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At least 469 records · Page 26Linked to original sources
PubMed2004

The role of late/slow replication of the FRA16D in common fragile site induction.

The FRA16D, at 16q23, spans the WWOX gene and is one of the most highly expressed common fragile sites observed when DNA replication is perturbed by aphidicolin. Several lines of evidence suggest that fragile sites are regions of DNA that are unusually sensitive to interference during replication. We have determined that the FRA16D alleles replicate in a synchronous fashion and that replication of these sequences occurs primarily in late S phase extending into G2 phase. Exposure to aphidicolin, an inhibitor of DNA polymerase alpha, results in a modest increase in cells with replication of FRA16D sequences in early S phase. This may represent initiation of replication in early S phase coupled with slow replication progression, or, alternatively, these cells may have passed through mitosis, entered the G1-S phase of the next cell cycle, and initiated replication/repair. Our results support a model in which common fragile sites are sequences that may initiate replication in early-mid S phase but are slow to complete replication, and the chromosomal breaks and gaps observed in metaphase cells result from unreplicated DNA.

Acid Anhydride Hydrolases↗
PubMed1992

A histologic and flow cytometric study of Kaposi's sarcoma.

One hundred forty-three biopsies of Kaposi's sarcoma (KS) from 96 patients were assessed histopathologically, and mitoses were counted. Ninety-seven samples from 66 patients were analyzed by flow cytometry. Six of 97 (5.8%) KS lesions were DNA aneuploid with a clustering around a DNA index of 1.5 (range, 1.4 to 1.6). The median percentage of S-phase plus G2-phase cells (%S + G2) was 16.7%. Increasing mitotic counts and %S + G2 were seen with progression of the phase and pattern of disease. Nodular KS and spindle cell predominant KS had the highest mitotic counts and %S + G2, with nodular KS larger than 4 mm having a higher mitotic count than those smaller than 4 mm. These findings suggest a low level of DNA aneuploidy in KS and important changes in the level of cell proliferation with the phase and pattern of the disease. However, flow cytometry does not solve the dilemma of whether KS is a hyperplastic or neoplastic process.

Aged↗
PubMed2002

[The influence of epidermal growth factor and insulin on proliferation and DNA synthesis in ciliates Tetrahymena pyriformis].

The influence of the epidermal growth factor (EGE) (10(-8) M), insulin (10(-6) M) and EGF (10(-8) M) in combination with insulin (10(-6) M) on proliferation and DNA synthesis in the nuclei of ciliates Tetrahymena pyriformis GL was studied. Insulin and EGF, known to stimulate growth of many types of mammalian cells revealed a mitogen influence on the unicellular eukaryotes. This effect involves stimulation of DNA synthesis, rising synchronization of cell division (upon the influence of EGF), and increase in cell number during the exponential growth. The mitogen effect may be evoked by cell progression in G1-phase under the action of growth factors and, consequently by earlier entry of cells into S-phase of the first cell cycle. Insulin repressed division of cells that entered into the generative cycle. These cells were delayed in late S-phase and G2-phase of the cycle. Part of these cells perished, while other cells could successively overcome the cell block to start their division by the 4th hours of cultivation. A collateral cytotoxic effect of insulin was found, being most prominent in early periods of Tetrahymena cultivation.

Animals↗
PubMedDate not supplied

Alcohol inhibits cell mitosis in G2-M phase in cell cycle in a human lymphocytes in vitro study.

Alcohol abuse is associated with the loss of immunocompetence, which leads to decreasing resistance to infections. No single mechanism can be accountable for the detrimental effects of alcohol on the body's defense mechanism. We present data demonstrating that, in cultured lymphocytes, 10-40 mM alcohol in the media caused 18-90% decrease in cell mitosis (p < 0.001). There was a linear decrease in cell mitosis upto 40 mM alcohol; at 100 mM cell mitosis virtually ceased. This study aimed to determine in which phase of the cell cycle did alcohol mediate its effects. The results showed that DNA synthesis was not affected with up to 50 mM alcohol, suggesting that G1-S phase in the cell cycle remained unaffected. At 100 mM alcohol, DNA synthesis decreased significantly (p < 0.01). From the results of this study, we conclude that a subpharmacological dose level of alcohol (10 mM) significantly inhibited cell mitosis and the inhibitory effect of alcohol was mediated in the G2-M phase in the cell cycle. The G1-S phase was unaffected.

