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At least 19 recordsLinked to original sources
PubMed1997

EGF- and CPA-induced mitogenic stimuli are differentially down-regulated by TGF-beta1 in cultured rat hepatocytes.

Down-regulation of the mitogenic activity of the rodent liver carcinogen cyproterone acetate (CPA) and of epidermal growth factor (EGF) were compared in cultured rat hepatocytes. Both hepatomitogens produce an increase in the expression of proliferating cell nuclear antigen (PCNA) and in [3H]thymidine incorporation in a dose-dependent manner. In combination, the two mitogens induced an additive mitogenic response. Concomitant exposure to the growth inhibitory cytokine transforming growth factor beta1 (TGF-beta1) resulted in a differential dose-dependent down-regulation of PCNA-expressing cells. The corresponding down-regulation of CPA-induced PCNA expression required a 3- to 5-fold higher TGF-beta1 concentration than for EGF-induced expression. In contrast, CPA-exposed hepatocytes become vulnerable to and EGF-exposed cells protected against the apoptosis-inducing activity of TGF-beta1 (>0.1 ng/ml). Under culture conditions that mimicked a pericentral-equivalent microenvironment (low oxygen tension, low glucagon concentration), PCNA expression was 3-fold lower and CPA-specific resistance was no longer detectable. It is concluded that EGF and CPA induce their growth stimuli preferentially in the periportal area of the liver but in different hepatocyte sub-populations, which differ in their down-regulation of premitotic events by TGF-beta1. At low TGF-beta1 concentrations, EGF-stimulated cells shift back into a resting cell cycle phase, whereas CPA-treated hepatocytes are eliminated by apoptosis at higher TGF-beta1 concentrations.

Animals↗
PubMed1997

Terminal complement complexes induce cell cycle entry in oligodendrocytes through mitogen activated protein kinase pathway.

Sublytic complement attack through C5b-9 assembly induces oligodendrocytes (OLG) to express proto-oncogenes and to enter the cell cycle from resting G0/G1 phase to S phase. We have investigated whether cell cycle induction by C5b-9 is mediated by mitogen activated protein kinase (MAPK) pathway in OLG. C5b-9 but not C5b6 induced activation of both ERK1 and c-jun NH2 terminal kinases 1 (JNK1) in OLG. The increased ERK1 and JNK1 activities are transient, reaching a maximum around 20 min following exposure to C5b-9. Activation of Raf-1 and MEK1, upstream kinases of ERK1, was shown by increased Raf-1 kinase activity in anti-Raf-1 immunoprecipitates of OLG treated with C5b-9 and ERK1 activity that can be inhibited by PD098,059, a specific MEK1 inhibitor. Requirement for the ERK1 pathway in DNA synthesis was then evaluated using PD098,059. Enhanced DNA synthesis induced by serum complement was completely abolished when OLG were pretreated with PD098,059. On the other hand, c-fos mRNA expression induced by complement was inhibited only 50% by PD098,059, while the c-jun mRNA level was not affected by this MEK1 inhibitor. Interestingly, p70 S6 kinase, an important ribosomal kinase in mitogenesis, was also activated by C5b-9. These findings indicated that the MAPK pathways appears to play a major role in inducing OLG to enter the S phase of the cell cycle from the resting G1/G0 phase.

Animals↗
PubMedDate not supplied

[The dependence of the cytokinetic effects of alkylating antitumor preparations on the phase of the hepatocyte cell cycle in regenerating liver].

Effects of alkylating antitumor drugs on resting (G0 phase of cell cycle) and proliferating (G1, S, G2 and M phases) hepatocytes were studied in regenerating mouse liver. Cell cycle kinetics (fraction of labeled mitoses, labeling and mitotic indices) were determined by 3H-thymidine autoradiography. Dipin and fotrin as a DNA-damaging agents attack mainly resting (G0) and proliferating (G1) cells. Effect of the damage results in the inhibition of DNA synthesis and G2 phase arrest in the following mitotic cycle. An alkylating drug phopurin as well as ara-C both suppress the mitotic progression in proliferating hepatocytes and do not influence the resting cells.

Alkylating Agents↗
PubMed1991

Mutations in the thymidine kinase gene that allow expression of the enzyme in quiescent (G0) cells.

