The Database for Analysis of Quantitative Characteristics of Chromosome Aberration Frequencies in the Culture of Human Peripheral Blood Lymphocytes

The data on spontaneous chromosome aberration rates in cultures of human peripheral blood lymphocytes obtained in the past 30 years have been collected to form a database. The database contains the results of analysis of more than 330 000 metaphases in lymphocytes from more than 1200 subjects. The frequency of aberrant metaphases in the control group has been estimated at 0.0213 ± 0.00085. No differences between sexes have been found with respect to either the total chromosome aberration rate or the rates of individual aberration types. The total chromosome aberration rate did not depend on age; however, it has been found that the number of fragments increased and the number of exchanges decreased with age. Smoking has been found to increase the frequency of chromosome aberrations in individuals with occupational hazards, but not in those who are not occupationally exposed to radiation or chemicals. Alcohol consumption increased the frequency of paired fragments, whereas the frequencies of other aberrations did not differ from the control values.     

Variation in Chromatin Structure of the rDNA Transcribed Region in Human Peripheral Blood Lymphocytes

The rDNA transcribed region (TR) was tested for its accessibility for RsaI recognizing 15 TR sites, DNase I, and photoinducible arylazide (N-(4-azido-2-hydroxybenzoyl)-N,N"-diaminoheptane acetate) in isolated nuclei, and for the arylazide in intact cells. Arylazide readily entered the cells and did not appreciably affect the chromatin structure. Its photolysis products efficiently modified DNA in accessible sites. Single-strand breaks made by DNase I were not transformed into two-strand ones in rDNA TR, suggesting the necessity of denaturing electrophoresis for such an analysis. About 70% of all rDNA copies proved poorly accessible for endonucleases and arylazide, the accessibility being higher in their 18S and 5.8S rRNA gene regions than in the regions of the external transcribed spacers (ETSs) and the 28S rRNA gene. Proteinase K disrupted this structure, and the corresponding copies were extracted from nuclei. This explained whyin situ hybridization occasionally fails to reveal rDNA in the nucleolar fibrillar center (FC) on electron microscopic preparations. In other rDNA copies, TR (excluding 5"-ETS) was accessible for nucleases and arylazide. These copies were not extracted from nuclei treated with proteinase K. Some of their RsaI sites were protected by tightly bound proteins. Seven such regions were identified in TR. Possible association of the molecular structure, nucleolar location, and functional state of rDNA is discussed.     

An Improved Culture System of Mouse Peripheral Blood Lymphocytes for Analysis of Radiation-Induced Chromosome Aberrations

There is an incentive to develop a culture system of mouse peripheral blood lymphocytes (PBLs) to serve as models for studying genotoxic effects in humans exposed to mutagens, including ionizing radiation. However, many past approaches have been laborious, complex and only partly reproducible. In the present study, we established an improved culture system of mouse PBLs by removing blood and/or plasma, which was found to inhibit in vitro mitotic stimulation or proceeding cell cycles of lymphocytes. We compared the reactions of isolated PBLs to mitogens between the classical method and the present improved one. Then, we applied this method to the cytogenetic analysis using chemically induced premature chromosome condensation (PCC) as well as the conventional analysis, and demonstrated that the frequency of excess fragments observed in PCC cells might be useful to quantify the radiation-induced damages on chromosomes.     

利用猪血清短期培养人体外周血淋巴细胞的研究

用猪血清代替小牛血清作人体外周血淋巴细胞短期培养,进行染色体分析及淋巴细胞转化的研究。在94例姐妹染色单体差别染色的培养中,培养液分别加入小牛血清与猎血清,细胞有丝分裂指数分别为3.91％和3.78％;21例常规法培养中,分裂指数分别为9.10％和9.21％;5例淋巴细胞转化试验,小牛血清和猪血清培养的转化率分别为69.52％和69.59％;16例阉割后公、母猪血清加入培养基中,细胞分裂指数(SCD法)分别为2.96％和3.14％。以上各项对照试验,均无显著性差异(P>0.05)本研究证实,猪血清可用于人体外周血淋巴细胞短期培养。     

低强度HeNe激光辐照人体外周血对淋巴细胞染色体影响的研究

本文通过低强度Ｈｅ－Ｎｅ激光以能量密度分别为１４．３１Ｊ／ｃｍ＾２（辐照５’）、２８．６２Ｊ／ｃｍ＾２（辐照１０）、５７．２４Ｊ／ｃｍ＾２（辐照２０）１１４．５２Ｊ／ｃｍ＾２（辐照４０）燠夫体外周血后,检测其淋染色畸变率（ＣＡ）,激光照射血样（能量密度由低到高）未照射血样ＣＡ分别平均为４．２９‰、３．９６‰、３．８１‰、３．５９０‰、４．１９‰、Ｘ＾检测无显著差异,说明低强度的Ｈｅ－Ｎｅ激光辐照人     