Adult↗
PubMedDate not supplied

[Structural characteristics and availability of DNA for solvent molecules in nuclei of Physarum polycephalum].

The rate constants of 1H--3H exchange between water and C8H-groups of purinic residues of DNA in solution and in nuclei of synchronous Physarum polycephalum at different phases of the cell cycle were measured. The nuclear membranes and nonhistone proteins were shown not to create additional hindrances and not to diminish the availability of DNA in nuclei for solvent molecules. A small increase of the rate of the 1H--3H exchange between water and the DNA of nuclei upon transition from the S-phase to the late G2-phase seems to reflect the process of chromatin decondensation connected with activation of the transcription and local changes of the secondary structure of DNA at the late G2-phase of the cell cycle.

Cell Cycle↗
PubMed1999

Sodium butyrate induces G2 arrest in the human breast cancer cells MDA-MB-231 and renders them competent for DNA rereplication.

When exposed to sodium butyrate (NaBut), exponentially growing cells accumulate in G1 and G2 phases of the cell cycle. In the human breast cancer cell line MDA-MB-231, an arrest in G2 phase was observed when the cells were released from hydroxyurea block (G1/S interface) in the presence of NaBut. The inhibition of G2 progression was correlated with increased contents both of total p21(Waf1) and of p21(Waf1) associated with cyclin A and with an inhibition of cyclin A- and B1-associated histone H1 kinase activities measured in cell lysates, as well as with dephosphorylation of the RB protein. A decrease in the cell contents of cyclins A and B1 was also observed but this decrease was preceded by p21(Waf1) accumulation. When NaBut was removed from the culture medium of cells blocked in G2 phase, p21(Waf1) level decreased and, instead of proceeding to mitosis, these cells resumed a progression toward DNA rereplication. These results suggest that the induction of p21(Waf1) by NaBut leads to the inhibition of the sequential activation of cyclin A- and B1-dependent kinases in this cell line, resulting in the inhibition of G2 progression and rendering the cells competent for a new cell division cycle.

Breast Neoplasms↗
PubMed1991

Effects of caffeine and cycloheximide during G2 prophase in control and X-ray-irradiated human lymphocytes.

The effect of caffeine and cycloheximide during the G2 phase on frequency of chromosomal aberrations and G2 duration was studied in control and X-ray-irradiated human lymphocytes in vitro. Caffeine treatments alone increase the frequencies of chromatid breakage and decrease the average G2 duration in control and X-ray-irradiated lymphocytes (40 R). Both caffeine effects are reversed by 0.5 micrograms/ml cycloheximide in combination treatments. Cycloheximide treatments alone prolong G2 duration in control as well as in X-ray-irradiated lymphocytes although no improvement in chromosome repairing by this inhibitor of protein synthesis was observed under the conditions of our experiments. We propose that the cycloheximide effect is associated with a low level of mitotic factors, required for the entrance into mitosis, which is maintained at a higher level in caffeine treatment alone. Finally, G2 delay has generally been associated with certain genome damage. The fact that the caffeine and cycloheximide effects on X-irradiated lymphocytes are also present in control lymphocytes (without X-rays) suggests that control of the G2 duration constitutes one of the mechanisms involved in DNA repair operating during the G2 phase.

Antimutagenic Agents↗
PubMed2002

Retardation of the G2-M phase progression on gene disruption of RNA binding protein Sam68 in the DT40 cell line.

Sam68 is an RNA binding protein that is tyrosine-phosphorylated by Src during mitosis and has been postulated to have a role in cell cycle control by modulating RNA metabolism. To elucidate the function of this protein, we isolated a Sam68-deficient DT40 cell line by gene disruption. The Sam68-deficient cells exhibited markedly decreased growth and the growth retardation was due to elongation of the G2-M phase, however, the kinase activity associated with Cdc2 remained unaltered. Our results indicate that Sam68 may play a critical role in G2-M progression in a manner independent of the control of Cyclin/Cdc2 kinase activity.