Thymidine kinase (TK) is a nucleotide salvage pathway enzyme whose activity is highly dependent on the growth state and cell cycle phase of a cell. Cells in the resting or quiescent (G0) phase express very low levels of TK mRNA and protein. When quiescent cells are stimulated to enter the cell cycle by the addition of serum, TK mRNA, activity and polypeptide increase coordinately after about 10-15 h, at the beginning of S phase. When growth-independent heterologous promoters are substituted for the natural TK promoter, TK mRNA can be expressed in quiescent cells. Despite the presence of TK mRNA in such G0 cells, there is little expression of TK polypeptide; the normal increase in enzyme at S phase is observed following serum stimulation. Deletion of the introns and 3' untranslated sequences does not affect the expression of the TK gene in serum stimulation experiments. In contrast, deletion of the C-terminal 40 amino acids or fusion of a small segment of a beta-galactosidase to the C-terminus overcomes the block to expression of the TK polypeptide in G0 cells. These C-terminal alterations are the same as those which lead to constitutive expression of TK during the cell cycle of proliferating cells, suggesting that mechanisms which control the levels of TK in cycling cells may also operate in quiescent cells.

Animals↗
PubMed1998

Peripheral blood lymphocytes of bipolar affective patients have a histone synthetic profile indicative of an active cell state.

1. Although abnormalities of the immune system have been described in depression, no information exists regarding the biochemical parameters which could characterize the physiological state of lymphocytes from patients with bipolar affective disorder. 2. Lymphocytes of normal control subjects are known to be in the Go resting phase of the cell cycle. Histone synthesis is characteristically different during the Go, G1/G2 and the S phases of the cell cycle. As such, it can be used as a biochemical marker with which to distinguish between cycling and noncycling cells. 3. In order to investigate the possibility of whether or not the lymphocytes of patients with bipolar affective disorder are in an activated state, typical of cycling cells, total histone and histone variant synthesis were analysed in peripheral blood lymphocytes of a group of 12 patients with bipolar affective disorder and 7 normal controls. 4. According to the histone variant synthesis pattern, lymphocytes of patients in normothymia have values similar to those of controls, i.e., of noncycling cells, while patients in either the depressed or the manic phase have values intermediate to those of resting and cycling cells. 5. This study shows that histone synthesis can perhaps be used as a biochemical parameter of possible significance in differentiating amongst the three phases of the illness.

Adult↗
PubMed2006

Chromosome or chromatin condensation leads to meiosis or apoptosis in stationary yeast (Saccharomyces cerevisiae) cells.

When starved of essential nutrients, yeast cells cease mitotic division and enter an alternative state called the 'stationary phase'. In this paper, we report that stationary cells enter two major pathways: meiosis and apoptosis. Using transmission electron microscopy, five types of cell were identified in the stationary phase: (1) cells with chromosome condensed nuclei; (2) cells with normal, homogeneously stained nuclei; (3) sporulated cells; (4) apoptotic cells, in which chromatin, but not individual chromosomes, was condensed; and (5) dead cells, in which nuclei and cytoplasm were degraded. Further evidence using live cell imaging and mutation analysis suggested that cells with condensed chromosomes underwent meiosis, whereas chromatin condensed cells underwent apoptotic cell death. Cells with homogeneous nuclei are believed to be in the true resting state and undergo cell death when starvation continues. Chromosome or chromatin condensation may serve as a hallmark of life or death for stationary cells.

Apoptosis↗
PubMed1999

Nonproductive human immunodeficiency virus type 1 infection in nucleoside-treated G0 lymphocytes.

Productive infection by human immunodeficiency virus type 1 (HIV-1) requires the activation of target cells. Infection of quiescent peripheral CD4 lymphocytes by HIV-1 results in incomplete, labile, reverse transcripts. We have previously identified G1b as the cell cycle stage required for the optimal completion of the reverse transcription process in T lymphocytes. However, the mechanism(s) involved in the blockage of reverse transcription remains undefined. In this study we investigated whether nucleotide levels influence viral reverse transcription in G0 cells. For this purpose the role of the enzyme ribonucleotide reductase was bypassed, by adding exogenous deoxyribonucleosides to highly purified T cells in the G0 or the G1a phase of the cell cycle. Our data showed a significant increase in the efficiency of the reverse transcription process following the addition of the deoxyribonucleosides. To define the stability and functionality of these full reverse transcripts, we used an HIV-1 reporter virus that expresses the murine heat-stable antigen on the surfaces of infected cells. Following activation of infected quiescent cells treated with exogenous nucleosides, no increased rescue of productive infection was seen. Thus, in addition to failure to complete reverse transcription, there was an additional nonreversible blockage of productive infection in quiescent T cells. These experiments have important relevance in the gene therapy arena, in terms of improving the ability of lentivirus vectors to enter metabolically inactive cells, such as hematopoietic stem cells.

