Effect Of 5-bromodeoxyuridine Research Articles

Failure of 5-bromodeoxyuridine to affect the synthesis of α-amylase in rat parotid gland

The effect of 5-bromodeoxyuridine (BUdR) on the synthesis of α-amylase in rat parotid gland was studied. Taking into account that the half-life of the α-amylase mRNA, as determined in a separate experiment, is about 2.5 days, the following procedure was used. The gland was stimulated to proliferate by Isoproterenol (IPR) treatment of the animals. During the period of DNA replication up to 10–14 mol.% of the DNA thymine was substituted with 5-bromouracil (BU) by administration of BUdR adsorbed on activated charcoal. Two days later the gland was depleted of α-amylase by a second IPR treatment and the recovery of the enzyme was followed. The results showed that 10–14 mol.% substitution of BU for thymine did not affect the synthesis of α-amylase.

Some effects of 5-bromodeoxyuridine on polyoma-transformed mouse cells

The tumor-forming potential of polyoma-transformed mouse cells was found to be diminished but not eliminated after growth in the presence of 5-bromodeoxyuridine (BUdR). Several in vitro characteristics of the cells were also investigated and those which best correlated with the tumor studies were the ability to form colonies either in agar suspension or on monolayers of non-transformed cells. Transformed cells treated with BUdR grew well in culture when plated above a certain critical number of cells per unit area and poorly when plated at lower densities. Non-transformed cells exposed to BUdR for two or three passages were unable to divide.

Effect of 5-Bromodeoxyuridine on Pre-lmplantation Mouse Embryo Development

Effect of 5-Bromodeoxyuridine on Pre-lmplantation Mouse Embryo Development

Effect of 5-bromodeoxyuridine on the size distribution of DNAs isolated from sea urchin embryos.

5-BROMODEOXYURIDINE (BUdR), an analogue of thymidine, has been shown to selectively inhibit cell differentiation in various cell types and development of embryos, without affecting cell maintenance capabilities1. Gontcharoff and Mazia2 demonstrated that exposure of newly fertilised sea urchin eggs or early cleavage stage embryos to BUdR resulted in embryos which did not gastrulate or produce echinochrome pigment. From this laboratory evidence has been reported3 for a BUdR-induced modification of DNA replication in developing sea urchin embyros. Pulse-chase experiments in sea urchin gastrulae demonstrated that newly synthesised low molecular weight (8-15S) nuclear DNA labelled with 3H-BUdR was not readily chased by unlabelled BUdR into higher molecular weight DNA. In contrast, 8-15S DNA labelled with 3H-thymidine was converted into higher molecular weight DNA during a relatively short chase period with unlabelled thymidine. This data suggested the incorporation of BUdR into newly synthesised low molecular weight DNA molecules in embryos might prevent or slow the rate of normal linkage of such molecules into higher molecular weight DNA.

EFFECT OF 5-BROMODEOXYURIDINE ON GROWTH, ENCYSTMENT, AND EXCYSTMENT OF ACANTHAMOEBA CASTELLANII

EFFECT OF 5-BROMODEOXYURIDINE ON GROWTH, ENCYSTMENT, AND EXCYSTMENT OF <i>ACANTHAMOEBA CASTELLANII </i>

The effect of 5-bromodeoxyuridine on steroidogenesis in mouse adrenal tumor cells

Three days of growth in 5-bromodeoxyuridine inhibited both baseline and ACTH-stimulated steroidogenesis by 50% in mouse adrenal tumor cells in tissue culture. Steroidogenesis returned to control levels after these cells were reincubated in Eagle's minimum essential medium containing thymidine for 10 days. The inclusion of an equimolar concentration of thymidine in the culture medium prevented BrdU inhibition of steroidogenesis. Further studies demonstrated that incorporation of [ 3H]BrdU into nucleic acid was proportional to the reduction in steroidogenesis. Adrenal cells grown in the presence of hydroxyurea and [ 3H]BrdU did not incorporate [ 3H]BrdU into nucleic acid and released control levels of corticosteroids. These studies suggested that BrdU inhibited steroidogenesis by being incorporated into DNA in place of thymidine. Growth in BrdU inhibited the specialized function of adrenal tumor cells to a much greater extent than RNA and protein synthesis. Pregnenolone restored steroidogenesis in BrdU-treated cultures to control levels, suggesting that the step in the steroid biosynthetic pathway that is inhibited by BrdU is prior to pregnenolone formation.

Effect of 5-bromodeoxyuridine on chromatin transcription in confluent fibroblasts.