Animals↗
PubMed1994

Bio-anticlastogenic effects of unsaturated fatty acids included in fish oil--docosahexaenoic acid, docosapentaenoic acid, and eicosapentaenoic acid--in cultured Chinese hamster cells.

Bio-anticlastogenic effects of unsaturated fatty acids--cis-4,7,10,13,16,19-docosahexaenoic acid (DHA), cis-7,10,13,16,19-docosapentaenoic acid (DPA), and cis-5,8,11,14,17-eicosapentaenoic acid (EPA)--on chemically induced chromosome aberrations were studied in cultured Chinese hamster cells. The induction of chromosome aberrations by the crosslinking agents mitomycin C (MMC) and cisplatin (DDP), the SN-1 type alkylating agents N-ethyl-N'-nitro-N-nitrosoguanidine (ENNG), methyl nitrosourea (MNU), and ethyl nitrosourea (ENU), and the SN-2 type alkylating agent ethyl methanesulfonate (EMS), but not by the SN-1 type alkylating agent N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) and the SN-2 type alkylating agent methyl methanesulfonate (MMS), was suppressed by post-treatment with DHA, DPA, and EPA. Since there was no opportunity to inactivate mutagens by desmutagenic mechanisms under the post-treatment schedule used, the results demonstrate the bio-anticlastogenicity of unsaturated fatty acids. Suppression by the unsaturated fatty acids was observed when cells were treated during the G2 phase, suggesting that G2 events were responsible for the bio-anticlastogenic effects. Two saturated fatty acids with the same number of carbons as the studied unsaturated fatty acids--docosanoic acid and eicosanoic acid--did not affect chromosome aberration induction, suggesting the necessity of unsaturation for fatty acid bio-anticlastogenicity.

Alkylating Agents↗
PubMed1996

UCN-01: a potent abrogator of G2 checkpoint function in cancer cells with disrupted p53.

BACKGROUND: Arrest of the cell cycle in G2 phase following DNA damage helps protect cell viability by allowing time for DNA repair before entry into mitosis (M phase). Abrogation of G2 arrest sensitizes cells to the effects of DNA-damaging agents. UCN-01 (7-hydroxystaurosporine), a protein kinase C inhibitor that may block G2 checkpoint regulation, has been reported to enhance the cytotoxicity of mitomycin C, a known DNA-damaging agent. PURPOSE: We studied the effect of UCN-01 on G2 checkpoint control in human lymphoma CA46 cells, whose sensitivity to various DNA-damaging agents and G2 response to DNA damage have been characterized. We also assessed the ability of UCN-01 to enhance the cytotoxicity of gamma irradiation in CA46 cells and human colon carcinoma HT-29 cells, both of which are mutant for p53 function. The influence of p53 function on UCN-01-mediated abrogation of the G2 checkpoint and enhancement of DNA-damaging agent cytotoxicity was studied in transfected human breast carcinoma MCF-7 cells that either expressed or did not express the human papillomavirus type-16 E6 protein. MCF-7 cells have normal p53 function, and the E6 protein binds p53 protein and promotes its destruction. METHODS: The effect of UCN-01 on cell cycle arrest induced by gamma irradiation was studied in CA46 cells and in transfected MCF-7 cells by use of flow cytometry. A histone H1 phosphorylation assay was employed to measure cyclin B1/Cdc2 kinase activity in extracts derived from irradiated and nonirradiated CA46 cells that had been either treated or not treated with UCN-01; the phosphorylation status of Cdc2 kinase protein in the same extracts was determined by use of western blotting. The effect of UCN-01 on the cytotoxicity of gamma irradiation in CA46 and HT-29 cells was determined by use of MTT (thiazolyl blue) and clonogenic (colony-forming) assays, respectively; a clonogenic assay was also used to measure the effect of UCN-01 on the cytotoxicity of cisplatin in transfected and nontransfected MCF-7 cells. RESULTS: G2 arrest induced in CA46 cells by gamma irradiation was minibited by treatment with UCN-01 in a dose-dependent manner; arrest in G2 was completely abrogated by exposure to 300 nM UCN-01. Biochemical markers indicative of the G2/M transition, including the activation of cyclin B1/Cdc2 kinase and the suppression of Cdc2 threonine-14 and tyrosine-15 phosphorylation, were detected in irradiated cells treated with UCN-01. UCN-01 enhanced the cytotoxicity of gamma irradiation in CA46 and HT-29 cells. MCF-7 cells with functional p53 protein were more resistant to G2 checkpoint abrogation by UCN-01 than MCF-7 cells with disrupted p53 function. UCN-01 markedly enhanced the cell-killing activity of cisplatin in MCF-7 cells defective for p53 function. CONCLUSIONS AND IMPLICATIONS: UCN-01 is a potent abrogator of G2 checkpoint control in cancer cells with disrupted p53 function. UCN-01 might be capable of enhancing the effectiveness of DNA-damaging agents in the treatment of tumors with cells lacking normal p53 function.