Animals↗
PubMed2003

Molecular mechanisms of G0/G1 cell-cycle arrest and apoptosis induced by terfenadine in human cancer cells.

Terfenadine (TF), a highly potent histamine H1 receptor antagonist, has been shown to exert no significant central nervous system side effects in clinically effective doses. In this study, we demonstrated that TF induced significant growth inhibition of human cancer cells, including Hep G2, HT 29, and COLO 205 cells, through induction of G(0)/G(1) phase cell-cycle arrest. The minimal dose of TF induced significant G(0)/G(1) arrest in these cells was 1-3 microM. The protein levels of p53, p21/Cip1, and p27/Kip1 were significantly elevated, whereas the kinase activities of cyclin-dependent kinase 2 (CDK2) and CDK4 were inhibited simultaneously in the TF-treated cells. On the other hand, significant apoptosis, but not G(0)/G(1) arrest, was induced in the HL 60 (p53-null) or Hep 3B (with deleted p53) cells when treated with TF (3-5 microM). To clarify the roles of p21/Cip1 and p27/Kip1 protein expression, which was involved in G(0)/G(1) arrest and apoptosis induced by TF in human cancer cells, antisense oligodeoxynucleotides (ODNs) specific to p21/Cip1 and p27/Kip1 were used, and the expression of the p21/Cip1 and p27/Kip1 were monitored by immunoblotting analysis. Our data demonstrated that the percentage of the apoptotic cells detected by annexin V/PI analysis in the TF-treated group was clearly attenuated by pretreatment with p27/Kip1-specific ODNs. These results indicated that p27/Kip1 (but not p21/Cip1) protein indeed played a critical role in the TF-induced apoptosis. We also demonstrated that the TF-induced G(0)/G(1) cell-cycle arrest effect was not reversed by TF removal, and this growth inhibition lasted for at least 7 d. Importantly, the occurrence of apoptosis and cell growth arrest was not observed in the TF-treated normal human fibroblast, even at a dose as high as 25 microM. Our study showed the molecular mechanisms for TF-induced cell growth inhibition and the occurrence of apoptosis in human cancer cells.

Apoptosis↗
PubMed2001

Ras-dependent cell cycle commitment during G2 phase.

Synchronization used to study cell cycle progression may change the characteristics of rapidly proliferating cells. By combining time-lapse, quantitative fluorescent microscopy and microinjection, we have established a method to analyze the cell cycle progression of individual cells without synchronization. This new approach revealed that rapidly growing NIH3T3 cells make a Ras-dependent commitment for completion of the next cell cycle while they are in G2 phase of the preceding cell cycle. Thus, Ras activity during G2 phase induces cyclin D1 expression. This expression continues through the next G1 phase even in the absence of Ras activity, and drives cells into S phase.

Animals↗
PubMed2004

Cell cycle stage analysis of rabbit foetal fibroblasts and cumulus cells.

Synchronization of the cell cycle stages in G0/G1 phase is one of the key factors determining the success of nuclear transplantation. Serum deprivation, contact inhibition and chemical inhibitors are widely used methods for this purpose. In this study, cell cycle stages of foetal fibroblasts and cumulus cells were determined using flow cytometry [fluorescence-activated cell scan (FACS)]. Foetal fibroblasts (in vitro cultured for 72-120 h) and fresh cumulus cells were analysed in Experiment 1. Fifty to 55% proliferating fibroblasts remained in G0/G1 phase compared with 78% in confluent culture (p <0.05). In contrast to foetal fibroblasts, fresh cumulus cells maintained 90% of the population in the G0/G1 stage. When serum was retrieved from the proliferating fibroblasts from day 1 to day 5 (Experiment 2), proportions of G0/G1 cells increased from the initial ratio of 53 to 87% at day 4 of starvation, which was significantly higher than the non-starved proliferating cells (p <0.05). In Experiment 3, fibroblasts were treated with aphidicolin (0.1 microg/ml, 6 h), demicolcine (0.5 microg/ml, 10 h), or a combination of these two chemicals to synchronize the cell cycle stages. Surprisingly, no differences or significantly lower in the proportions of G0/G1)phase cells were detected (25-50%) compared with the uncontrolled growing cells (53%). These results suggested that fresh cumulus cells rest their cell cycle in G0/G1 stage. Serum deprivation became effective in the first 24 h and reached the highest proportions during days 4-5 after deprivation. Chemical synchronization of the cell cycle stage of rabbit foetal fibroblasts to G0/G1 phase appeared less effective compared to serum deprivation.

