EFFECT OF 5-BROMODEOXYURIDINE ON EXPRESSION OF CULTURED CHONDROCYTES GROWN
            <i>in vitro</i>

Holthausen, H.; Chacko, Samuel; Davidson, Eugene A.; Holtzer, H.

doi:10.1073/pnas.63.3.864

Cited by 43 publications

(9 citation statements)

References 14 publications

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“…The findings of Schulte-Holthausen et al (24) and Marzullo (25) of strikingly decreased activities of UDP-glucose dehydrogenase, UDP-N-acetylgalactosamine-4-epimerase, and UDP-glucose epimerase in BrdU-treated chondrocytes are at variance with our data for limb-bud cells. Experiments in this laboratory indicate that xylose overcomes the inhibition of chondroitin sulfate synthesis in BrdU-treated chondrocytes as well as in limb-bud cells.…”

contrasting

confidence: 57%

Control of Chondrogenesis in Limb-Bud Cell Cultures by Bromodeoxyuridine

Levitt

Doreman

1973

Proc. Natl. Acad. Sci. U.S.A.

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Initial exposure of cultured limb-bud cells (stage 23-24) to 5-bromo-2'-deoxyuridine (BrdU) irreversibly inhibits differentiation to cartilage under three different culture conditions. The inhibition of chondroitin sulfate synthesis is partially reversed by D-xylose in limb-bud cells after treatment with BrdU. The activities of four enzymes involved in chondroitin sulfate production were reduced in BrdU-treated cultures, but the magnitude of decrease was far less than the decrease in glycosaminoglycan synthesis. The slight increase in the turnover rate of sulfated glycosaminoglyeans in BrdU-treated mesenchyme was not sufficient to account for the marked decrease in chondroitin sulfate content. The results suggest that BrdU treatment interferes with normal synthesis of chondroitin sulfate core protein in cultured limb-bud cells, bitt does not greatly diminish enzyme activities or UDP-sugar levels necessary for production of polysaccbaride chains.Culture of several types of differentiated cells in the presence of the thymidine analogue 5-bromo-2'-deoxyuridine (BrdU) results in decreased synthesis of macromolecules characteristic of cell specialization (1-4). Differentiated cell expression is resumed upon removal of BrdU. In contrast to this reversible effect, BrdU irreversibly inhibits differentiation of certain precursor cell populations to specific phenotypes (5, 6). Irreversibility apparently depends upon treatment of cells at stages of development before "determination" has been stabilized. Differentiated chondrocytes are characterized by synthesis of large quantities of chondroitin sulfate proteoglycan. Production of chondroitin sulfate is sharply decreased in both differentiated chick chondrocytes growing in the presence of BrdU and cultures of chick limb-bud cells (stages 23-24) (7) initially exposed to BrdU then grown in its absence for 1A4 week&(5). Brett and Robinson (8) Glycosaminoglycan Synthesis and Turnover. On the ninth day of growth, cultures were labeled with 5 uCi of H235SO4,10 MACi of [3H]acetate, or 5 ,uCi of [I4C]acetate per ml of medium for 6 hr. For analysis of total glycosaminoglycan synthesis, cells and media were pooled and total glycosaminoglycans were isolated as described (9). For turnover measurements, fresh medium was added after a 6-hr pulse with 5 MCi of H2'5SO4 per ml on the ninth day of culture. Glycosaminoglycans were purified from cells plus fresh medium in 9-day, 11-day, and 13-day cultures, and radioactivity was determined by liquid scintillation counting.Enzyme Assays. Cells were scraped from culture dishes, washed twice with the appropriate buffer, and disrupted by 25 1-sec bursts with -a Branson Sonifier at 70 W. Lysis and homogenization were monitored with trypan blue stain. Xylosyltransferase (10) and N-acetylgalactosamine transferase assays (11) were performed on 10,000 X g supernatant

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confidence: 57%

Control of Chondrogenesis in Limb-Bud Cell Cultures by Bromodeoxyuridine

Levitt

Doreman

1973

Proc. Natl. Acad. Sci. U.S.A.