低强度He-Ne激光辐照人体外周血对淋巴细胞染色体影响的研究

本文通过低强度 Ｈｅ Ｎｅ 激光以能量密度分别为 １４３１ Ｊ／ｃｍ ２ （辐照 ５′）、２８６２ Ｊ／ｃｍ ２ （辐照 １０′）、５７２４ Ｊ／ｃｍ ２ （辐照 ２０′）１１４５２ Ｊ／ｃｍ ２ （辐照 ４０′）照射人体外周血后,检测其淋巴细胞染色体畸变率（ Ｃ Ａ）,激光照射血样（能量密度由低到高）及未照射血样 Ｃ Ａ 分别平均为 ４２９‰、３９６‰、３８１‰、３５９‰、４１９‰, Ｘ２ 检验无显著差异（ Ｐ＞ ００５）,说明低强度的 Ｈｅ Ｎｅ 激光辐照人体细胞对细胞染色体无致畸变效应。而且随着能量密度的增大,染色体的畸变率有降低的趋势,因此认为是低强度的 Ｈｅ Ｎｅ 激光促使细胞内分子的相互作用和能量转换,从而使染色体损伤的修复增强,其机理有待于进一步的研究。     

香蕉苗试管繁殖染色体数量畸变的研究

对在常规培养以及高６－ＢＡ、高腺嘌呤、高继代数、长继代时间等培养条件下,分化芽的染色体数目变化的规律以及一些苗期变异性状与染色体数量变异的关系进行了研究。主要结果有：（１）以上因素均能不同程度地促使染色体数量畸变率增高。其中,高腺嘌呤和高继代数加长继代时间２种处理效果最明显;（２）细胞畸变以非整倍体为主,可以在培养中逐代积累、增加。植株畸变以混倍体为主,非整倍体次之。植株畸变率明显低于细胞畸变率;（３）观察到一些苗的畸变的类型以及其对应的细胞学变化。结果显示,高继代代数的苗畸变率显著增高;在继代培养中,采取用核酸盐或高激动素以及片面地选用分化率高的芽丛作进一步扩大培养,以提高分化率的做法是危险的;根据苗期的形态特征,可将部分变异苗清除     

Variation of B Chromosome Associated with Tissue Culture in Wheat-rye Cross

In vitro variation of B chromosomes was studied by examining the callus cells derived from the immature embryos from a cross of Chinese Spring wheat （Triticum aestivum L.） and Fin 7416 rye （Secale cereale L.） carrying two B chromosomes. In 40-d-old callus cells, the numbers of B chromosomes ranged from one to four in 65.6% of the cells observed. The distribution of B chromosome numbers was associated with the ploidy levels of the normal chromosomes （A chromosomes）. The frequency of the cells with high numbers of B chromosomes （i.e., three or four B chromosomes） in the amphiploid cells with 56 A chromosomes was greater than those in the haploid cells with 28 A chromosomes. Although structural changes in the rye A chromosomes were observed, cytological observation and genomic in situ hybridization demonstrated that the rye B chromosomes were conserved in morphological appearance following tissue culture.     

In vitro Cytodifferentiation of Human Blood Lymphocytes

The proliferation of human blood lymphocytes after incubation with either antigen or nonspecific mitogens indicates a process of differentiation. This assumption is supported by several findings:
The small lymphocytes profoundly change their cytological structure, as transformed cells following stimulation by PHA, Con. A, tuberculin and MLC had the ultrastructural characteristics of immunoblasts. Blastoid transformed cells with a highly developed vacuolar apparatus were observed in those cultures stimulated by PHA and ALS.
This differentiation is paralleled by the development of some functions which characterize T-derived lymphocytes. It was demonstrated that the lymphocytes stimulated by these agents secrete mediators (i.e. a migration inhibition factor) and acquire killer properties. The steps of the cytotoxic process were studied using electron microscopy.     

