of Carbon Ions

The relationship between the penetration depth and the level and distribution of chromosomal aberration of the root tip cells were investigated by exposure of the superposed tomato seeds to 80 MeV/u carbon ions. The results showed that on the entrance of the beam the chromosomal aberration level was low. Damage such as breaks aud gaps were dominant. At the Bragg peak~ the chromosomal aberration level was high. The yields of dicentrics~ rings and disintegrated small chromosomes increased but the yields of breaks and gaps decreased. These results are consistent with the distribution of tile physical depth dose profile of carbon ions. It is effective to deposit the Bragg peak on the seeds to induce hereditary aberration in the mutation breeding with heavy ions.

Analysis of Unstable Chromosome Aberrations in Simulation of Partial Body Exposures with Gamma Rays

Accidentally or occupationally exposures to ionizing radiation generally involving the partial-body exposures and this may pose significant health hazards that are indicated by chromosome aberration （CA） induction. In this experiment, the quantification of the frequencies of CA was carried out based on cytogenetic analyses of peripheral blood samples obtained from 4 healthy volunteers as a result of simulation of partial-body exposures. The percentages of mixtures of blood samples irradiated in vitro with 2 Gy of gamma rays were 10, 25, 50, 75 and 100.0%. Lymphocytes were cultured and first-division metaphase cells were collected after culture times of 48 h and then harvested with standard procedures. The results showed that frequencies of unstable CA were depended on the percentage/portion of irradiated blood. All frequencies of observed CA was lower than that of calculated from 100% exposed blood, except in one case, indicating a phenomena of＂dilution＂ of unirradiated into irradiated lymphocytes though there could be a bystander effects taken place. The increasing in frequency for 25-100% portions was also comparable with other similar experiments. The quantification of CAs in lymphocytes is an important methodology of dose assessment for partial-body exposure to ionizing radiation, however, the scenario of exposure should be determined.

Effects of CO_2 laser on chromosomal aberration in the root tip cells of rice

Seeds of japonica rice cv.Zhenuo 2 with twodifferent physiological states(dry seeds withwater content 13% and wet seeds soaked in thewater for 36 h)were irradiated by COlaser infour different power-densities and durations re-spectively.The treatment irradiated with 200GY ofCo γ-rays was considered as control.The flesh root tips were cut and fixed inCarnoy’s fluid for cytological examination.

Studies on Ultra-dry Storage and Genetic Stability of Vegetable Seeds

[Objective] To study the long-term ultra-dry storage method and genetic stability of vegetable seeds.[Method] Seeds of Lycopersicum esculentum,Raphanus satuvus and Apium graveolen.were chosen as material.The changes of seed vigor,viability and genetic stability after ultra-storage were discussed by studying the seed potentiality,shoot length,germination percentage and the chromosome aberration rate of root tip cells.[Results] Maintaining the low moisture content,different vegetable species had different storage effects of the long-term storage seeds under normal temperature.The Lycopersicum esculentum and Raphanus satuvus seeds were more suitable to ultra-dry storage at normal temperature,and could keep good genetic stability,while the seeds of Apium graveolen had bad performance.[Conclusion] This study established the foundation of studying ultra-dry storage of vegetable seeds.

Copy number changes of target genes in chromosome 3q25.3-qter of esophageal squamous cell carcinoma: TP63is amplified in early carcinogenesis but down-regulated as disease progressed

AIM: By using comparative genomic hybridization, gain of 3q was found in 45-86% cases of esophageal squamous cell carcinoma (EC-SCC). Chromosome 3q25.3-qter is the minimal common region with several oncogenes found within this region. However, amplification patterns of these genes in EC-SCC have never been reported. The possible association of copy number changes of these genes with pathologic characteristics is still not clear. METHODS: Real-time quantitative PCR (Q-PCR) was performed to analyze the copy number changes of 13 candidate genes within this region in 60 primary tumors of EC-SCC, and possible association of copy number changes with pathologic characteristics was analyzed by statistics. Immunohistochemistry (IHC) study was also performed on another set of 111 primary tumors of EC-SCC to verify the association between TP63 expression change and lymph node metastasis status. RESULTS: The average copy numbers (±SE) per haploid genome of individual genes in 60 samples were (from centromere to telomere): SSR3: 4.19 (±0.69); CCNL1: 5.24 (±0.67); SMC4L1: 2.01 (±0.16); EVI1: 2.02 (±0.12); hTERC. 5.28 (±0.54); SKIL 2.71 (±0.14); EIF5A2. 1.95 (±0.12); ECT2: 9.18 (±1.68); PIK3CA: 8.13 (±1.17); EIF4G1: 1.07 (±0.05); 557: 3.07 (±0.25); TP63: 2.51 (±0.22); TFRC. 2.42 (±0.19). Four clusters of amplification were found: SSR3 and CCLN1 at 3q25.31; hTERC and SKIL at 3q26.2; ECT2 and PIK3CA at 3q26.31-q26.32; and 55T, TP63 and TFRC at 3q27.3-q29. Patients with lymph node metastasis had significantly lower copy number of TP63 in the primary tumor than those without lymph node metastasis. IHC study on tissue arrays also showed that patients with lymph node metastasis have significantly lower TP63 staining score in the primary tumor than those without lymph node metastasis. CONCLUSION: This study showed that different amplification patterns were seen among different genes within 3q25.3-qter in EC-SCC, and several novel candidate oncogenes (SSR3, SMC4L1, ECT2, and SST) were identified. TP63 is amplified in early stage of EC-SCC carcinogenesis but down-regulated in advanced stage of disease.

