RNA transport from isolated nuclei of cultured Djungarian hamster cells

The in vitro system of RNA transport containing isolated nuclei of Djungarian hamster cells transformed by SV-40 virus was studied. A functional test for cytoplasmic contaminations of the nuclei was proposed. The release of the newly synthesized RNA was found to be dependent on the duration of incubation, temperature and pH of the incubation medium as well as on the presence of spermine, spermidine, dithiothreitol, Mg2+, EDTA, exogenous RNA, nucleoside triphosphates and cytosol. The results obtained indicate that RNA release is an active process with activation energy of 12 kcal/mol. ATP, GTP, CTP and UTP have equal ability to serve as energy sources for the release of RNA. The nucleoside triphosphatase activity of the nuclei was the same in the presence of these four nucleoside triphosphates.     

Developmental abnormalities induced by 6-mercaptopurine in the hamster.

Pregnant hamsters were given varying doses of 6-mercaptopurine (6MP) at various times during gestation. The fetuses were examined for both gross and histological malformations which showed that the toxic and teratogenic effects of 6MP were dose and time dependent. The most severe gross malformations induced by 6MP were cleft palate, micrognathia and agnathia, microglossia, short limbs, and gut herniation. Grossly normal appearing fetuses, greated during late gestation, showed malformations at the tissue and cellular level. The effects of 6MP in hamster was compared with other species, and with other growth-supressive agents, and it was deduced that the teratogenicity of 6MP is species and tissue specific. Also, it was recommended that histological observations be made an integral part of the teratological safety analysis.     

Vertical development of the secondary palate in hamster embryos following exposure to 6-mercaptopurine

Cellular aspects of vertical development of the secondary palate were examined in control and 6-mercaptopurine (6MP)-treated hamster embryos. Cross-sectional area of the palatal shelf was measured and the numbers of both epithelial and mesenchymal cells counted. Also, in 6MP-treated palates the damaged mesenchymal cells, characterized by the presence of dense bodies, were counted. DNA synthesis in both control and treated fetuses was measured by 3H-thymidine incorporation. The results indicated that both the shelf area and cell numbers increased with age in control and 6MP-treated palates. However, in controls the mesenchymal cell density and DNA synthesis showed two peaks that were absent following 6MP treatment. Unlike controls, in treated embryos the damage to mesenchymal cells became increasingly pronounced between days 10:00 and 10:12 but subsided by day 11:00 of gestation. It is suggested that a major force in the development of the initial primordia and early vertical development of the palatal shelf may be provided by a spurt of DNA synthesis in the mesenchymal cells resulting in their increased number. After 6MP treatment, depression of DNA synthesis and consequent reduction in the mesenchymal cell number and density followed by cell damage lead to retardation in the vertical development of the palatal shelves.     

Induction of gene amplification in Djungarian hamster cells by various chemical carcinogens

The influence of 9 different carcinogens on gene amplification was studied in DM-15 Djungarian hamster cells. The effect was assessed by resistance to colchicine or methotrexate. It was found that tumour promotors (12-0-tetradecanoylphorbol-13-acetate (TPA), mezerein, tween-80) and some carcinogens possessing both initiating and promoting activity (20-methylcholanthrene, 7,12-dimethylbenz(a)antracene, aflatoxin B1) dramatically increased the number of colchicine and methotrexate-resistant cells. 4-0-methylTPA, a non-promoting analog of TPA, and alkylating carcinogens (ethylmethanesulphonate and nitrosomethylurea) did not induce gene amplification. It was suggested that the ability of carcinogens to induce gene amplification correlated with their ability to induce the second promotion stage.     

Spontaneous fibrosarcoma in a Djungarian hamster (phodopus sungorus)

A 1.5-y-old female Djungarian hamster (Phodopus sungorus) presented with a large subcutaneous mass surrounding the right shoulder. Radiography revealed dislocation of the right humeral articulation and osteolytic lesions of the right scapula. Histologically, the mass was composed of spindle to stellate cells arranged in fascicles interwoven with delicate collagen fibers, and neoplastic cells infiltrated the bone, skeletal muscle, and subcutaneous tissues. Neoplastic cells stained intensely positive for vimentin and negative for S100 protein, neurofilament, and desmin. A minority of neoplastic cells (10% to 20%) stained moderately for smooth muscle actin. The mass was diagnosed as a fibrosarcoma. Although fibrosarcomas are relatively common in dogs and cats, this is the first report of fibrosarcoma in a domestic Djungarian hamster.     