Variation of B Chromosome Associated with Tissue Culture in Wheat-rye Cross

In vitro variation of B chromosomes was studied by examining the callus cells derived from the immature embryos from a cross of Chinese Spring wheat ( Triticum aestivum L.) and Fin 7416 rye ( Secale cereale L.) carrying two B chromosomes. In 40-d-old callus cells, the numbers of B chromosomes ranged from one to four in 65.6% of the cells observed. The distribution of B chromosome numbers was associated with the ploidy levels of the normal chromosomes (A chromosomes). The frequency of the cells with high numbers of B chromosomes (i.e., three or four B chromosomes) in the amphiploid cells with 56 A chromosomes was greater than those in the haploid cells with 28 A chromosomes. Although structural changes in the rye A chromosomes were observed, cytological observation and genomic in situ hybridization demonstrated that the rye B chromosomes were conserved in morphological appearance following tissue culture.     

Cryptic Variation in the Human Mutation Rate

The mutation rate is known to vary between adjacent sites within the human genome as a consequence of context, the most well-studied example being the influence of CpG dinucelotides. We investigated whether there is additional variation by testing whether there is an excess of sites at which both humans and chimpanzees have a single-nucleotide polymorphism (SNP). We found a highly significant excess of such sites, and we demonstrated that this excess is not due to neighbouring nucleotide effects, ancestral polymorphism, or natural selection. We therefore infer that there is cryptic variation in the mutation rate. However, although this variation in the mutation rate is not associated with the adjacent nucleotides, we show that there are highly nonrandom patterns of nucleotides that extend ~80 base pairs on either side of sites with coincident SNPs, suggesting that there are extensive and complex context effects. Finally, we estimate the level of variation needed to produce the excess of coincident SNPs and show that there is a similar, or higher, level of variation in the mutation rate associated with this cryptic process than there is associated with adjacent nucleotides, including the CpG effect. We conclude that there is substantial variation in the mutation that has, until now, been hidden from view.     

Variation in Short Tandem Repeats of Human Y Chromosome

Nine rare (biallelic) mutations and six short tandem repeats (STR) mapping to the nonrecombining portion of the Y chromosome were genotyped in 734 males from different geographical regions inhabited by the contemporary Armenian population. The analysis of molecular variance (AMOVA) showed that 48.9% of total STR genetic variation was explained by the differences between the haplogroups isolated based on biallelic polymorphism, whereas only 1.3% of genetic variation could be attributed to the differences between the geographic groups.     

Transport of the Immunosuppressant 15-Deoxyspergualin in Human Peripheral Blood Lymphocytes

15-deoxyspergualin (DSG) is a potent immunosuppressive compound currently in clinical trials. In this study, we have characterized the uptake and intracellular localization of DSG in human peripheral blood lymphocytes (PBL′s). DSG is transported into human PBL′s and reaches an estimated maximum concentration of approximately 500μM in 6 hours. The majority of the [³H]-DSG remains in the cytoplasm of cells and that which is associated with the nucleus is only loosely associated. DSG was transported by HeLa cells, as well, suggesting uptake is not specific for hematopoietic cells. Positively charged amino acids and polyamines, which are structurally similar to DSG, were unable to compete for DSG transport suggesting that DSG is transported into cells via a pathway distinct from amino acids or polyamines.     

The Geographic Distribution of Human Y Chromosome Variation

We examined variation on the nonrecombining portion of the human Y chromosome to investigate human evolution during the last 200,000 years. The Y-specific polymorphic sites included the Y Alu insertional polymorphism or ``YAP' element (DYS287), the poly(A) tail associated with the YAP element, three point mutations in close association with the YAP insertion site, an A-G polymorphic transition (DYS271), and a tetranucleotide microsatellite (DYS19). Global variation at the five bi-allelic sites (DYS271, DYS287, and the three point mutations) gave rise to five ``YAP haplotypes' in 60 populations from Africa, Europe, Asia, Australasia, and the New World (n = 1500). Combining the multi-allelic variation at the microsatellite loci (poly(A) tail and DYS19) with the YAP haplotypes resulted in a total of 27 ``combination haplotypes'. All five of the YAP haplotypes and 21 of the 27 combination haplotypes were found in African populations, which had greater haplotype diversity than did populations from other geographical locations. Only subsets of the five YAP haplotypes were found outside of Africa. Patterns of observed variation were compatible with a variety of hypotheses, including multiple human migrations and range expansions.     