Mutagenic effects of chromium trioxide on root tip cells of Vicia faba

In this study on the mutagenic effects of different concentrations of chromium trioxide (CrO3) on Vicia faba root tip, micronucleus assay and chromosome aberration assay were used to determine the mitotic indexes, micronucleus rate and chromosome aberration rate of Vicia faba root tip cells. The results showed that the effects of CrO3 concentration on the mitotic indexes were complicated. CrO3 increases the micronucleus rate of Vicia faba root tip cells. It was found that within certain range of CrO3 concentration the micronucleus rate increased systematically with increased concentration of CrO3, but that the micronucleus rate decreased at higher level of CrO3 and that CrO3 also caused various types of chromosome aberration at a rate which increased systematically with increased concentration of CrO3. We concluded that CrO3 has significant mutagenic effect on Vicia faba root tip cells.

Genotoxic Effects of PAH Containing Sludge Extracts in Chinese Hamster Ovary Cell Cultures

Objective Many studies have been conducted in order to evaluate the genotoxicity of chemicals and waste materials, which utilized in vivo test protocols. The use of animals for routine toxicity testing is now questioned by a growing segment of society. Methods Keeping the above fact in mind, we have conducted in the present study the genotoxicity evaluation of oily sludge samples generated from a petroleum refinery and petrochemical industry and ETP sludge from petroleum refinery using DNA damage, chromosomal aberration, p53 protein induction and apoptosis in short term in vitro mammalian Chinese Hamster Ovary cell cultures. Results It is evident from the results that the oily sludge compounds derived from petroleum refinery and petrochemical industry could cause DNA damage, chromosomal aberration, p53 protein accumulation and apoptotic cell death on exposure to oily sludge extracts in the presence of metabolic activation system (S-9 mix), however, ETP sludge extract could not cause significant genotoxicity in comparison to oily sludge extract and negative control. Conclusion The effect may be attributed to polycyclic aromatic hydrocarbons present in the samples as evidenced from GC-MS.

Evaluation of Cytogenetic Effects of Isoproturon on the Root Meristem Cells of Allium sativum

Objective\ To validate the use of Allium sativum as a reliable test model for genotoxicity, isoproturon, a substituted phenylalkylurea herbicide, was evaluated on the root meristem cells by this plant system. Method\ Test concentrations were selected by determining EC50 and root tips were exposed to various concentrations for 6 or 24 hr. EC50 concentration was calculated to be 70.8 ppm for the root growth. In addition to root growth retardation exposure to isoproturon induced morpholoogical changes like discolouration and stiffness of roots. Results\ Exposure to various experimental concentrations of isoproturon (35-280 ppm), including EC50, significantly and dose-dependently inhibited the mitotic index and induced chromosome breaks/mitotic aberrations at 6 or 24 hr. Conclusion\ The frequency of aberrations was found to be decreased in a dose dependant manner at 24 or 48 hr post exposure, however in comparison of control cells the frequency of aberrations was considerably high which indicates genotoxicity potentials of isoproturon. Further, present study also suggests that Allium sativum is a sensitive, efficient, and reliable test system for measuring the genotoxicity potential of environmental chemicals.

