Comparison of silver staining of nucleolus organizer regions in human lymphocytes and fibroblasts

Summary Ag-staining of the nucleolus organizer region (NOR) was studied in the acrocentric chromosomes identified by Q-banding in repeated lymphocyte and skin fibroblast cultures from three different individuals. A similar pattern of Ag-stainability of NORs was found in the two tissues in each individual. Small differences concerning, in each case, only one of the acrocentric chromosomes were found between repeated lymphocyte cultures, as well as between lymphocyte and fibroblast cultures of the same individual without indication of any prevalence of one tissue type in a certain direction. The possibility that these differences are caused by different stages of NOR activation is discussed.     

Increased satellite association induced by 5′ Bromodeoxyuridine treatment of Phytohemaglutinin-stimulated blood lymphocytes

5'-Bromodeoxyuridine (BrdU) present in the course of late S and G2 phases of the cell cycle in PHA-stimulated human lymphocyte cultures causes the despiralization and elongation of some chromosome regions, including short arms of acrocentric chromosomes. BrdU present at a concentration of 250 microM during the last 10 h in lymphocyte cultures from 19 healthy subjects did not affect the number of silver-stained NORs, but raised significantly the number of satellite associations of acrocentric chromosomes. The mere substitution of thymine by BrdU in DNA strands as a reason for increased number of satellite associations seems a less plausible explanation than the modification of DNA-protein complexes of NOR regions, which could alter the degree of their spiralization and cause the increased tendency of acrocentric chromosomes to associate in the subsequent metaphase.     

Frequency of acrocentric chromosomal associations in children immunized with smallpox vaccine]

The frequency of associations of acrocentric chromosomes (AAC) diminished on the 7th day after vaccination in children primary vaccinated, primary revaccinated and secondary revaccinated against smallpox. This decrease reached its maximum by the 30th day and returned to its starting point after 6th months after vaccination. The degree of reduction of the frequency of AAC in every immunized children group correlated with the degree of increasing of antihemagglutinin titre. The relation of the number of group D chromosomes involved in AAC to the number of group G chromosomes varied in various individuals, these variations remaining after immunization. It was supposed that in PHA-stimulated lymphocyte cultures the degree of reduction of AAC frequency after vaccination against smallpox is a cytochemical marker of proliferation intensity of T-lymphocytes induced for immunopoiesis.     

Preferential association of nucleolar organizing human chromosomes as revealed by silver staining technique at mitosis

Summary The frequency of different types of satellite associations of nucleolar organizing human chromosomes (i.e. acrocentric chromosomes; 13, 14, 15, 21, and 22) is reported using 10 normal individuals by Ag-staining technique. The preferential involvement of acrocentric chromosomes in satellite association is suggested. Only acrocentric chromosomes with active NORs (i.e. Ag-stained) were found in association while unstained (inactive NORs) chromosomes were never seen in satellite association. In general as number of NORs expression increase, the frequency of association per cell was also increased. A possible mechanism and the clinical consequences of such an unusual phenophenon is described.     

Nondisjunction in human sperm: comparison of frequencies in acrocentric chromosomes.

Acrocentric chromosomes may be particularly predisposed to nondisjunction because of the frequency of trisomy for these chromosomes in human spontaneous abortions and liveborns. Studies of aneuploidy in human sperm have provided data on only a few acrocentric chromosomes, with evidence that chromosome 21 has a significantly increased frequency of disomy. To determine whether other acrocentric chromosomes have a higher frequency of nondisjunction or if chromosome 21 is anomalous, disomy frequencies for chromosomes 13 and 22 were studied by fluorescence in situ hybridization (FISH) analysis of 51,043 sperm nuclei from five normal men for whom the frequency of disomy for chromosomes 15 and 21 was known. The mean frequency of disomy for chromosome 13 (0.19%) did not differ significantly from that for other autosomes; however, the frequency of disomy 22 (1.21%) was significantly elevated (P < 0.001, Mantel-Haenszel chi(2) test). The G-group chromosomes (Nos. 21 and 22) also showed a significantly increased frequency of disomy (0. 75%) compared to acrocentric D-group chromosomes (viz., chromosomes 13 and 15; 0.15%) (P < 0.001, Mantel-Haenszel chi(2) test) and other autosomes (chromosomes 1, 2, 4, 9, 12, 13, 15, 16, 18, and 20; 0. 13%) studied in the same men (P < 0.001, Mantel-Haenszel chi(2) test).     