Quantitative Characteristics of Chromosome Aberration Frequency in the Human Population of a Large Western Siberian Industrial Region

The results of a 15-year study of chromosome aberration frequency in cultured peripheral blood lymphocytes from subjects living in the Kuzbass industrial region are presented. The database for the analysis of the main parameters of chromosome aberrations contains data on 925 subjects, with the total number of cells examined being 92 900. It has been found that the total frequency of aberrant metaphases in the database is 3.73 ± 0.1%, whereas this frequency for the sample of subjects from industrial areas of this region (the basic control group) is significantly lower (2.86 ± 0.26%). It has been demonstrated that the sex and age of the subjects do not affect substantially the frequency of any type of chromosome aberrations. Tobacco smoking is associated with a slight increase in chromosome damage frequency; however, the difference between smokers and nonsmokers is insignificant even among subjects exposed to occupational hazards. The possible causes that have determined the increased basic and background chromosome aberration frequencies in the population of the Kemerovo oblast compared to these parameters for the European part of Russia and the CIS are discussed.     

Leucocyte Culture and Chromosome Preparations from Pig Blood

Blood was drawn into heparinized tubes from any large vein and allowed to settle 2-3 hr at 3-5 C. The cell sample consisting of 1 ml drawn from the buffy coat and 2 ml from the plasma was planted in the following medium: Medium 199 (Difco), 10 ml; penicillin G sodium, 1000 USP units; dihydrostreptomycin, 1 mg; and Bacto-PHA-M (Difco), 0.2 ml. Incubation, with twice daily shaking, was at 37 C for 68-70 hr; colchicine to give 4 μ ml was then added and incubation continued for 3-4 hr. The bulk of the medium was removed by centrifugation, the cells washed once in Hanks' salt solution, centrifuged, and all but 0.5 ml of the fluid decanted; 1.5 ml of distilled water at 37 C was added, the cell suspension incubated at 37 C 5-15 min, followed by centrifugation and fixation in methanolacetic acid 3:1 (3 changes) as usual. Spreads were made by applying 4-5 drops of cell suspension to ice-cold slides and burning off the fixative. Giemsa stain was used. The method has proved very satisfactory for determining chromosome numbers in the domestic pig. This number, as determined in 690 cells from Poland China and Duroc gilts and crosses of these breeds was 38 in 611 (88.6%) of the cells.     

Mechanism of the Cytotoxic Action of Gold Polyacrylate on Human Blood Lymphocytes

Biophysics - This work concerns the mechanism of the action of gold polyacrylate (aurumacryl) on human blood lymphocytes. The effects of the medication on cell viability and DNA structure and the...     

Leukocyte Culture and Chromosome Preparations from Adult Dog Blood

Plasma is obtained from dog blood after 3 hr settling in a syringe. Portions of the plasma (0.5-1.0 ml) are added to 4 ml of a medium consisting of 17 parts of BME Spinner, 3 parts of calf serum, 0.5 parts of glutamine, 0.5 parts of penicillin-streptomycin, and 0.1-1.0 parts of Scarlet Runner bean phytohemagglutinin. Colchicine, 0.1 ml of 10:1 stock solution, is added after 72 hr and incubation continued for 2 hr, then centrifuged 5 min at 700 rev/min. The supernatant is discarded, 3 ml of distilled water added, and the cell suspension centrifuged again. The supernatant is discarded and the fixative, consisting of 45% glacial acetic acid allowed to act for 0.5 hr. Acetic-orcein stains of smears were very satisfactory.     

Rate and Time of Dna Synthesis of Human Leukaemic Blasts In Bone Marrow and Peripheral Blood

Abstract. We have evaluated DNA synthesis rate (S rate) and time (T_s) and tritiated thymidine labelling index (LI) of peripheral blood (PB) and/or bone marrow (BM) leukaemic blasts (Bl) in nineteen cases of acute leukaemia (twelve non-lymphoblastic, AnLL, and seven lymphoblastic, ALL), in one case of non-Hodgkin's leukaemic lymphoma and in a case of plasma cell leukaemia.
The LI of PB-BI was significantly lower than that of BM-Bl (range 0.1-6.2% and 1.9-19.5%, respectively; P < 0.01). the S rate was higher for PB-Bl than for BM-Bl (range 3.5-11-3 and 2.5-9.5 mol × 10-¹⁸/min; P < 0.02) and the T^s of PB-Bl was shorter than that of BM-Bl (range 7.6-22.1 and 10.8-34.7 hr, respectively; P < 0.02). In eight cases where S rates of both BM-Bl and PB-Bl were available, a linear correlation ( r = 0.82; P < 0.01) was found between the two parameters. This suggests that the DNA synthetic rate is a property of the leukaemic cell line in individual patients and differs from case to case. It further indicates that the environmental influences on the DNA synthesis rate in BM or PB are always of the same order of magnitude. From the results of this study we speculate that the DNA synthesis rate of leukaemic blasts is slowed down in the BM by environmental factors such as cell density.