Identification ofTrypanosoma theileri as a contaminant in primary cultures of bovine retina

Summary T. theileri has been isolated from primary cultures of bovine retina and subcultered successfully for 2 passages in sub-confluent cultures. When cultures reached confluency no trypomastigotes or epimastigotes could be detected and attempts to recover trypanosomes from these cultures were unsuccessful. The presence of intracellular forms could not formally be excluded.We wish to thank Dr S. Dershko, Division of Meat Inspection, Health of Animals Branch, Department of Agriculture (Canada) and Intercontinental Packers, Ltd. for their assistance in obtaining the freshest possible specimens. The skillful technical assistance of Mrs J. Graham is also gratefully acknowledged. This investigation was supported by the M. R. C. of Canada.     

Use of aggregating cell cultures for toxicological studies

Relatively simple techniques are now available which allow the preparation of large quantities of highly reproducible aggregate cultures from fetal rat brain or liver cells, and to grow them in a chemically defined medium. Since these cultures exhibit extensive histotypic cellular reorganization and maturation, they offer unique possibilities for developmental studies. Therefore, the purpose of the present study was to investigate the usefulness of these cultures in developmental toxicology. Aggregating brain cell cultures were exposed at different developmental stages to model drugs (i.e., antimitotic, neurotoxic, and teratogenic agents) and assayed for their responsiveness by measuring a set of biochemical parameters (i.e., total protein and DNA content, cell type-specific enzyme activities) which permit a monitoring of cellular growth and maturation. It was found that each test compound elicited a distinct, dose-dependent response pattern, which may ultimately serve to screen and classify toxic drugs by using mechanistic criteria. In addition, it could be shown that aggregating liver cell cultures are capable of toxic drug activation, and that they can be used in co-culture with brain cell aggregates, providing a potential model for complementary toxicological and metabolic studies.     

Use of aggregating cell cultures for toxicological studies

Summary Relatively simple techniques are now available which allow the preparation of large quantities of highly reproducible aggregate cultures from fetal rat brain or liver cells, and to grow them in a chemically defined medium. Since these cultures exhibit extensive histotypic cellular reorganization and maturation, they offer unique possibilities for developmental studies. Therefore, the purpose of the present study was to investigate the usefulness of these cultures in developmental toxicology. Aggregating brain cell cultures were exposed at different developmental stages to model drugs (i.e., antimitotic, neurotoxic, and teratogenic agents) and assayed for their responsiveness by measuring a set of biochemical parameters (i.e., total protein and DNA content, cell type-specific enzyme activities) which permit a monitoring of cellular growth and maturation. It was found that each test compound elicited a distinct, dose-dependent response pattern, which may ultimately serve to screen and classify toxic drugs by using mechanistic criteria. In addition, it could be shown that aggregating liver cell cultures are capable of toxic drug activation, and that they can be used in co-culture with brain cell aggregates, providing a potential model for complementary toxicological and metabolic studies.     

Histamine production in mixed culture between allograft donor and recipient lymphocytes]

An increased production of histamine has been demonstrated during mixed cultures between allograft donor and recipient lymphocytes. This phenomenon could result from the action of a non-dialysable factor released by recipient cells in the presence of donor cells. This factor is able to increase histamine production from normal spleen cells. Little or no increase in histamine production is found during primary and mixed lymphocyte cultures (without previous allograft).     

Quantitative studies on in vitro transformation of hamster chondrocytes

Summary Hamster chondrocytes could be transformed in a quantitative assay system which used X-irradiated feeder layer cells. Morphological transformation occurred on addition of, 4NQO, but not in control cultures. Differentiation was classified into 3 types (good, poor and none); normal and transformed colonies contained similar proportions of the 3 types.This work was supported by a grant from the Ministry of Education, Science and Culture, Japan.We thank Miss M. Tanaka for her technical assistance.     