In vitro induction of numerical changes in the karyotype of normal and transformed Djungarian and Chinese hamster cells]

The comparative study of the frequency of colcemid-induced aneuploidy and polyploidy in cultured normal and transformed cells of Djungarian hamster is described. The occurrence of variants with changed chromosome number is much higher in populations of SV40-transformed cell line (4/21) than in normal embryonic cultures. In transformed lines of Djungarian and Chinese hamsters (4/21 and V-79) the frequency of cells with changed chromosome number was found to be dependent on the culture density: the percentage of polyploids was 4-5-fold higher when the number of seeded cells was 2-fold lower. The highest number (18-29%) of hypermodal cells was produced at drug concentrations of 0.02-0.025 mkg/ml. The percengate of polyploids under these conditions reached 10-20. At further increase of colcemid concentrations the proportion of polyploid cells increased. In Djungarian hamster embryonic cell cultures there were single cells with changed chromosome numbers at a concentration of the drug of 0.015-0.1 mkg/ml.     

Changes in the genotype and phenotype determining the resistance of Djungarian hamster somatic cells to adriablastin

The development of adriablastin resistance in Djungarian hamster DM-15 cells is accompanied by the appearance of small chromatin bodies (SCB) and long homogeneously staining regions (HSRs) in the chromosomes--the structures that contained amplified genes. The pattern of karyotypic alterations (the appearance of additional chromosome 4, and emergence of SCB, formation of the HSRs in one of three of chromosome 4, transposition of the HSRs from chromosome 4 to other chromosomes) during the development of adriablastin resistance is identical to that found in these cells before, namely during the development of colchicine resistance. Adriablastin- and colchicine-resistant cells have similar changes in plasma membrane permeability for 3H-colchicine, 3H-actinomycin D, 3H-puromycin, 3H-cytochalasin B, and 3H-vinblastine. Apparently, adriablastin resistance has the same mechanism as colchicine resistance, being connected with gene amplification and decreased plasma membrane permeability for these drugs.     

6-(2,4-Dinitrophenyl)-mercaptopurine, a stronger immunosuppressive drug than 6-mercaptopurine to primary humoral T-cell dependent immune response in mice]

The suppressive effect of 6-(2,4-Dinitrophenyl)-mercatopurine (DNP-MP) and 6-mercaptopurine (MP) was investigated on the early primary immune response of mice against the T-cell dependent antigens DNP49-bovine gamma globuline (BGG), sheep red blood cells (SRBC) or FITC8-BCG and the T-cell independent DNP22-Ficoll. The number of IgM antibody forming cells (AbFC) to the hapten determinants and to the SRBCs per 10(6) spleen cells was determined. DNP-MP reduced the number of AbFCs after the immunisation with the T cell dependent antigens always stronger than the MP, independently of the antigen type by which the mice had been immunised. The Anti-DNP22-Ficoll immune response was suppressed equally by both immunosuppressive drugs. DNP-MP is not a specific immunosuppressive drug for the anti-DNP-B-lymphocytes. Helper T-cells and macrophages are discussed as target cells for the stronger unspecific action of DNP-MP.     

Cloning and characteristics of DNA sequences amplified in Djungarian hamster cells with multidrug resistance

A number of DNA clones containing the amplified DNA sequences were isolated from the genomic library of multidrug-resistant (MDR) Djungarian hamster cells using the DNAC0t 10-250 hybridization probe. Five independent nonoverlapping clones were obtained that covered more than 100 kb of the amplified genomic region. These clones were used as hybridization probes in blot-hybridization with DNA from 7 independently derived MDR Djungarian hamster cell lines selected for the resistance to colchicine or actinomycin D. Some clones contained the DNA sequences amplified in all of the cell lines tested while the others contained the cell line specific amplified sequences. Hybridization in situ was used to localize the amplified DNA in metaphase chromosomes of a MDR cell line that contained about 140 copies of these sequences. The approximate size of an amplicon calculated on the basis of the obtained data is about 1-2 X 10(3) kb.     