Frequency of Ag-stained nucleolus organizer regions in the acrocentric chromosomes of man

Summary The Ag-stainability of the nucleolus organizer region (NOR) was studied in the acrocentric chromosomes identified by Q-banding of cultured lymphocytes in 51 karyotypically normal persons (31 males and 20 females). A consistent pattern of Ag-positive NORs was found in each individual. Ninety percent of the individuals have a modal number of 8–10 Ag-positive NORs per cell. The frequency of Ag-positive NORs is similar in all five acrocentrics. A statistically nonsignificant lower frequency is found in chromosome 22. Ag-negative NORs on both homologues were found in four cases. The observed frequency distribution of individuals with homozygous NOR-positive, heterozygous, and homozygous negative acrocentric chromosomes was in accordance with the Hardy-Weinberg law in all five pairs of the acrocentric chromosomes as well as in total. No sex difference was observed in our material.A.-V. Mikelsaar is visiting exchange scientist of the Österreichische Bundesministerium für Wissenschaft und Forschung     

Sequential staining for G- and C-banding of chromosomes in the analysis of the morphology of the short arms of human acrocentric chromosomes]

Sequential staining for G- and C-banding of acrocentric chromosomes of 8 persons showed that the large heterochromatin region occurred more frequently in chromosome 15 than in chromosomes 13 and 14, and in chromosome 22 more frequently than in chromosome 21. There proved to be no correlation between the size of the heterochromatic region and the short arm of the acrocentric chromosomes. The frequency of occurrence of the satellites in the 8 persons was approximately the same for all the acricentric pairs. The C-banded satellite region of the homologous chromosomes is often heteromorphic.     

Frequency of acrocentric chromosome associations in human blood lymphocytes when using different methods for its assessment

The indeces of frequency of associations of acrocentric chromosomes in lymphocytes of human peripheral blood were compared with the help of different criteria of their estimation: by argentofile connections and by specific location of acrocentrics in metaphase plate. It was shown by the methods of variation statistics that the specificity of orientation of associative acrocentric chromosomes towards each other by short arms to the distance equal to the length of G-chromosome long arm is not accidental. Therefore, the method elaborated on the basis of the estimation criteria reflects participation of acrocentric chromosomes association in the formation of general nucleus in interphase more completely than the Ag-method, as preservation of Ag-material depends on the extent of its resorbtion in mitosis.     

Relationship of the frequency of acrocentric chromosome associations and the immune response in children primarily vaccinated against smallpox]

The frequency of acrocentric chromosomes associations in the lymphocytes of periferal blood decreased on the 7th and 30th day after vaccination. The rate of the decrease was in direct dependence on the intensity of the immunological response, that probably was due to shortening the length of the interphase of the lymphocytes induced for immunopoiesis. Frequency of the acrocentric chromosomes associations did not differ from the control level 6 months after vaccination.     

Variation in pattern and frequency of acrocentric association in normal and trisomy-21 individuals

Summary Chromosomally normal and trisomy-21 individuals were studied for the ability of their nucleolus-organising chromosomes to form satellite associations in G-banded lymphocyte metaphases. Two types of parameter, absolute association frequency and relative association frequency, were used. There was no significant difference between females and males or between Caucasoids and Mongoloids for either type of association parameter in the controls, nor was there significant correlation between age (17–40 years) and either type of parameter in the controls.The pattern of two chromosome associations is accounted for by two related models in both normal and trisomic individuals. These models imply that there is an extensive polymorphism for associating ability and that this ability may be zero in individual chromosomes. Homologous do not associate preferentially with each other. The absolute frequency of acrocentric association is lower in trisomy 21 individuals than disomic controls, but the relative involvement of chromosome 21 (after correction for the trisomic state) is higher than in the controls.     