3T6 mouse fibroblasts were grown in 5-bromodeoxyuridine (BrdU) so that approximately 20% of the thymine residues in DNA were replaced by BrdU. BrdU replacement caused an alteration in the relative incorporation of labeled nucleotide precursors into RNA. The RNA synthesized by cells grown in BrdU has a lower proportion of adenine and a higher guanine complement. This was shown for (a) nascent RNA made in vivo by confluent monolayers of cells in culture; (b) RNA synthesized in vitro on a chromatin template with either homologous or heterologous RNA polymerase; and (c) RNA synthesized in vitro on a DNA template with a highly purified RNA polymerase. The product was completely digested by RNase. The relative decrease in the incorporation of adenine into RNA was reserved when BrdU-treated cells were allowed to proliferate in BrdU-free medium.

Modification of radiation-induced genetic damage in Drosophila melanogaster male germ cells with nucleic acid precursors : III. Effects on the premeiotic cells

Using a 2-day brood pattern, the effect of 5-bromodeoxyuridine (BUdR) or 5-bromodeoxycytidine (BCdR) pre-treatment on the radiation-induced yield of sex-linked recessive lethals and translocations was studied in the spermatocytes and late gonial cells (p.i. DNA synthesis cells) of D. melanogaster. The p.i. DNA synthesis cells were irradiated ( I.2 kR γ-radiation) in the pre-meiotic or post-meiotic stage. Irradiation of p.i. DNA synthesis cells in the pre-meiotic stage resulted in enhanced lethal frequency with BUdR (3.0%) and BCdR (2.9%) over the other pre-treatment conditions: saline (S), thymidine (TdR) and deoxycitydine (CdR) in the spermatocytes but not in the late gonial cells. The radiosensitizing property was evident with BCdR even when the p.i. DNA synthesis cells were irradiated in the post-meiotic stage; but not with BUdR pre-treatment. Probable reasons for the contradicting results reported in the literature were discussed.

The effect of 5-bromodeoxyuridine on the differentiation of chick embryo pigment cells

Cultures of ( a) dispersed presumptive melanoblasts from chick somites and ( b) organ cultures of retinal melanoblasts were grown in control medium and medium containing 5-bromodeoxyuridine (BUdR). The somite cell cultures at time zero had no evidence of melanogenic organelles; the eye cultures, from embryos of the same stage, stage 16–18, contained cells showing the initial stages of melanogenesis. In the presence of BUdR, presumptive trunk melanoblasts failed to pigment, while controls became heavily pigmented, whereas cells in the retinal pigment epithelium made melanin but in reduced amounts when compared with controls. These results suggest different methods of control over synthetic programs depending upon whether synthesis has, or has not, been initiated when the cells are exposed to BUdR.

Modification of radiation-induced genetic damage in Drosophila melanogaster Male germ cells with nucleic acid precursors III. Effects on the premeiotic cells

Using a 2-day brood pattern, the effect of 5-bromodeoxyuridine (BUdR) or 5-bromodeoxycytidine (BCdR) pre-treatment on the radiation-induced yield of sex-linked recessive lethals and translocations was studied in the spermatocytes and late gonial cells (p.i. DNA synthesis cells) of D. melanogaster . The p.i. DNA synthesis cells were irradiated ( I .2 kR γ-radiation) in the pre-meiotic or post-meiotic stage. Irradiation of p.i. DNA synthesis cells in the pre-meiotic stage resulted in enhanced lethal frequency with BUdR (3.0%) and BCdR (2.9%) over the other pre-treatment conditions: saline (S), thymidine (TdR) and deoxycitydine (CdR) in the spermatocytes but not in the late gonial cells. The radiosensitizing property was evident with BCdR even when the p.i. DNA synthesis cells were irradiated in the post-meiotic stage; but not with BUdR pre-treatment. Probable reasons for the contradicting results reported in the literature were discussed.

The effects of 5-bromodeoxyuridine (BrdU) on morphogenesis of the early chick embryo.

The effects of BrdU (3×10-4 M) on morphogenesis of the chick embryo explanted at the definitive streak stage and cultured for 24 hours were studied. Compared to controls treated embryos often showed (1) an open neural tube and (2) less numerous somites. Heart development was not significantly affected by BrdU. The damage caused by BrdU was not permanent, i.e., the embryos retained the ability to undergo fairly normal morphogenesis when, after 4-5 hours of BrdU treatment, they were subcultured on a medium with excess thymidine.

Human neuroblastoma cell culture: effect of 5-bromodeoxyuridine on morphological differentiation and levels of neural enzymes.

