An improved system for exposure of cultured mammalian cells to gaseous compounds in the chromosomal aberration assay

A gas exposure system using rotating vessels was improved for exposure of cultured mammalian cells to gaseous compounds in the chromosomal aberration assay. This system was composed of 12 square culture vessels, a device for preparation of air containing test gas, and positive and negative control gases at target concentrations and for supplying these gases to the culture vessels, and a roller apparatus in an incubator. Chinese hamster lung cells (CHL/IU) were grown on one side of the inner surface of the square culture vessel in the MEM medium. Immediately prior to exposure, the medium was changed to the modified MEM. Air in the culture vessel was replaced with air containing test gas, positive or negative control gas. Then, the culture vessels were rotated at 1.0 rpm. The monolayered culture cells were exposed to test gas during about 3/4 rotation at upper positions and alternatively immersed into the culture medium during about 1/4 rotation at lower positions. This system allowed the chromosomal aberration assay simultaneously at least at three different concentrations of a test gas together with positive and negative control gases with and without metabolic activations, and duplicate culture at each exposure concentration. Seven gaseous compounds, 1,3-butadiene, chlorodifluoromethane, ethyl chloride, methyl bromide, methyl chloride, propyne, and vinyl chloride, none of which has been tested to date, were tested on CHL/IU for the chromosomal aberration assay using this gas exposure system. All the compounds except chlorodifluoromethane showed positive responses of the structural chromosomal aberrations, whereas polyploidy was not induced by any of these gases. This improved gas exposure system proved to be useful for detecting chromosomal aberrations of gaseous compounds.     

A unique human mutant B-lymphoblastoid cell line (ataxia telangiectasia) which exhibits increased siste-chromatid exchange retaining hypersensitivity to neocarzinostatin and bleomycin

Chromosome aberrations and sister-chromatid exchanges (SCEs) were examined in 4 ataxia telangiectasia (AT)-derived B-lymphoblastoid cell lines (B-LCLs) (AT-S, AT-SHI, AT-SHI B13A and AsHa) following treatments with neocarzinostatin (NCS) and bleomycin. All of these cell lines exhibited extremely high frequencies of chromosome aberrations with the NCS and bleomycin treatments. Among them, AsHa, a mutant B-LCL originating from an AT patient, showed high frequencies of SCEs under high bromodeoxyuridine (BrdU) concentrations retaining hypersensitivity to NCS and bleomycin with regard to chromosome aberrations. A clear BrdU dose-dependent increase in SCEs (9.85 SCEs/cell at 40 μg/ml, 36.65 SCEs/cell at 100 μg/ml on average) in this mutant was observed. When AsHa mutant cells were treated with NCS (0.02 μg/ml) and/or bleomycin (5.0 μg/ml) under 40 μg/ml BrdU (minimum BrdU concentration for sister-chromatid differential staining), SCE levels increased from 9.85 (baseline level) to 21.1 with NCS and 20.5 with bleomycin, in a dose-dependent manner. These observations indicate that AsHa is a unique AT-derived mutant cell clone with a high SCE character retaining the original hypersensitivity to bleomycin and NCS.     

Chromosomal aberrations in V79 hamster cells induced by hyperosmotic solutions of NaCl

In this study V79 hamster cells were treated with hypertonic NaCl solutions. A pulse treatment of 30 min made it possible to use hypertonic solutions of up to 1500 mM. Hypertonic treatment induced high frequencies of chromosomal aberrations and the aberration pattern led to the conclusion that hypertonicity acts similarly to S-phase-independent mutagens. We also tested the influence of several parameters on aberration induction and tried to standardize the experimental protocol. The mechanisms involved in aberration production after hypertornic treatment are unknown, we discuss a change in chromatin structure, inhibition of repair processes or protein damage responsible for chromosomal aberrations.     

The adaptation of a two-compartment cell renewal system to external demands

Summary The inner enamel epithelium (IEE) covers the labial tooth aspect as a one cell layer which, when cut sagittally, appears as a longitudinal cell column extending from the tooth origin toward the periphery. Following sudden tooth shortening, the IEE responds by an increased cell production which later declines below normal values. The perturbation affects all cell kinetic parameters; the progenitor compartment, which initially increases, diminishes in size toward end of the experiment. The cell cycle transition times, which initially decline, rise toward the end of the experiment. The mean normal daily cell production rate of 70 cell % (i.e. 70 cells are produced by 100 progenitors) increases to 111 cell % and then declines to a low of 51 cell %. The IEE response typifies the behavior of other cell renewal systems such as intestinal epithelium and epidermis.     