N-Band polymorphism of human acrocentric chromosomes and its relevance to satellite association

Summary With the aid of Q- and N-banding techniques we investigated the relationship between the length of satellite stalks, the appearance of N-bands and the frequency of satellite association of individual acrocentric chromosomes in the cells of seven individuals, including one male with a satellited and small Y-chromosome. The appearance of N-bands seemed to be a constant and characteristic property of individual acrocentric chromosomes, independent of the status of concentration of the chromosomes at metaphase. The homolog with longer satellite stalks had larger N-bands and participated in satellite association at a higher frequency than the one with shorter stalks. It appeared that N-bands were present along the whole length of the satellite stalk, the size of which could possibly reflect the amount of rDNA present in the nucleolar organizers in human chromosomes.     

Inheritance of acrocentric association patterns.

The individual association frequencies of acrocentric chromosomes identified by fluorescent markers were analyzed in 17 individuals from 3 families. The frequency of association appears to be a characteristic property of an individual chromosome, since certain marker chromosomes showed an increase in frequency of association in each family member in which they appeared. Most of the marker chromosomes with increased frequency of association had a longer nucleolar constriction than their homologs.     

Genetic determination of NOR activity in human lymphocytes from twins

Summary Activity of nucleolar organizer regions (NORs) was studied in cultured blood lymphocytes from 20 monozygotic (MZ) and 20 dizygotic (DZ) twin pairs. The number of Agstained NORs, the degree of staining, and the frequency of acrocentric associations were used as criteria of the NOR activity, the acrocentric chromosomes being identified by G-banding. Analysis of intrapair concordance as well as of intrapair variance showed the number of Ag+NORs and the size of Ag-deposits to be highly heritable traits. Intrapair differences in acrocentric association frequency were not significantly higher in DZ compared with MZ twins.     

Satellite association in Down's syndrome

Summary The frequency of association of acrocentric chromosomes is examined in 20 Down's syndrome children, their parents, and 60 controls. Chromosome 21 enters into satellite associations most frequently, and chromosome 15 least. The parents of Down's syndrome children do not show any increased tendency for satellite association of chromosome 21 or indeed any other acrocentric.     

Attraction between centric heterochromatin of human chromosomes.

An analysis of the pattern of association of acrocentric chromosomes with nonacrocentric chromosomes in human lymphocyte metaphases was performed. This pattern in nonrandom with respect to chromosome length and intrachromosomal distribution. There is a general preference for the centric regions, most pronounced at the proximal segments of the long arms of chromosomes 1, 9, and 16, which is interpreted to reflect heterochromatin attraction during interphase. Comparison of the association patterns of homologous chromosome 1's differing with regard to the size of their heterochromatic regions corroborates this interpretation. The possible significance of heterochromatin attraction for the formation of spontaneous and induced chromosome anomalies is discused.     

Detection of nucleolus organizer regions in chromosomes of human,chimpanzee, gorilla,orangutan and gibbon

Nucleolus organizer regions were detected by the Ag-AS silver method in fixed metaphase chromosomes from human and primates. In the human, silver was deposited in the secondary constriction of a maximum of five pairs of acrocentric chromosomes: 13, 14, 15, 21 and 22. The chimpanzee also had five pairs of acrocentric chromosomes stained, corresponding to human numbers 13, 14, 18, 21 and 22. A gibbon had a single pair of chromosomes with a secondary constriction, which corresponded to the nucleolus organizer region. In each case the Ag-AS method detected the sites which have been shown by in situ hybridization to contain the ribosomal RNA genes. An orangutan had eight pairs of acrocentric chromosomes stained with Ag-AS, probably corresponding to human numbers 13, 14, 15, 18, 21 and 22, plus two others. Two gorillas had silver stain over two pairs of small acrocentric chromosomes and at the telomere of one chromosome 1. The larger gorilla acrocentric chromosomes had no silver stain although they all had secondary constrictions and entered into satellite associations.     