Microbacterium oxydans, a symbiont of Djungarian hamster, displaying probiotic properties

A resident microorganism (strain Kho-17) was isolated from secretions of the specific glandular structures at the angles of mouth of Djungarian hamster (Phodopus campbelli). According to cultural, morphological, and physiological properties as well as to the phylogenetic analysis basing on the sequences of 16S rRNA gene and analysis of the cell wall the strain was assigned to the species Microbacterium oxydans. The bacterium isolated displayed probiotic properties when administered orally as a suspension of live cells for 20 days to Syrian hamster (Mesocricetus auratus), which manifested themselves in increased body weight and weights of several organs and stimulation of both cell-mediated and humoral immunities.     

Metabolism and mechanisms of action of 9-(tetrahydro-2-furyl)-6-mercaptopurine in Chinese hamster ovary cells

The mechanisms of action of 9-(tetrahydro-2-furyl)-6-mercaptopurine (THFMP) have been studied in Chinese hamster ovary (CHO) cells in tissue culture. THFMP is relatively unstable in physiological buffers, being facilely converted to 6-mercaptopurine (6-MP) even in the absence of cells. Consequently, THFMP undergoes metabolic conversions characteristic of 6-MP, namely formation of 6-thioIMP and incorporation into DNA as 6-thioguanine (6-TG) nucleotide. A number of purines are capable of preventing the toxicity of THFMP in wild-type cells in a manner similar to that of 6-MP. However, exogenous purines and pyrimidines did not prevent the toxicity of THFMP to cells deficient in the enzyme, hypoxanthine-guanine phosphoribosyltransferase (EC 2.4.2.8; HGPRTase). Cells lacking HGPRTase were 20–40-fold resistant to 6-TG and 6-MP but were only 2–4-fold resistant to THFMP. Furthermore, the time-course for killing CHO cells deficient in HGPRTase was different from that in wild-type cells containing the enzyme. There was no apparent effect of THFMP on the utilization of precursors for DNA, RNA or protein synthesis in the enzyme-deficient mutant cell line. The results suggest that THFMP is converted non-enzymatically to 6-MP and shares its mechanisms of action in wild-type cells containing HGPRTase, i.e., inhibition of de novo purine biosynthesis and incorporation into DNA as 6-TG nucleotide. However, the mechanism of action of THFMP in cells lacking HGPRTase is probably unique and is presently unknown.     

Seasonal adaptation of brown adipose tissue in the Djungarian hamster

Summary The composition and oxidative capacity of brown adipose tissue (BAT) were investigated in Djungarian hamsters kept under natural photoperiod, either indoors at neutralT _a (23°C) or under outdoor conditions. BAT comprises up to 5% of the body weight in summer/indoor hamster, with lipid representing 86% of the total tissue mass. Tissue mass and thermogenic capacity are inversely related during seasonal adaptation: 30% decrease of total DNA, accompanied by extensive lipid depletion, reduces the amount of BAT by almost 60% during acclimatization from summer/indoor to winter/outdoor conditions. Mitochondrial protein in BAT is increased by a factor of 2.6 concomitantly, and by a factor of 4 when related to body weight (body weight reduction 36%).Cytochrome oxidase activity in different brown fat deposits varies by up to 150% in summer/indoor hamsters; depending on the fat pad, the enzyme activity is increased 200%–700% during adaptation to winter/outdoor conditions.Natural photoperiod is decisive in determining the seasonal adaptation of DNA content in BAT and of body weight. Short photoperiod alone may lead to depletion of lipid content of BAT and thus decrease the tissue mass practically to the lowest seasonal level, even though both parameters may be also influenced byT _a. One third of the maximum adaptive increase of tissue mitochondria may be attributed to seasonal changes in photoperiod and up to two thirds toT _a. Photoperiod establishes a fixed fundament of slow-reacting functional adaptation of BAT, whereas the effect of decreasedT _a depends on the rate and duration of cold influence.Abbreviations BAT brown adipose tissue - NST nonshivering thermogenesis - T _a ambient temperature