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“…In the presence of BrdU, presumptive myoblasts do not form myotubes or striated myofibrils (3,4). Chondrocytes cease production of cartilage enzymes (5,6), melanoma cells cease pigment production (7), and amnion cells cease mucopolysaccharide synthesis (8). This analogue also blocks the expression of the differentiated phenotype in cultures of pancreatic tissue (9,10), and prevents the induction of tyrosine aminotransferase in hepatoma cells (11).…”

mentioning

confidence: 99%

Lac Operator Analogues: Bromodeoxyuridine Substitution in the lac Operator Affects the Rate of Dissociation of the lac Repressor

Lin

Riggs

1972

Proc. Natl. Acad. Sci. U.S.A.

141

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As measured by a decreased rate of dissociation, lac repressor binds 10-times tighter to 5-bromodeoxyuridine-substituted lac operator than it does to normal lac operator. This result is obtained both in the absence and in the presence of isopropylthiogalactoside, an inducing ligand. These data are significant with regard to the mechanism of sequence-specific protein-DNA interaction, and also suggest a possible explanation for the effects of bromodeoxyuridine on the expression of differentiated functions in eukaryotic cells. Recently, the effect of BrdU on eukaryotic cells has received a great deal of attention, and. our results may be relevant to this work. BrdU selectively blocks the expression of differentiated functions. In the presence of BrdU, presumptive myoblasts do not form myotubes or striated myofibrils (3, 4). Chondrocytes cease production of cartilage enzymes (5, 6), melanoma cells cease pigment production (7), and amnion cells cease mucopolysaccharide synthesis (8). This analogue also blocks the expression of the differentiated phenotype in cultures of pancreatic tissue (9, 10), and prevents the induction of tyrosine aminotransferase in hepatoma cells (11). As pointed out by Holtzer and Abbott (12), BrdU appears to suppress selectively the synthesis of "luxury" molecules without grossly depressing the synthesis of "essential" molecules. Within the last year, several groups have also reported (13)(14)(15)(16)(17)(18)

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“…These observations, that the suppression of melanogenesis and the normalization of transformed phenotypes are coupled by BrdU treatment, are consistent with our results showing that cells of tyrosinase-negative cell lines always have diminished transformed phenotypes. In other primary-cultured cells (5,11), similar BrdU effects, the suppression of specific differentiated functions and the alteration of such cellular phenotypes as cell flattenisg with an increase in adhesiveness to substratum, have been reported to be induced at the same time. This suggests a close relationship between differentiative functions and cellular (or transformed) phenotypes.…”

Section: Discussionmentioning

confidence: 77%

A Correlation between Melanogenic Functions and Transformed Phenotypes in Cultured B16 Mouse Melanoma Cells

Sato

Kinoshita

Takeuchi

1980

Cell Struct. Funct.

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Cell differentiation not only results in the production of specific molecules for differentiated functions, but is accompanied by concomitant alterations in other cellular phenotypes including cell morphology and cell behavior (4,6,7,12). This also seems to be the case for the suppression of differentiated functions. For instance, when melanoma cells are treated with 5-bromodeoxyuridine (BrdU), their differentiated marker, melanin synthesis, is suppressed, and such transformed phenotypes as tumorigenicity, plasminogen activator production and cell-to-substratum adhesiveness are concurrently modified (2, 15). How these alterations in the various cellular phenotypes cause the expression or suppression of differentiated functions is not clear.In the present study, we isolated tyrosinase-negative variants which appeared spontaneously in a cultured melanotic B16 mouse melanoma cell strain and compared them with tyrosinase-positive cell lines obtained from the same cell strain. The variants were characterized by their melanogenic ability. We used other cellular phenotypes including cell morphology, anchorage dependency and cell-to-substratum adhesiveness to examine whether any significant relationship between the loss or expression of differentiated functions and the alteration of transformed phenotypes Abbreviations:BrdU, 5-bromodeoxyuridine; DOC, sodium desoxycholate; OD, optical density; DOPA, 3,4-dihydroxyphenylalanine.199

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EFFECT OF 5-BROMODEOXYURIDINE ON EXPRESSION OF CULTURED CHONDROCYTES GROWN in vitro

Cited by 43 publications

References 14 publications

Control of Chondrogenesis in Limb-Bud Cell Cultures by Bromodeoxyuridine

Control of Chondrogenesis in Limb-Bud Cell Cultures by Bromodeoxyuridine

Lac Operator Analogues: Bromodeoxyuridine Substitution in the lac Operator Affects the Rate of Dissociation of the lac Repressor

A Correlation between Melanogenic Functions and Transformed Phenotypes in Cultured B16 Mouse Melanoma Cells

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