Summary5-Bromodeoxyuridine (5-BrdU) caused morphological “differentiation” of human neuroblastoma cells in culture, as shown by the formation of long neurites. This clone had a high level of choline acetyltransferase (ChA), however, 5-BrdU treatment allowed the expression of TH and COMT without any change in the cyclic AMP level. The addition of an equimolar of thymidine together with 5-BrdU did not reduce the BrdU-effects on morphological differentiation and TH level. The ChA activity decreased in BrdU-treated cells. This study indicates that the ChA and TH activities can be expressed in the same neuron.This paper was supported by DRG-1182 from Damon Runyon Memorial Fund for Cancer Research and by U.S. Public Health Service IP-CA-12247–01AL. We thank Ms. April Montgomery for technical help.

Adenosine 3′,5′-Monophosphate in Glial Tumor Cells: ALTERATIONS BY 5-BROMODEOXYURIDINE

Abstract The effect of 5-bromodeoxyuridine (BrdUrd) on the ability of the glial tumor cell line, C-6, to elevate its adenosine 3',5'-monophosphate (cAMP) content in response to norepinephrine has been examined. Increasing concentrations of BrdUrd were found to inhibit the norepinephrine response progressively, with a maximal inhibition of approximately 80%. No effect of BrdUrd was seen on basal cAMP levels. Analysis of individual enzymes demonstrated that BrdUrd increased the cAMP phosphodiesterase activities but had no effect on the adenylate cyclase or its sensitivity to norepinephrine. In addition, BrdUrd caused a change in the shape of the cells. The effect on shape also occurred in confluent cells, whereas the effect on the norepinephrine response required dividing cells, suggesting that incorporation of the BrdUrd into DNA was required. N6, O2'-Dibutyryl adenosine 3',5'-monophosphate also increased the cAMP phosphodiesterase activities of the cells, and caused a similar change in their morphology. Although BrdUrd had no effect on the adenylate cyclase, variations in the culture conditions were found to have a large effect on the enzyme's norepinephrine sensitivity. Following subculturing, there was little elevation of cAMP in response to norepinephrine until after the cells began to enter the stationary growth phase. Phosphodiesterase activity was found to remain constant throughout this period. The results suggest that cell-to-cell contact may regulate certain cellular functions.

Effect of 5-bromodeoxyuridine on the structure of differentiating choroid plexus cells in vitro

5-Bromodeoxyuridine caused structural alterations in a culture of choroid plexus cells of 13-day-old chick embryos, when added to the medium in a concentration of 0.122 mg/ml. The changes consisted of: increase of filamentous profiles, enlargement and segregation of the nucleoli, increase of vesicles in the cytoplasm, diminution of the tubules of the endoplasmic reticulum, and polymorphy of mitochondria. Under the same experimental conditions most of the connective tissue cells of 13-day-old chick embryos become necrotic.

Regulation of cartilage enzymes in cultured chondrocytes and the effect of 5-bromodeoxyuridine

When sternal cartilage from 15-day chick embryos is dissociated and cultured in normal media, the activity of the two cartilage enzymes, UDPG-epimerase and UDPG-dehydrogenase, falls to a level of about 25% of the original value within 3 days, but remains stable at this level thereafter. No change, on the other hand, occurs in the activity of the non-tissue-specific enzyme, phosphoglucose isomerase. The specific downward regulation of the cartilage enzymes seems to be attributable to the mitotic activity induced by the culture conditions. The addition of low concentrations of 5-bromodeoxyuridine to the culture medium, while having no effect on the isomerase or on cell division, causes a further drop in the activity of the cartilage enzymes from the 25% level to a value about 5%, but not further than this. In the case of the epimerase the latter low level is ubiquitously found in all tissues at all stages, indicating that only the cartilage-specific excess of this enzyme is susceptible of suppression by low concentrations of bromodeoxyuridine. The reduction of isomerase activity caused by higher concentrations of the analog is concomitant with a reduction of cell division and seems to be attributable to cell death. A scheme is presented summarizing the results and interpretations of the present paper and of a previous one.

DNA Damage and Repair Following Irradiation: The Effect of 5-Bromodeoxyuridine in Cultured Chinese Hamster Cells

Cell cultures, in exponential growth, were pretreated with noncytotoxic concentrations of the thymidine analogue 5-bromodeoxyuridine ( $1.0-3.0\times 10^{-7} M$ ) for ∼12 generations. Two forms of DNA damage and repair following radiation were evaluated by analysis of the sedimentation patterns of the DNA released during lysis on preformed alkaline sucrose density gradients. DNA released from cells grown in the presence of 5-bromodeoxyuridine was more susceptible to degradation by alkali (no irradiation). In irradiated cells, the production and repair of single-strand breaks in DNA were not significantly affected by 5-bromodeoxyuridine pretreatment. The low-dose radiation effect on DNA-i.e., the acceleration of the release in alkali of denatured DNA from a complex of associated cellular material by doses less than ∼700 rad-also may not have been affected by the incorporation of 5-bromodeoxyuridine. However, the repair of this low-dose radiation effect on the DNA complex was reduced in ...