Alkaloids↗
PubMed1975

Cell cycles in the early development of axolotl.

In axolotl embryos, during the course of development, the generation time increases, chiefly on account of a lengthening of the phase of DNA synthesis, the longest in the cell cycle. At the stages of the middle blastula and early gastrula, the G1 phase is shorter than in the G2 phase, while at the neurula stage, the G1 phase is far longer than the G2 phase. Cells of the chordamesoderm pass through the cycle nonuniformly during gastrulation: a period with a very low index of labeled nuclei and slow entry of cells into the S phase gives way to a period of their intensive entry into the S-phase.

Ambystoma↗
PubMed1989

[In vivo antitumor activity of mitoxantrone and the flow cytometric analysis of its influence on cell cycle transition--comparison with doxorubicin and aclarubicin on ascitic hepatoma AH109A cells].

Mitoxantrone was compared with doxorubicin and aclarubicin of its in vivo antitumor activity and influence on cell cycle transition by use of rat ascitic hepatoma AH109A. Antitumor activity determined by the cell growth curve was similar in mitoxantrone and doxorubicin, but the sensitivity of AH109A to aclarubicin was lower than that to the other two drugs. Doxorubicin and mitoxantrone showed all phase arrests with 1/10 of maximally tolerated dose (MTD), and with lower concentrations a strong arrest at G2 phase was observed, thus, mitoxantrone appeared to have a similar antitumor activity on AH109A to that of doxorubicin. Aclarubicin, with 1/10 MTD, demonstrated only a transient arrest at G2 phase, cells arrested at G2 phase entering into the next phase. With below 1/10 MTD, there was no appearance on histograms, and the influence on AH109A cell cycle transition by aclarubicin was considered to be little in comparison with doxorubicin and mitoxantrone.

Aclarubicin↗
PubMed2006

Primary and compensatory roles for RB family members at cell cycle gene promoters that are deacetylated and downregulated in doxorubicin-induced senescence of breast cancer cells.

When treated with DNA-damaging chemotherapy agents, many cancer cells, in vivo and in vitro, undergo a terminal growth arrest and acquire a senescence-like phenotype. We investigated the molecular basis for this in breast cancer cells following a 2-hour treatment with 1 muM doxorubicin. Treated cells arrested in G1 and G2 phases of the cell cycle, with concomitant reductions in S-phase and G2-M regulatory genes. p53 and p21 protein levels increased within hours after treatment and were maintained for 5 to 6 days but were reduced 8 days posttreatment, though the cells remained growth arrested. Levels of p130 rose after drug treatment, and it was the primary RB family member recruited to the S-phase promoters cyclin A and PCNA and G2-M promoters cyclin B and cdc2, remaining present for the entire 8-day time period. In contrast, p107 protein and promoter occupancy levels declined sharply after drug treatment. RB was recruited to only the PCNA promoter. In MCF-7 cells with p130 knockdown, p107 compensated for p130 loss at all cell cycle gene promoters examined, allowing cells to retain the growth arrest phenotype. Cells with p130 and p107 knockdown similarly arrested, while cells with knockdown of all three family members failed to downregulate cyclin A and cyclin B. These results demonstrate a mechanistic role for p130 and compensatory roles for p107 and RB in the long-term senescence-like growth arrest response of breast cancer cells to DNA damage.

