Heteroploid Cell Lines Research Articles

A Permanent Heteroploid Human Cell Line with Type B Glucose-6-phosphate Dehydrogenase

SummaryDet. 562 is a permanent heteroploid human cell line that is similar to HeLa and other heteroploid cell lines in morphology, growth, and virus susceptibility.It differs from similar permanent human cell lines in having G-6-PD type B isozyme. The implications of this finding are discussed. The importance of cell line monitoring is stressed.

Quinacrine fluorescent karyotypes of human diploid and heteroploid cell lines.

Quinacrine-fluorescence karyotypes were prepared on a series of human cell lines. WI-38 karyotypes were indistinguishable from those obtained from cultured XX blood leukocytes. WI-L2 lymphoblastoid cells were 46, XY and pseudodiploid, with a single chromosome 21 and an additional submetacentric marker whose short arm may have been derived from a No. 21. Three HeLa cell lines had several marker chromosomes in common. Three of these marker chromosomes were also present in a D98/AG line, which therefore must have been of HeLa cell origin. Another cell line of presumptive human origin was shown to be a contaminant, of mouse origin.

Isoenzyme stability in human heteroploid cell lines

Isoenzyme stability in human heteroploid cell lines

Effect of thyroxine on the cell generation cycle parameters of cultured human cells

A heteroploid cell line of human kidney cells was grown with or without the hormone 1-thyroxine. When the cells were in exponential growth the cell generation cycle characteristics were determined in control and hormone-treated populations. The life cycle of the cells was reduced from 27 to 20 h when hormone was present while the dead cell fraction remained the same. The shortening of the life cycle is due to a reduced time in the pre-DNA replication period (G1). The time to complete other stages of the life cycle appears to be virtually unchanged.

Heterokaryon formation between alkaline phosphatase variants in a human heteroploid cell line

The fusion between cells from a human line with high alkaline phosphatase levels (AP +) and from a clonal subline deficient in the same enzyme has been induced with Sendai virus. The proportion of cells with various numbers of nuclei has been determined and the polynucleated cells have been scored for numbers of AP + and AP − nuclei with the aid of the histochemical staining. The heterokaryons are readily recognizable as they show the enzyme in a region of their cytoplasm near to AP + nuclei. A higher proportion of homokaryons over heterokaryons within each class of multinucleated cells have been noticed. However the overall distribution of AP + and AP − nuclei in the multinucleated cells reflects the original proportion of the two types of mononucleated cells indicating the absence of any interaction between the two genomes in the immediate product of fusion.

VERO cells (Cercopithecus aethiops kidney)--growth characteristics and viral susceptibility for use in diagnostic virology. (Brief report).

An investigation of some of the characteristics of the VERO cell line (Cercopithecus aethiops kidney) is reported, in which the suitability of the cells for use in routine diagnostic virology was examined. VERO cells will: 1. grow to monolayers as rapidly as other heteroploid cell lines, but will maintain as usable monolayers in conventional maintenance medium for a significantly longer time; 2. grow a selection of ten representative viruses to a higher titre than will two other heteroploid lines or two cell strains; 3. support the growth of a very wide range of viruses with development in most instances of a characteristic cytopathic effect.

Glucose-6-phosphate dehydrogenase isoenzymes in human cell cultures determined by sucrose-agar gel and cellulose acetate zymograms.

SummaryCell lines and strains were studied with respect to possession of G-6-PD isoenzymes, using cellulose acetate strips and sucrose-agar gel electrophoretic media. The latter provided higher resolution of A and B bands. The A and B types of G-6-PD were found to occur in human, diploid cultured cell populations in accordance with racial origin of the donor; while certain other heteroploid cell lines were found to possess A type G-6-PD irrespective of donor race.

Lysosomes and virus induced chromosome breakage

The role of lysosomal enzymes as a possible mechanism in virus induced chromosome breakage was evaluated by correlating acid phosphatases and chromosome breaks in virus infected cell cultures and by testing the effect of cortisone on frequencies of virus induced chromosome damage. Virus-cell systems used were adenovirus-12 in human diploid cell lines and measles virus in heteroploid cell lines. The pattern of acid phosphatase stainings in adenovirus-12 treated diploid cells, as well as in measles virus treated heteroploid cells, was identical to the uninfected control cells. An increase of single chromosome breaks was seen in cells treated with adenovirus-12 and an increase of chromosome pulverization in cells treated with measles virus. On the contrary, in parallel experiments poliovirus (type 3) treated cells showed an increase in acid phosphatase staining indicating a rupture of lysosomes but no chromosome changes could be found in these cells. Cortisone in concentrations from 2.5 μg/ml to 250 μg/ml was found to have no effect on the frequencies of virus induced chromosome damage in either of the systems studied. Thus, no evidence was found to support the hypothesis that the action of lysosomal enzymes is the mechanism in virus induced chromosome breakage.