Effect of 5-bromodeoxyuridine on hyaluronic acid synthesis of a clonal hybrid line of mouse and chinese hamster in culture.

AbstractA clonal hybrid line, B‐6, derived from the fusion of mouse and Chinese hamster cell lines, has an inheritable ability to synthesize and secrete hyaluronic acid (HA). In order to elucidate the control mechanism of expression of this differentiated function, we studied the effect of 5‐bromodeoxyuridine (BUDR), a thymidine (TDR) analog on the growth and HA synthesis of this hybrid line. The presence of BUDR at 0.5–1 μg/ml of medium had no effect on cell multiplication, but reduced HA production by approximately 25 to 40%. Treatment with 5 to 25 μg/ml of the analog resulted in about 15% decrease of the total cell growth, or 30% decrease of the viable cell growth, while mucopoly‐saccharide production was reduced by more than 70%. These results indicate that BUDR affects HA synthesis more strongly than it does cell growth.Inhibition was found to be detectable after two days of culture with BUDR. This effect was prevented by the simultaneous addition of TDR in a concentration more than twice that of the analog. Furthermore the inhibitory effect was reversible, since a gradual but complete recovery of HA synthesis occurred after removal of BUDR from the medium. In contrast, the growth rate of B‐6 cells was discovered to be more sensitive than HA synthesis to the inhibitory action of either excess TDR or another analog, 5‐fluorodeoxyuridine (FUDR).

Repair of X-ray damage to the DNA in Micrococcus radiodurans: the effect of 5-bromodeoxyuridine

Radiation damage to the DNA of Micrococcus radiodurans is eliminated or modified by processes which effect the rejoining of DNA strand breaks and degradation of the DNA. The enzyme system which is responsible for DNA degradation also appears to be implicated in the removal (excision) of damaged DNA constituents. Following anoxic X-irradiation the survival of the bacterium exceeds 95% at doses below 500 krads. Progressive replacement of the thymine in the cellular DNA by 5-bromouracil results in the progressive removal of the shoulder from the anoxic X-ray survival curve with little change in the slope. There does not appear to be a concomitant effect upon either the extent of DNA strand breakage or the properties of the system rejoining the breaks. However, the loss of the shoulder from the survival curve is paralleled by interference with DNA degradation. When the level of the 5-bromouracil incorporation is such that the shoulder has been eliminated from the survival curve the degradation-excision system is inoperative.

Effect of 5-bromodeoxyuridine on expression of cultured chondrocytes grown in vitro.

Exposure of cultured cartilage cells to 5-bromodeoxyuridine results in a rapid loss of the ability to synthesize the chondroitin 4-sulfate-protein complex characteristic of the extracellular matrix of differentiated cartilage. UDP-glucose dehydrogenase, UDP-N-acetyl hexosamine 4-epimerase and the enzyme system responsible for catalyzing 3'-phosphoadenosine-5'-phosphosulfate synthesis all progressively decline in activity, while several other cellular processes seem unaffected. However, there are marked morphological changes which may be associated with plasma membrane components or the synthesis of the protein-polysaccharide complex.

The effects of 5-bromodeoxyuridine on the differentiation of chondrocytes in vitro

BUdR at low levels (10 −7 M) can completely but reversibly inhibit the expression of differentiation by chondrocytes grown as single cell derived clones in a modification of Ham's medium F10. The effect of BUdR can be reversed by thymidine and uridine. Uridine itself stimulates differentiation, while the lack of thymidine and the drug aminopterin, which inhibits thymidine synthesis, interfere with expression of differentiation. Uridine reversal of the BUdR inhibition is blocked by aminopterin. Neither BUdR, aminopterin, or thymidine deprivation have significant effects on plating efficiency of the cells. BUdR-treated cells grow as apparent “fibroblasts” in the presence of the drug, but can revert to differentiated chondrocytes upon its removal. It is suggested that there exists a type of DNA synthesis necessary for cell differentiation and different from that involved in S phase cell DNA replication. Selective gene amplification such as is found in dipteran salivary chromosomes and during nucleolar production in oogenesis is suggested as possibly playing an important role in the maintenance and expression of the differentiated state in growing cells.