Preliminary observations of a bacteriophage infecting Xenorhabdus luminescens (Enterobacteriaceae)

A bacteriophage infective to Xenorhabdus luminescens, a bacterial symbiont of heterorhabditid nematodes, was recovered from insects that supported poor nematode development. Plaque tests showed the phage particles to be infective only to primary and not secondary colonies of X. luminescens. The phage was not infective to X. nematophilus primaries or secondaries. The bacteriophage particles ranged 80-90 nm in length, with the head ranging from 40 to 50 nm in diameter. Restriction analysis was performed on isolated bacteriophage DNA. This first report of a bacteriophage from Xenorhabdus species has practical implications since it could be detrimental to cultures of Heterorhabditis nematodes that are being produced throughout the world for the biological control of insects.     

Quantitative studies on in vitro transformation of hamster chondrocytes.

Hamster chondrocytes could be transformed in a quantitative assay system which used X-irradiated feeder layer cells. Morphological transformation occurred on addition of, 4NQO, but not in control cultures. Differentiation was classified into 3 types (good, poor and none); normal and transformed colonies contained similar proportions of the 3 types.     

Altered or increased transfer-RNA methylation in the course of Interferon action on cells in culture?

The induction of the antiviral state by Interferon might reflect the decrease of the rate of biosynthesis, the degradation or the alteration of one or several tRNAs. This could result in rate-limiting concentrations for codons common in viral RNA but rare in host mRNA. Altered methylation of tRNA could be the basis of such a phenomenon. However, we could not find an altered extent of methylation of total tRNA or an altered pattern of methylation, if mixed tRNAs were chromatographed on MAK- or BD-cellulose columns, despite a large range of conditions of pretreatment of chick embryo fibroblast cultures with interferon.     

Regulation of lymphokine production in peripheral blood mononuclear cell cultures

An increase in the production of macrophage migration inhibitory factor, chemotactic factor for neutrophils, and skin reactive factor, was observed in lymphocyte cultures if the cells were allowed to age in culture for 24 h. The increased lymphokine production was reduced by adding concanavalin A-stimulated and mitomycin C-treated suppressor cells. It is suggested that the lymphokine production could be regulated by suppressive mononuclear cells.     

Altered or increased transfer-RNA methylation in the course of Interferon action on cells in culture?

Summary The induction of the antiviral state by Interferon might reflect the decrease of the rate of biosynthesis, the degradation or the alteration of one or several tRNAs. This could result in rate-limiting concentrations for codons common in viral RNA but rare in host mRNA. Altered methylation of tRNA could be the basis of such a phenomenon. However, we could not find an altered extent of methylation of total tRNA or an altered pattern of methylation, if mixed tRNAs were chromatographed on MAK- or BD-cellulose columns, despite a large range of conditions of pretreatment of chick embryo fibroblast cultures with interferon.Work supported by the Swiss National Science Foundation, grants 3.1050 and 3.540.     

Variations in alpha-aminoisobutyric acid transport during in vitro differentiation of cultured myocardial cells from newborn rats]

The transport of alpha-aminoisobutyric acid has been studied as a function of age in heart cell cultures derived from new-born Rats. The specific rate of alpha-aminoisobutyric acid transport decreased slightly once confluency had been reached and then, from day 8 on, increased abruptly. This last phase may be related to the expression of specialized functions encountered in older cultures.     

A short-term culture method for chromosome preparation from somatic tissues of adult mussel (Mytilus edulis)

In order to improve the efficiency of mussel chromosome preparation, a tissue culture procedure has been developed. Mantle and foot explants were grown in tubes in media composed of Eagle's Basal Medium supplemented either with salts or seawater, enriched with egg yolk, adjusted to pH 7.50, and containing penicillin and streptomycin. After 4 days of incubation at 18°C, antibiotics were renewed and after 6–7 days, cultures were ready for harvesting and preparation of microscopical slides. The cultures were a source of actively dividing cells and consistent metaphase spreads were obtained. Evidence from BrdU incorporation suggested that cells could undergo several rounds of replication. The chromsome spreads were good enough for karyotyping and to successfully silver stain the nucleolar organizer regions.     