Chromosomal responses to ionizing radiation reminiscent of and adaptive response in cultured Chinese hamster cells

When Chinese hamster V79 cells were internally exposed to low level chronic β-rays from incorporated tritiated thymidine (³H-dThd), the showed an “adaptive” response to the induction of chromosomal damage by subsequent higher acute doses of γ-rays.

The yield of sister-chromatid exchanges (SCEs) in the ³H-dThd pretreated cells was less than the yield induced by γ-rays alone (protective effects), and the micronucleus frequency was less than the sum of the induced frequencies by ³H-dThd and γ-rays separately (below-additivity effects). No adaptation to the micronucleus induction by γ-rays was observed after the ³H-adapted cells had divided once and when 3-aminobenzamide (3AB) was given before the challenge doses. The cross-resistance study revealed that the ³H-adapted cells were resistant to SCE induction but not to the micronucleus inductions by the challenge doses of reactor radiations. The results suggest that the SCE adaptation and the micronucleus adaptation or clastogenic adaptation are probably caused by different, inducible adaptive repair pathways.     

玉米赤霉烯酮在小鼠体内的致突变性研究

用小鼠骨髓细胞微核试验和染色体畸变试验评价了赤霉病麦毒素之一玉米赤霉烯酮的体内致突变效应。玉米赤霉烯酮的试验剂量为0,0.1,1.0,10.0,100.0mg/kg体重,以环磷酰胺为阳性对照。实验结果表明,玉米赤霉烯酮各试验剂量组的微核发生率和染色体畸变率与阴性对照组相比均无显著性统计学差异,亦未发现有明显的性别差异。     

玉米赤霉烯酮在小鼠体内的致突变性研究*

用小鼠骨髓细胞微核试验和染色体畸变试验评价了赤霉病麦毒素之一玉米赤霉烯酮的体内致突变效应。玉米赤霉烯酮的试验剂量为0,0.1,1.0,10.0,100.0mg/kg体重,以环磷酰胺为阳性对照。实验结果表明,玉米赤霉烯酮各试验剂量组的微核发生率和染色体畸变率与阴性对照组相比均无显著性统计学差异,亦未发现有明显的性别差异。     

The serine 2481-autophosphorylated form of mammalian Target Of Rapamycin (mTOR) is localized to midzone and midbody in dividing cancer cells

Using a high-resolution, automated confocal high-content imaging system, we investigated the sub-cellular localization of the Serine 2481-autophosphorylated form of mTOR (PP-mTOR^Ser2481) during mitosis and cytokinesis in human cancer cells. PP-mTOR^Ser2481 exhibited a punctate nuclear distribution in interphase cancer cells, with the number of PP-mTOR^Ser2481 nuclear speckles positively relating with the proliferative capacity of cancer cells. PP-mTOR^Ser2481 expression dynamically rearranged within the cytoplasm in a close association near and between separating chromosomes during early stages of mitosis. Towards the end of anaphase and in telophase, PP-mTOR^Ser2481 drastically focused on the midzone and ultimately in the centre of the midbody at the presumptive cleavage furrow. In cells at cytokinesis, PP-mTOR^Ser2481 appeared as a doublet facing each other at the apical ends of two daughter cells. Three-dimensional analysis confirmed that PP-mTOR^Ser2481 positioned at a ring structure wrapped round by microtubule bundles to connect daughter cells. These results reveal for the first time that PP-mTOR^Ser2481 may be unexpectedly involved in the terminal stages of cytokinesis.     

Cytogenetical analysis on aneuploids obtained from pollenclones of rice (Oryza sativa L.)

Summary Rice aneuploids were obtained from 1,715 pollenclones with a mean frequency of 10.2% in anther culture (1983 to 1985). Among the aneuploids obtained, the frequency of primary trisomics ranged from 5.4% to 6,7%, tetrasomics from 1.1% to 1.7% monosomics from 0.9% to 1.3%, nullisomics from 0.5% to 1% and double trisomics from 0.5% to 0.7%. The chromosome complements of those aneuploids were identified by pachytene analysis on the absolute length of the extra chromosomes. Pollen clonal aneuploids showed a different range of variation in agronomic characters from dihaploids of the same origin but the phenotypic variations ressembled those found in aneuploids created by conventional breeding methods. The meiotic chromosome behavior of PMC revealed various chromosomal aberrations of aneuploids: loose pairing, trivalents, univalents, straggling chromosomes, bridges and laggards.     