Acetylation↗
PubMed1990

Cell cycle synchronization: reversible induction of G2 synchrony in cultured rodent and human diploid fibroblasts.

In accord with a set of prespecified principles of cell synchrony induction, a three-step procedure was developed to arrest cells reversibly in the G2 phase of the cell cycle. Cultures of Chinese hamster ovary (CHO) cells were presynchronized in early S phase by sequential treatment with isoleucine deficiency and hydroxyurea blockades; then they were switched to medium supplemented with either of two agents that inhibit DNA topoisomerase II activity by different mechanisms, Hoechst 33342 at 7.5 micrograms/ml for 12 hr or VM-26 at 0.5 micrograms/ml for 8 hr. Up to 95% of the cells accumulated in G2 phase under those conditions. After switch of Hoechst 33342-treated cells to drug-free medium, the cells divided as a highly synchronized cohort of cells within 3 hr. Up to 85% of the cells in a culture of human diploid dermal fibroblasts (HSF-55 cells) could be accumulated in G2 phase by placing cells presynchronized in early-S phase in medium containing Hoechst 33342 at 0.1 micrograms/ml for 10 hr. Reversal of G2 arrest in the HSF-55 cultures resulted in cells dividing synchronously over 3.5 hr. By varying the concentration of Hoechst 33342 and the duration of the treatment period, it was possible to alter the position within G2 phase at which cells accumulated. This synchronization protocol should greatly facilitate study of G2/M biochemical events in mammalian cells, in particular, those associated with cdc2 gene regulation of the onset of mitosis.

Animals↗
PubMed1986

A two-parameter flow cytometry protocol for the detection and characterization of the clastogenic, cytostatic and cytotoxic activities of chemicals.

Cultured, freshly-isolated rat fibroblasts were exposed in vitro to vincristine sulphate (VC), amethopterin (AM), bleomycin (BL), benomyl (BE) and practolol (PR). Cells treated for 5 h were subjected 24 h later to a two-parameter (DNA/protein) flow cytometry analysis. The fluorochromes used were sulphorhodamin 101 and DAPI. From DNA and protein histograms, alterations in cell-cycle kinetics, variations in the amount of DNA in individual G1-phase cells and the enhancement of or increased variation in the protein content of the exposed cells were determined. Each of the 5 chemicals induced a specific dose-dependent pattern of changes in the DNA and protein histograms. DNA dispersion was enhanced with VC, AM, BL and BE but not with PR. The cell cycle was blocked in the G2 phase with VC, at early S phase with amethopterin and, depending on the dose, at the G1 or G2 phase with bleomycin or at the S phase or G2 phase with benomyl. Practolol inhibited cells slightly in the S phase at the highest exposure level. Protein analysis allows cytotoxic activity (loss of proteins) or induced unbalanced growth (protein accumulation) of test compounds to be recognized. The results obtained imply that the proposed two-parameter DNA/protein analysis by flow cytometry is a suitable method for prospective testing of chemicals for their induction of structural or numerical chromosome aberrations. Simultaneously, a broad range of cytotoxic, cytostatic and cell-cycle perturbing activities of the test agents can be recognized.

Animals↗
PubMed1981

[Structural study of the chromatin of the myxomycete Physarum polycephalum in the mitotic cycle].

The chromatin structure of Physarum polycephalum was studied with electron microscope at different phases of its mitotic cycle. At the S-phase and during mitosis, the chromatin has a nucleosomal structure. At the early G2-phase the chromatin structure changes, long regions of non-beaded structure being found in the chromatin fibers. At the late G2-phase, the major part of chromatin loses its globular organization, with chromatin fibres without a pronounced subunit structure prevailing in the preparations. Biochemical data show that the amount of chromatin resistant to staphylococcal nuclease varies during the mitotic cycle. The amount of nuclease-resistant chromatin is equal to 80% at the S-phase, to decrease up to 50-60% by the early G2-phase. Successive changes of chromatin structure at different levels of its transcriptional activity are found. Lability of nucleosomes is shown to increase with the increase in the transcriptional activity of chromatin, thus leading presumably to the chromatin structural alterations during the mitotic cycle.