Growth and glycolysis in 2 cell lines derived from embryos of salmonid fish.

SummaryMeasurements of growth and glycolysis in 2 established heteroploid fibro blast-like cell lines derived from embryos of coho salmon and steelhead trout, growing at 23°, have been compared wit...

Fluctuation of alkaline phosphatase activity in synchronized heteroploid cell cultures: effects of prednisolone.

AbstractIn two heteroploid cell lines synchronized with thymidine double block, activity of alkaline phosphatase decreased during the 12 hour period preceding mitotic peak. A return to high values was observed during the next 12 hours of synchronous cycle. Prednisolone (11β, 17α, 21‐trihydroxy‐1,4‐pregnadiene‐3,20‐dione), when added to such cell cultures increased alkaline phosphatase activity in one of the cell lines (Henle embryonic intestine) but had the opposite effect on another line (HeLa‐S3) in which the enzyme activity was decreased. Neither effect could be demonstrated if the hormone was added at the end of S phase or if cells were arrested in metaphase by vinblastine sulfate.

Studies on the dynamics of heteroploid cell populations

The heteroploid cell lines may provide useful models for studying the dynamics of aneuploid cells populations which are frequently observed in tumours. Two important features to be analyzed in heteroploid cells populations are: chromosomal changes and selection processes. Two different methods are being used for investigating these problems: the first consists of analyzing the clonal distribution of cell characters which may be used as markers for their association with specific karyotypic changes. The second method consists of inducing chromosomal changes with various agents including 8-azaguanine and selecting for certain chromosomal variants. One example for each of these methods is given.

Enzyme activity during the growth and aging of human cells in vitro.

AbstractAcid phosphatase, alkaline phosphatase, and lactic dehydrogenase activities have been compared in normal human diploid cell strains and in SV40‐transformed heteroploid cell lines derived from them. A higher level of acid phosphatase activity was observed in diploid cultures derived from adult lung than in cultures derived from fetal lung of similar passage levels. The alkaline phosphatase activity of normal diploid fibroblasts was significantly higher than that of SV40‐transformed cell lines derived from them. Generally, the lactic dehydrogenase activities of all these cell cultures were similar.Human diploid cells in culture “age,” in the sense that their ability to proliferate decreases with time during serial subcultivation. Evaluation of the activities of these three enzymes during the “aging” process showed that, although alkaline phosphatase and lactic dehydrogenase activities were similar in “young” and “senescent” cells, acid phosphatase showed a small but significant increase in the senescent cells.

Enhancement of rhinovirus plaque formation in human heteroploid cell cultures by magnesium and calcium.

Fiala, Milan (University of Washington, Seattle), and George E. Kenny. Enhancement of rhinovirus plaque formation in human heteroploid cell cultures by magnesium and calcium. J. Bacteriol. 92:1710-1715. 1966.-A reproducible macroplaque assay for six M and three H strains of rhinoviruses has been developed in several human heteroploid cell lines. Plaques were produced only with suitable solidifying agents: purified agar (Ionagar, Agarose) or methylcellulose. Plaque development was greatly enhanced by increasing Mg(+2) to 30 to 40 mm. Diethylaminoethyl (DEAE) dextran also increased plaque sizes, and the effects of Mg(+2) and DEAE dextran were additive. In addition, Ca(+2) substituted for Mg(+2). The suitability of human heteroploid cell lines for rhinovirus plaque assay varied greatly, ranging from insensitivity through partial to complete sensitivity. This assay was six to seven times more sensitive than an end point tube assay. These results indicate that potentiation of plaque formation by Mg(+2) known for some enteroviruses can also be extended to the rhinovirus group of picornaviruses.

SPECIES DISTRIBUTION OF HUMAN TISSUE ISOANTIGENS. I. DETECTION OF HUMAN TISSUE ISOANTIGENS IN CHIMPANZEES

Human tissue isoantigens were detected on chimpanzee leukocytes and on a continuous heteroploid cell line of chimpanzee origin. The antisera used to detect these isoantigens were taken from multiparous women, multiparous chimpanzees, and chimpanzees immunized with human leukocytes and platelets. These sera were known to contain antibodies to human leukocyte isoantigens. The methods used to demonstrate the isoantigens were leukocyte agglutination and inhibition of leukocyte agglutination, as well as mixed agglutination and inhibition of this reaction.