墨江蜈蚣与少棘蜈蚣的比较研究──Ⅱ．药效和毒理

本文比较了墨江蜈蚣与少棘蜈蚣的药效学和毒理学作用,实验包括抗惊厥试验、对致病性真菌和细菌体外生长的影响、急性毒性试验和染色体畸变实验等。结果两种蜈蚣的３％醋酸提取液对所试的真菌有抑制作用但无杀真菌作用,而对细菌无抑制作用。两种蜈蚣的毒性很低,在给小鼠５０ｇ／ｋｇ的剂量下都无法测出ＬＤ５０。在２０５ｍｇ／ｋｇ条件下的致突变率与未给药的突变率接近,表明两者都无遗传毒性。     

Absence of chromosomal damage in human lymphocytes exposed to microwave radiation with hyperthermia

Specimens of human blood were exposed to 0, 4, 40, 100, and 200 Wkg-1 of 2.45 GHz microwave radiation for 20 minutes. The blood temperature was carefully controlled so that it rose from 37 to 40 degrees C. Cultured lymphocytes were examined for induced chromosomal damage but no effect in excess of background was observed.     

CO2激光对蚕豆诱变效应试验研究

７０年代以来,全国已有２０多个省市,近百个单位开展激光在农业上应用的研究,并对２０多种作物２００多个品种进行激光育种的研究。试验表明：激光在刺激作物生长,早熟增产以及诱发变异、改良品种等都有一定的效果,并培育出一些新品种应用于生产。然而,激光对作物的作用机制,特别是遗传机理方面的研究报道尚少。本试验目的是探讨ＣＯ２激光对蚕豆根尖细胞染色体畸变的影响及田间农艺性状的变异情况,分析细胞染色体畸变与农艺性状变异的相关性及其遗传传递情况,为作物激光诱变育种提供依据。本试验采用功率密度为３９４ｍＷ／ｃｍ２的ＣＯ２激光照射蚕豆萌动种子,观察其对蚕豆根尖细胞染色体畸变及幼苗生长的影响,探讨ＣＯ２激光照射后蚕豆植株农艺性状的变异及其遗传传递情况。试验结果表明：ＣＯ２激光不同时间照射蚕豆萌动种子,均能使根尖细胞产生染色体畸变,且畸变类型相似;同时,对Ｌ１幼苗生长有明显的抑制作用,Ｌ２植株高度、分枝数及结荚数等农艺性状均产生不同程度的变异。其中分枝数这一性状的变异能传递给Ｌ３,其余性状的变异则属生理损伤所致。     

4NQO对蚕豆诱变效应的研究

本试验利用４ＮＱＯ为诱变剂处理蚕豆种子,研究其诱变效应。结果表明;４ＮＱＯ对蚕豆Ｍ１的发芽率、苗高及染色体有较强的影响,可诱发蚕豆Ｍ１细胞染色体畸变。畸变类型以微核为主,是一种有效的诱变剂。     

氮离子束注入大麦种子的细胞生物学效应

本文研究了用３０ＫｅｖＮ＋离子束注入大麦干种子后其Ｍ１代的细胞生物学效应。研究结果表明,低剂量的Ｎ＋离子注入对大麦种子的萌发及Ｍ１胚根、胚芽细胞的有丝分裂有明显的促进作用。离子注入均能诱发胚根细胞和胚芽细胞的染色体畸变和核畸变,呈现微核、双核、小核、桥、断片和落后染色体等多种类型。并在２×１０１６Ｎ＋／ｃｍ２－８×１０１６Ｎ＋／ｃｍ２剂量范围内,注入剂量与畸变率之间有显著的正相关,但到１×１０１７Ｎ＋／ｃｍ２后畸变率却反而下降。研究结果还显示胚芽细胞较胚根细胞对氮离子束更为敏感。     

Function of the loop residue Thr792 in human DNA topoisomerase II alpha

We studied the mutation effect of one of the putative loop residues Thr792 in human DNA topoisomerase II alpha (TOP2 alpha). Thr792 mutants were expressed from high or low copy plasmids in a temperature sensitive yeast strain deficient in TOP2 (top2-1). When expressed from a high copy plasmid, mutants with small side chains complemented the yeast defect; however, from a low copy plasmid, only wild-type, Ser, and Cys substitution mutants complemented the yeast defect. Interestingly, at the permissive temperature other mutants (e.g., Val, Gly, and Glu substitutions) showed the dominant negative effect to the top2-1 allele, which was not observed by the control alpha 4-helix mutants. T792E mutant was 10-fold less active than wild-type and the T792P had no decatenation activity in vitro. These results suggest that Thr792 in human TOP2 alpha is involved in enzyme catalysis.     