Animals↗
PubMed1998

On the proposal of a G0 phase and the restriction point.

Zetterberg and Larsson proposed that the restriction point divides the G1 phase into two parts. Cells before this point are able to leave the division cycle and enter a G0 phase; cells past this point are unaffected by a short period of low serum. Additional results of Zetterberg and Larsson--1) cycloheximide treatment affects cells in the same way as low serum, and 2) a delay in the second division after serum starvation in the cells not initially affected in their first division--indicate that their experiments are consistent with serum removal affecting cells in all phases of the cell cycle equally. Their experiments are consistent with the continuum model of the mammalian division cycle. There is no need to postulate a restriction point or a G0 phase to explain the serum starvation results.

Cell Cycle↗
PubMed1996

Cell cycle-dependent tumor necrosis factor apoptosis.

To determine if tumor necrosis factor (TNF)-mediated apoptosis affects cells at defined stages of the cell cycle, WEHI-164/2F (WEHI) cells were synchronized at G0-G1 after 3-day cultures in medium containing RPMI 1640 and 0.5% FCS (RPMI-0.5% FCS). The arrested WEHI cells (60-75% in G0-G1) showed increased sensitivity to TNF killing, measured as 48-h 3-(5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide assays, and 15-h apoptosis by propidium iodide staining and flow cytometry analysis. The TNF killing kinetics of G0-G1-arrested cells was similar to controls, and TNF did not accelerate or retard cell cycle progression of the arrested cells after feeding with fresh RPMI-0.5% FCS. However, TNF inhibited WEHI DNA synthesis as early as 1 h after treatment, and inhibition was proportionate to sensitivity to TNF-induced apoptosis. WEHI cells treated with TNF showed a higher percentage of cells in S phase with concomitant decrease in G0-G1 and G2-M. When cultured for 3-18 h in fresh RPMI-0.5% FCS to allow progression of the G0-G1-arrested cells toward the G1-S boundary, WEHI cells became more sensitive to TNF killing, especially at the 3-9 h time points. Moreover, TNF did not degrade [125I]5-iodo-2'-deoxyuridine-labeled WEHI DNA if the labeled cells were precultured for 9 h in fresh RPMI-0.5% FCS to allow them to pass S phase before the addition of TNF. These results show that TNF-induced apoptosis of WEHI cells is connected to cell cycle events; WEHI targets receive the TNF cytotoxic signal mainly at the G1-S boundary and begin to die by apoptosis as they exit from S phase.

Apoptosis↗
PubMed1998

Endogenous E2F-1 promotes timely G0 exit of resting mouse embryo fibroblasts.

Much evidence strongly suggests a positive role for one or more E2F species in the control of exit from G0/G1. Results described here provide direct evidence that endogenous E2F-1, as predicted, contributes to progression from G0 to S. By contrast, cycling cells lacking an intact E2F-1 gene demonstrated normal cell cycle distribution. Therefore, E2F-1 exerts a unique function leading to timely G0 exit of resting cultured primary cells, while at the same time being unnecessary for normal G1 to S phase progression of cycling cells.

Animals↗
PubMed1993

Growth inhibition of human keratinocytes by 1,25-dihydroxyvitamin D3 is linked to dephosphorylation of retinoblastoma gene product.

Human keratinocyte is one of the target cells for 1,25(OH)2D3, a biologically active form of vitamin D3. It induces the differentiation and growth inhibition of human keratinocytes. In order to understand the inhibitory mechanism of 1,25(OH)2D3, we examined its effect on cell cycle kinetics and retinoblastoma gene product (pRB), one of tumor suppressor gene products, in normal human keratinocytes. Cell cycle analysis demonstrated that 10(-6) M of 1,25(OH)2D3 induced cell cycle arrest in both G1/G0 (63.4% +/- 1.4 versus 52.7% +/- 1.2 in control, p < 0.05) and G2 + M (21.5% +/- 0.6 versus 10.9% +/- 0.8 in control, P < 0.05) phase. Addition of 10(-6) M of 1,25(OH)2D3 increased dephosphorylated pRB in a time dependent manner from 23% at 0 h to 58% at 48 h. Since the phosphorylation of pRB is supposed to be essential for the progression from G1 to S phase, the inhibition of pRB phosphorylation could be responsible for the G1/G0 growth arrest induced by 1,25(OH)2D3 in normal human keratinocytes.

Calcitriol↗
PubMed2003

Cell differentiation and proliferation--simultaneous but independent?