Chromatin↗
PubMed1976

Survival and cycle-progression delay of human lymphoma cells in vitro exposed to VP-16-213.

The lethal and kinetic effects of VP-16-213 were analyzed in a human lymphoid cell line (T1 cells) in vitro. When asynchronous T1 cells were exposed to increasing concentrations of VP-16-213 for 1 hour, an exponential survival curve with a Do (mean lethal dose equal to the concentration required to reduce survival by 63% on the exponential part of the survival curve) of 3 mug/ml was obtained. Increasing exposure time also reduced survival exponentially. Synchronized cells showed age-dependent sensitivity to VP-16-213 with the greatest lethal damage experienced by cells treated in S and G2 phase. The major kinetic response of the T1 cells to VP-16-213 was a delay in G2 phase, the extent and duration of which was a function of drug concentration, exposure time, and cell cycle stage of drug addition; thus, cells in S phase were most effectively blocked in the subsequent G2 phase. Continuous treatment with concentrations of VP-16-213 (greater than 0.5 mug/ml for greater than 3 hours) caused a retardation of S-phase transit with prompt recovery after drug release. Treatment with 10.0 mug/ml for greater than 3 hours resulted in a "frozen state" of the whole life cycle, inducing only minor compartment changes. DNA synthesis was inhibited in the majority of cells with an S-phase DNA content. There was no correlation between the extent of perturbation and lethal effects after treatment with VP-16-213.

Cell Division↗
PubMed2001

The Chk1-Cdc25C regulation is involved in sensitizing A253 cells to a novel topoisomerase I inhibitor BNP1350 by bax gene transfer.

Promotion of apoptosis may potentiate the sensitivity of tumor cells to chemotherapeutic agents, thus improving the efficacy of cancer treatment. The transfection of the proapoptotic bax gene, which results in the overexpression of bax protein, augments the growth inhibition of A253 cells by BNP1350. Increased drug response was associated with the induction of DNA fragmentation in the size of 30-200 Kb, generating a cleaved fragment of 18 kDa from full-length 21 kDa bax and the cleavage of PARP. A253/vec cells treated with 0.07 microM(IC50) of BNP1350 accumulated in G2 phase at 24 h after drug removal. In contrast, A253/Bax cells treated with an equimolar concentration of BNP1350 primarily displayed a G1 phase accumulation with a concurrent decrease in G2 phase. Certain cell cycle regulatory protein expression and activities were altered following drug exposure in both cell lines under similar conditions. Cdk2- and cdc2-associated H1 kinase activities were markedly increased in the A253/Bax cell line with marginal increased activity in the A253/vec cell line. A chk1 activity assay was performed with GST-cdc25C (200-256) or GST-cdc25C(S216A) (200-256) fusion proteins as the substrate. Increased chk1 activity was observed in the A253/vec cell line, with little change in the A253/Bax cell line, when exposed to equimolar concentrations of BNP1350 (0.07 microM). A Western blot of immunoprecipitated chk1 indicated that increased chk1 phosphorylation following DNA damage induced by BNP1350 was accompanied by the observed G2 accumulation in the A253/vec cell line, while only a slight increase in chk1 phosphorylation was seen in the A253/Bax cell line. A decreased expression of cdc25C was observed in the BNP1350-treated A253/Bax cells, but not in the A253/vec cell line. Following exposure to BNP1350, increased binding of 14-3-3 proteins to chk1 occurred in both cell lines, with more being observed in the A253/vec cell line. The data have shown that inhibition of the chk1 pathway accompanied by the abrogation of G2 arrest is involved in sensitizing A253 cells to BNP1350 by bax gene transfer. These findings suggest that bax gene transfer sensitizes A253 cells to BNP1350 through apoptosis promoting and G2/M DNA damage checkpoint regulatory pathways.

Apoptosis↗