γ－射线照射小鼠后F_1代中的染色体异常和显性致死突变研究

本文报道雌鼠及雌雄性小鼠分别受到０．１５－１．５６Ｇｙ~６０Ｃｏγ－射线照射后,在不同时间进行同笼,取其胚胎观察形态、大小等并制备每个胚胎的染色体标本。分析胚胎细胞染色体数目异常、结构畸变及显性致死效应。结果表明,染色体数目异常发生率与剂量呈直线关系。单纯雌鼠受到１．０７Ｇｙ照射后观察到平衡易位携带者的再现率为０．９９％。雌雄鼠均受到１．０７及１．５６Ｇｙ照射后其Ｆ_１子代中平衡易位携带者的再现率皆为０．３２％。雌雄鼠均受照射组中的显性致死率明显高于雌鼠受照射组,且剂量效应关系为直线相关     

Biological significance of chromosomal imbalance aberrations in gastrointestinal stromal tumors

Gastrointestinal stromal tumors (GISTs) are the most common mesenchymal tumors of the gastrointestinal tract. Current criteria for the diagnosis of malignant GISTs do not always reliably predict patient outcomes. In order to search for genetic markers with prognostic potential, chromosomal imbalance aberrations (CIAs) were analyzed in 28 subjects with GIST using comparative genomic hybridization and correlated with clinicopathological features. Except for a small rectal tumor, CIAs were identified in all GISTs, including 14 from the stomach, 11 from the small intestine, 1 from the esophagus, and 1 from the rectum. Losses were more common than gains. The median number of CIAs in high-risk GISTs was significantly higher than that in low-risk GISTs (5.60±2.59 vs. 3.38±2.55; p<0.05), especially for losses (4.60±1.84 vs. 2.63±2.13; p<0.01). Loss of 14q was the most common CIA in both low-risk and high-risk GISTs, and can be regarded as an early event of GIST development. Losses of 1p and 15q were also very common, often coexisting, and were slightly more frequent in high-risk GISTs than in low-risk GISTs. Other recurrent CIAs, including losses of 10q, 13q, 15q, 18q, and 22q and gains of 5p, 12q, 17q, and 20q were relatively less common in this series. Among these CIAs, losses of 13q, 10q (with minimal overlapping on q11–q22), and 22q were most likely the chromosomal loci potentially harboring the tumor suppressor gene(s) which may be related to early recurrence and/or metastasis during malignant transformation of GISTs.     

Tempol prevents genotoxicity induced by vorinostat: role of oxidative DNA damage

Vorinostat is a member of histone deacetylase inhibitors, which represents a new class of anticancer agents for the treatment of solid and hematological malignancies. Studies have shown that these drugs induce DNA damage in blood lymphocytes, which is proposed to be due to the generation of oxidative lesions. The increase in DNA damage is sometimes associated with risk of developing secondary cancer. Thus, finding a treatment that limits DNA damage caused by anticancer drugs would be beneficial. Tempol is a potent antioxidant that was shown to prevent DNA damage induced by radiation. In this study, we aimed to investigate the harmful effects of vorinostat on DNA damage, and the possible protective effects of tempol against this damage. For that, the spontaneous frequency of sister chromatid exchanges (SCEs), chromosomal aberrations (CAs), and 8-hydroxy-2-deoxy guanosine (8-OHdG) levels were measured in cultured human lymphocytes treated with vorinostat and/or tempol. The results showed that vorinostat significantly increases the frequency of SCEs, CAs and 8-OHdG levels in human lymphocytes as compared to control. These increases were normalized by the treatment of cells with tempol. In conclusion, vorinostat is genotoxic to lymphocytes, and this toxicity is reduced by tempol. Such results could set the stage for future studies investigating the possible usefulness of antioxidants co-treatment in preventing the genotoxicity of vorinostat when used as anticancer in human.