ANEUPLOIDY INDUCTION BY THE WATER EXTRACT OF TRIPTERYGIUM HYPOGLAUCUM(LEVEL) HUTCH IN MOUSE BONE MARROW CELLS

The aneuploldy inducing activity of a Chinese medicinal herb, Tripterygium hypoglaucum (level) Hutch (THH) were investigated by means of thhree cytogenetic end points, namely C-mitotic (CM) effects, micronucleus (MN) and parallel chromosome structural aberration (CA) analyses in vivo. The experiments were performed on mouse bone marrow cells. THH showed similar gentoxic effects to colchicine (COL) in CM, MN and CA analyses; positive CM effects were observed accompanied with increases of mitotic index and frequencies of C-mltotic cells as well as decreased frequencies of anaphase in all of the THH-treated groups. The compound showed a positive MN response in bone marrow polychromatic erythrocytes but was negative in CA analyses. The preliminary results suggested that THH is an aneuploldy inducer in mouse bone marrow cells under present experiment conditions.

Genotoxicity of Pesticide Waste Contaminated Soil and Its Leachate

Objective Improper land disposal of hazardous waste can result in leaching of hazardous constituents which may contaminate ground and surface water leading to adverse impact on human health and environment consequences. The present study utilized mammalian cell culture for the genotoxicity assessment of waste and its leachate. Methods Genotoxic potential and chemical analysis of pesticide derived tarry waste contaminated soil extract and its leachate was assessed using in vitro human lymphocyte cultures and GC-MS. Results The investigation revealed that the soil extract could cause significant to highly significant genotoxicity in the form of DNA strand break at 25 mL (P<0.01), 50 mL, 100 mL and 200 mL (P<0.001) and chromosomal aberration at 25 mL (P<0.01) and 50 mL and 100 mL (P<0.001). The leachate could cause significant DNA strand break and chromosomal aberration only at 100 mL and 200 mL (P<0.01) dose levels. Conclusion The genotoxicity observed is attributed to carbaril and tetra methyl naphthyl carbamate, the major ingredients of the extracts, as revealed by GC-MS.

Cytogenetic Effects of Pulsing Electromagnetic Field on Domestic Pig Lymphocytes in Vitro

The effects of pulsing electromagnetic fields(PEMFs)on cells are very important subjects in the field of bioelectromagnetics.In this experiment,the cytogenetic effects of PEMF on domestic pig lymphocytes were tested in vitro.Pig lymphocytes in RPMI 1640 medium were exposed to PEMFs of 100 kHz and 200 kHz for 12,24 and 48 hours.Chromosomal aberrations(aneuploidy,breaks,gaps,et al)were significantly increased in exposed cultures,and of these aberrations,56%chromosomal or chromatid breaks and 42%gaps induced by PEMFs were the points of pig chromosomal fragile sites.The baseline frequency of sister chromatid exchange(SCE)increased after exposing lymphocytes continuously to PEMFs of 100 kHz and 200 kHz for 48 hours.These results suggested that the exposure to PEMFs might induce a type of DNA lesion and chromosomal aberrations.

Combination of Cytogenetic Analysis and Molecular Screening in Patients with de novo Acute Myeloid Leukemia

Nowadays the role of genetic findings in determining the diagnosis,therapy and prognosis of acute myeloid leukemia(AML) has become more valuable.To improve and validate the detection of clonal chromosomal aberrations in leukemia,we designed a combined application of karyotyping with multiplex reverse transcription-polymerase chain reaction(RT-PCR) and fluorescence in situ hybridization(FISH),and addressed the expression and distribution of fusion genes among the subtypes of Chinese adult patients with de novo AML.Multiplex RT-PCR assays were performed on 477 samples from newly diagnosed AML patients,and cytogenetic data were obtained from 373 of them by R or G banding techniques and those in some cases were confirmed by FISH.The PCR products in some suspected cases were tested by two-directional sequencing.The results showed that except unqualified samples,fusion genes were detected by multiplex RT-PCR in 211 of 474 patients(44.51%),including AML1-ETO,CBFβ-MYH11,PML-RARα,PLZF-RARα,NPM-RARα,MLL rearrangements,BCR-ABL,DEK-CAN,SET-CAN,TEL-PDGFR,TLS-ERG,AML1-MDS1(EVI-1).In 373 patients,who took both multiplex RT-PCR and karyotype analysis,the detection rate of chromosomal aberrations by using multiplex RT-PCR and karyotyping was 160/373(42.89%) and 179/373(47.98%) respectively,and the combination could optimize the detection rate of clonal genetic abnormalities to 216/373(57.90%).The PCR results from 11 cases 'normal' in karyotyping but abnormal in RT-PCR for MLL rearrangements were confirmed by two-directional sequencing.It is concluded that karyotype studies remain the cornerstone for genetic testing;conventional cytogenetics and molecular-based methods are complementary tests for the detection of clonal genetic aberrations in AML,especially for the cryptic or submicroscopic aberrations.Once a genetic marker has been identified by combined analysis,it could be used to monitor residual disease during/after chemotherapy,by quantitative RT-PCR and/or FISH.