Preliminary observations of a bacteriophage infectingXenorhabdus luminescens (Enterobacteriaceae)

A bacteriophage infective toXenorhabdus luminescens, a bacterial symbiont of heterorhabditid nematodes, was recovered from insects that supported poor nematode development. Plaque tests showed the phage particles to be infective only to primary and not secondary colonies ofX. luminescens. The phage was not infective toX. nenatophilus primaries or secondaries. The bacteriophage particles ranged 80–90 nm in length, with the head ranging from 40 to 50 nm in diameter. Restriction analysis was performed on isolated bacteriophage DNA. This first report of a bacteriophage fromXenorhabdus species has pratical implications since it could be detrimental to cultures ofHeterorhabditis nematodes that are being produced throughout the world for the biological control of insects.     

Proteinzuwachs und Glukoseaufnahme isolierter Myokardzellen von Hühnchenembryo und Ratte in der Primärkultur

Summary Primary cultures of isolated myocardial cells of the chicken embryo (Ch) and of the new-born rat (R) present a characteristic behaviour of an increase of protein synthesis and glucose uptake: while in the Ch the increase of protein synthesis exceeds, in the R a high glucose uptake is shown. Both processes could be influenced by insulin.     

Development of in vitro toxicity tests with cultures of freshly isolated rat hepatocytes

Freshly isolated and cultured hepatocytes were analyzed by two-parameter flow cytometry. The combined analysis of DNA and cellular protein content allowed the contribution of ploidy classes and of subpopulations within a ploidy class to be defined. Analysis of hepatocytes during exposure to dimethylsulfoxide (DMSO), phenobarbital (PB), low oxygen tension (4% O2) or fetal calf serum (FCS), provided insight into the dynamic response of individual ploidy classes as a function of culture time. By analogy with the age-dependent ploidy shifts in vivo, hepatocyte-cultures shift towards adult animals during exposure to DMSO and towards young animals when cultured at low pO2 (4% O2). FCS and phenobarbital disturb this constitutive ploidy balance. FCS increased the 2 N cell population, where stem cells probably respond to the proliferative stimuli provided by growth factors in the serum. Phenobarbital affects the liver-specific 4 N hepatocytes, which agrees with effects seen in liver after exposure in vivo. It is suggested that drug-induced pathological alterations in ploidy in hepatocyte cultures could serve as indicators of compounds, such as liver tumor promoters, which interfere with cell differentiation in liver. The heterotypic cell-cell interaction of freshly isolated hepatocytes with isolated, in vitro cultured, rat liver epithelial cells in co-cultures proved to be a valuable concept in toxicity testing: aldrin epoxidase, an enzyme system involved in xenobiotic metabolism, was stabilized for more than two weeks. After exposure to the three chemicals, 2-acetylaminofluoren, procarbazine and cyproterone-acetate, a preferential toxicity for each compound and cell population was established. Thus heterotypic cell cultures can considerably increase the amount of information available from in vitro studies. The final concept, combining monitoring of cellular DNA (ploidy) and protein content in hepatocyte cultures during and after exposure to a given test compound at tissue oxygen tension with the heterotypic cell-cell interaction, would create a more in vivo-like culture system. This would enhance the predictability of hepatocyte cultures and contribute to a more widespread use of the test system and as a result help to reduce the number of whole-animal tests.     

Development of in vitro toxicity tests with cultures of freshly isolated rat hepatocytes

Summary Freshly isolated and cultured hepatocytes were analyzed by two-parameter flow cytometry. The combined analysis of DNA and cellular protein content allowed the contribution of ploidy classes and of subpopulations within a ploidy class to be defined. Analysis of hepatocytes during exposure to dimethylsulfoxide (DMSO), phenobarbital (PB), low oxygen tension (5% O₂) or fetal calf serum (FCS), provided insight into the dynamic response of individual ploidy classes as a function of culture time. By analogy with the age-dependent ploidy shifts in vivo, hepatocyte-cultures shift towards adult animals during exposure to DMSO and towards young animals when cultured at low pO₂ (4% O₂). FCS and phenobarbital disturb this constitutive ploidy balance. FCS increased the 2 N cell population, where stem cells probably respond to the proliferative stimuli provided by growth factors in the serum. Phenobarbital affects the liver-specific 4 N hepatocytes, which agrees with effects seen in liver after exposure in vivo. It is suggested that drug-induced pathological alterations in ploidy in hepatocyte cultures could serve as indicators of compounds, such as liver tumor promoters, which interfere with cell differentiation in liver. The heterotypic cell-cell interaction of freshly isolated hepatocytes with isolated, in vitro cultured, rat liver epithelial cells in co-cultures proved to be a valuable concept in toxicity testing: aldrin epoxidase, an enzyme system involved in xenobiotic metabolism, was stabilized for more than two weeks. After exposure to the three chemicals, 2-acetylaminofluoren, procarbazine and cyproterone-acetate, a preferential toxicity for each compound and cell population was established. Thus heterotypic cell cultures can considerably increase the amount of information available from in vitro studies.The final concept, combining monitoring of cellular DNA (ploidy) and protein content in hepatocyte cultures during and after exposure to a given test compound at tissue oxygen tension with the heterotypic cell-cell interaction, would create a more in vivo-like culture system. This would enhance the predictability of hepatocyte cultures and contribute to a more widespread use of the test system and as a result help to reduce the number of whole-animal tests.     