Karyotypes of squirrel monkeys (Saimiri sciureus) from different geographic regions

Three different karyotypes have been found so far among Saimiri originating from five different South American localities. All animals examined have the same diploid number (44) of chromosomes but the number of acrocentric and submetacentric chromosomes varies, presumably as a result of pericentric inversions. Saimiri originating from Iquitos, Peru, consistently have ten acrocentric chromosomes; animals originating from Leticia, Colombia, have 12 acrocentric chromosomes. Hybrids produced in our laboratory have the expected 11 acrocentrics and one unpaired submetacentric chromosome. Animals originating from Guyana have fourteen acrocentric chromosomes and the expected two fewer submetacentric chromosomes. Squirrel monkeys from Costa Rica, Panama, and Pucallpa, Peru, studied to this date conform to the Iquitos type with ten acrocentric chromosomes. These findings point to genetic differences which may result in variable responses to laboratory situations. The evolutionary factors involved in this rearrangement of chromosomes and possible influences on phenotypes are subjects of interest for future study. The importance of identifying the source of squirrel monkeys used in biomedical research is apparent if results from different laboratories are to be repeated or compared.     

Chromosomal banding pattern and idiogram of the cotton rat,Sigmodon arizonae (Rodentia,Muridae)

A procedure for obtaining G-bands on chromosomes of mammals is outlined. The procedure was utilized in an investigation of the idiogram and banding pattern of the mitotic chromosomes of the cotton rat, Sigmodon arizonae. The diploid number of this species is 22, and each pair of homologues is easily separated on the basis of size, centromeric position, and banding pattern. The autosomes are represented by four pairs of large submetacentric chromosomes, three pairs of medium to small submetacentric chromosomes, two pairs of large subtelocentric chromosomes and one pair of small acrocentric chromosomes. The X chromosome is acrocentric and averages from 5.42% to 5.46% of the haploid female complement. The Y chromosome is a minute acrocentric and easily separated from the smallest acrocentric autosome. The usefulnes of Sigmodon arizonae as a laboratory animal for cytogenetic studies is substantiated.     

The spermatogenesis of Dictyocaulus filaria (Nematoda, Trichostrongyloidea)

A cytological study was carried out, using male Dictyocaulus filaria, that revealed the diploid number of chromosomes was 2n = 11 and the sex determining mechanism was XO. The behaviour of the chromosomes in the different stages of meiosis was also investigated. Cross, open ring and rod bivalents were observed in diakinesis. The chromosomes appeared to be acrocentric since they acquired a radial disposition in Metaphase-II. The chiasma frequency was 1 and the nucleolus-organizing region was located at the ends of the chromosomes.     

Polymorphisms of Ag-stained nucleolar organizer regions in man

Summary Polymorphisms of the NORs as tested by Ag-staining of metaphase G-banded chromosomes were investigated in cultured blood lymphocytes of karyotypically normal individuals from the Moscow population.The study of cell-to-cell variability in the number of Ag-stained NORs carried out on 14 monozygotic twin pairs showed the phenomenon to have some features of real intercellular variation.In 40 unrelated individuals the individual acrocentric chromosomes were compared by the number of Ag-stained NORs, their degree of staining, and their participation in acrocentric association. Chromosome 21 was found to be significantly more active than four others by all the criteria, and chromosome 15 was less active compared with the others by the size of the Ag deposits and the frequency of participation in NOR associations. The frequency distribution of homozygotes and heterozygotes for Ag-stained NORs in the same group of 40 individuals was in accordance with the Hardy-Weinberg law.