Detection of Tissue Isoantigens on Primary and Serial Heteroploid Cell Lines of Human Origin by the Mixed Agglutination Technique

Summary The technique of mixed agglutination has been used to detect tissue isoantigens on human cells grown in tissue culture. Both primary human kidney cultures and heteroploid serial cell lines were shown to possess the isoantigens. The antisera used in these studies were from multiparous women and immunized chimpanzees. The sera were known to contain antibodies which reacted with human isoantigens by leukocyte agglutination. Absorption experiments showed the isoantigens detected on the serial heteroploid cultures were also present on human leukocytes and platelets.

Genetic studies of influenza viruses: II. Plaque formation by influenza viruses in a clone of a variant human heteroploid cell line

Plaque formation with representative strains of all major influenza A virus subtypes and with influenza B (Lee) virus has been obtained in a clone (clone 1-5C-4) of a human cell line of apparently unique susceptibility. The efficiency of plaque formation relative to egg infective dose (EIU) varies with the strain of virus, ranging from 2 to 70%. The clone (1-5C-4) has remained stable with respect to viral susceptibility for at least 36 transfers in mass culture during a period of 9 months. Certain strains of virus formed plaques of characteristic and distinctive type of potential value in genetic studies. Kinetic studies of the virus-cell interaction with NWS indicated that adsorption proceeded rapidly. However, about 40% of inoculated virus was unadsorbed. Adsorption not reversed with antiserum was a slower process and required 3–4 hours. Single-cycle infection with a high input multiplicity of virus required 20 hours for completion and resulted in a yield of 4 PFU/cell.

The limited in vitro lifetime of human diploid cell strains

The time at which human diploid cell strains can be expected to cease dividing in vitro (Phase III) is not a function of the number of subcultivations but rather of the number of potential cell doublings. Each clonable cell within the population is endowed with the same doubling potential (50 ±10). Cells of the same strain, but with different “doubling potentials”, were mixed. Phase III in such mixed populations occurs at that time when the “youngest” cell component is expected to reach Phase III. The “older” component has no effect on the time at which Phase III is expected to take place in the “younger” component. An ancillary conclusion that Phase III cannot be explained by the presence of a latent virus, mycoplasma or media composition is confirmed. Human diploid cell strains derived from adult lung have a significantly lower doubling potential in vitro than do fetal strains. The Phase III phenomenon may be related to senescence in vivo. The cellular theory of aging must be related to normal cells in vitro and not to heteroploid cell lines. The former have a finite period of multiplication; the latter are indefinitely cultivable. In vivo experiments also indicate that transplanted normal tissue has a finite lifetime. Chromosome anomalies occurring in Phase III may be related to such anomalies occurring in the cells of older animals, including man. The survival curves obtained with human diploid cell strains are comparable to “multiple-hit” or “multiple-target” curves obtained with other biological systems where an initial threshold dose is required before an exponential form of the curve is established. Whatever cell component(s) may be involved in the finite lifetime of human diploid cell strains, the ultimate accumulation of nondividing cells could be the result of accumulated damage to a single cellular target or to inactivation of many targets.

The serial cultivation of human diploid cell strains

The isolation and characterization of 25 strains of human diploid fibroblasts derived from fetuses are described. Routine tissue culture techniques were employed. Other than maintenance of the diploid karyotype, ten other criteria serve to distinguish these strains from heteroploid cell lines. These include retention of sex chromatin, histotypical differentiation, inadaptability to suspended culture, non-malignant characteristics in vivo, finite limit of cultivation, similar virus spectrum to primary tissue, similar cell morphology to primary tissue, increased acid production compared to cell lines, retention of Coxsackie A9 receptor substance, and ease with which strains can be developed. Survival of cell strains at − 70 °C with retention of all characteristics insures an almost unlimited supply of any strain regardless of the fact that they degenerate after about 50 subcultivations and one year in culture. A consideration of the cause of the eventual degeneration of these strains leads to the hypothesis that non-cumulative external factors are excluded and that the phenomenon is attributable to intrinsic factors which are expressed as senescence at the cellular level. With these characteristics and their extremely broad virus spectrum, the use of diploid human cell strains for human virus vaccine production is suggested. In view of these observations a number of terms used by cell culturists are redefined.