Despite studies over many years, it is still not clear to what extent cellular controls on proliferation and on differentiation are interrelated. For example, the idea that exit from cell cycle into G1/G0 is a necessary-or at least frequent-prelude to differentiation developed partly from studies of haemopoietic cell maturation, often using cell lines as models. The responses of cells to treatment with differentiating agents suggested that exit from cell cycle into G1/G0 occurs quite quickly, with functional differentiated characteristics acquired later, and so promoted the notion that cyclin-dependent kinase inhibitors (CDKIs) might be important initiators of normal differentiation. However, recent work has made it clear that differentiation and cell proliferation are regulated simultaneously but independently, that cells often start differentiating long before they stop dividing, and that the launching of differentiation is not restricted to any particular segment of the cell cycle. This combination of attributes allows expansion of cell numbers and acquisition of differentiated function to occur in parallel, generating abundant effector cells. Given this dichotomy, future studies to develop a more exact picture of the events that initiate and drive differentiation might be simplified by studying differentiation under experimental conditions that divorce it from concerns about cell cycle control.

Animals↗
PubMed1993

Application of a DNA double labelling method for the flow cytometric analysis of recruitment of non-cycling cells in a mixed population of P and Q cells.

In this paper we describe the application of a non-radioactive DNA double labelling and staining method to an analysis of cell proliferation kinetics by flow cytometry, aimed at the direct measurement of recruitment rates in cell cultures. The method is based on the application of two halogenated deoxyuridines: iododeoxyuridine (IdUrd) and chlorodeoxyuridine (CldUrd) which are incorporated into DNA synthesizing cells. By applying two commercially available monoclonal antibodies both deoxyuridines can be detected separately. To measure recruitment all proliferating cells in a plateau phase culture were labelled first with IdUrd applied during a time interval approximately equal to the cell cycle time. Subsequently, recruitment induced by a medium change was analysed by flow cytometric assessment of incorporation of CldUrd in cells which had not taken up IdUrd. Experiments designed to determine the toxicity of continuous labelling with IdUrd in different concentrations and of pulse labelling with CldUrd showed that there was no effect on the progression of cells through the cell cycle. The aim of this study is to test the sensitivity of the procedure to detect changes in proliferation kinetics, in particular the entrance of resting cells into the S phase. Although the cell culture model used is very simple, the results demonstrate clearly that a low rate of recruitment can be detected. It is suggested that the procedure described here is specific and sensitive enough to quantify changes in cell proliferation in tumours induced by various treatments and has advantages over other methods, which measure recruitment indirectly, or directly by using two radioactive thymidines.

Animals↗
PubMed1997

The effect of butylated hydroxytoluene on the chromosomal damage induced by bleomycin in Chinese hamster ovary cells.

The effect of butylated hydroxytoluene (BHT) on chromosomal damage induced by bleomycin (BLM) in CHO cells was studied. Treatments were performed in cells at quiescent state (90/95% at G0-G1), 2 h after subculture (G1) or 3 h before fixation (G2). Cells were treated for 20 min with BLM plus BHT and subsequently incubated in the presence of BHT until fixation. Results were compared with those obtained from untreated and DMSO-treated controls, from treatments with BLM or BHT alone, from treatments with BLM followed by treatment with DMSO until fixation, and from treatments with BLM plus BHT for 20 min without post-treatment with BHT. BLM induced chromatid- and chromosome-type aberrations in cells treated at G0/G1 or G1 and chromatid-type aberrations in cells treated at the G2 stage. Post-treatment with BHT strongly decreased the frequency of chromosome- but not of chromatid-type aberrations in G0/G1 and G1 and of chromatid-type aberrations in G2. These results are explained assuming that chromosome-type aberrations induced at G0/G1 and G1, and chromatid-type aberrations induced at G2 are originated by the induction of double-strand breaks by BLM through the formation of free radicals. Thus, the observed effect of BHT post-treatment could be considered as evidence that chromosome aberrations are induced by BLM following a two-step mechanism. On the other hand, it is necessary to differentiate between chromatid-type aberrations induced by BLM at G0/G1 and those produced by G2 treatment on the basis of replication errors for the former and DNA repair errors for the latter. In addition, the induction of chromatid-type aberrations by BHT itself at G0/G1 must be taken into account. As BHT acts as an S-dependent agent, chromatid-type aberrations observed after treatment with BLM and BHT in G0/G1 could arise from single-strand breaks induced by BLM and DNA primary lesions induced by BHT.

Animals↗