Cultures of simulations vs. cultures of calculations? The development of simulation practices in meteorology and astrophysics

While the distinction between theory and experiment is often used to discuss the place of simulation from a philosophical viewpoint, other distinctions are possible from a sociological perspective. Turkle (1995) distinguishes between cultures of calculation and cultures of simulation and relates these cultures to the distinction between modernity and postmodernity, respectively. What can we understand about contemporary simulation practices in science by looking at them from the point of view of these two computer cultures? What new questions does such an analysis raise for further studies? On the basis of two case studies, the present paper compares and discusses simulation activities in astrophysics and meteorology. It argues that simulation practices manifest aspects of both of these cultures simultaneously, but in different situations. By employing the dichotomies surface/depth, play/seriousness, and extreme/reasonable to characterize and operationalize cultures of calculation and cultures of simulation as sensitizing concepts, the analysis shows how simulation code work shifts from development to use, the importance of but also resistance towards too much visualizations, and how simulation modelers play with extreme values, yet also try to achieve reasonable results compared to observations.     

Metabolic studies of Hg-203 on Chlamydomonas reinhardi

Sterile cultures of Chlamydomonas reinhardi, WT+, were treated with Hg-203 at 25 degrees C to identify probably formed volatile mercury compounds. Experiments were performed with living and dead cells under aerobic or anaerobic conditions, respectively, and the mercury concentration was measured in the system algae/nutrient medium. We found a timerelated decrease of mercury concentration in the cell suspension and the cell-free nutrient medium due to a reduction of Hg++ to Hg0, probably caused by extracellular enzymes; monomethyl or dimethyl mercury could not be detected.     

Storage of irradiated human blood; a source of error in quantitative chromosome analysis

Human whole blood was irradiated with 2.5 Gy of 220 k Vp X-rays and stored before culture with 9.7 microM BrdU and 19.4 or 38.7 microM BrdU for 0, 24, 48 and 72 h. The frequency of dicentrics and ring chromosomes was determined in cells staining as first division (M1) metaphases with the fluorescence plus Giemsa technique. Storage had no influence on the observed aberration yields in 44 h cultures containing 9.7 microM BrdU. In 66 h cultures at 19.4 microM BrdU the observed yields after 2 and 3 days' storage were significantly lower as compared to cultures from fresh blood. No storage effect was revealed in 66 h cultures containing 38.7 microM BrdU. In cases where cytogenetic radiation dosimetry has to be carried out using blood samples which have been in transit for 2-3 days, the findings are of relevance for a correct determination of the chromosome damage in M1 cells.     

Regulation of lymphokine production in peripheral blood mononuclear cell cultures

Summary An increase in the production of macrophage migration inhibitory factor, chemotactic factor for neutrophils, and skin reactive factor, was observed in lymphocyte cultures if the cells were allowed to age in culture for 24 h. The increased lymphokine production was reduced by adding concanavalin A-stimulated and mitomycin C-treated suppressor cells. It is suggested that the lymphokine production could be regulated by suppressive mononuclear cells.This work was supported by the Scientific Research Council, Ministry of Health, Hungary (Code No. 421 0304011/S).