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C Dive

Publications and source records attributed to C Dive.

At least 37 records · Page 2Linked to original sources
PubMed1999

Survival signals within the tumour microenvironment suppress drug-induced apoptosis: lessons learned from B lymphomas.

The suppression of apoptosis is one mechanism by which tumours become drug resitant. Extracellular signals from the germinal centre (GC) of secondary lymphoid tissue can rescue B cells from physiological- and chemotherapy-induced apoptosis. Such survival signals include CD40 receptor ligation, interleukin-4 (IL-4) receptor stimulation and the interaction of the integrin ligand VCAM-1 with its receptor. The GC environment was modelled in vitro by providing B lymphoma cells with these survival signals. JLP119 B lymphoma cells underwent apoptosis after exposure to the topisomerase II inhibitor etoposide and this was dramatically reduced when the cells were cultured in the GC system. CD40 receptor ligation resulted in increased levels of Bcl-XL. Etoposide diminished the binding between Bax and Bcl-XL but this was restored by IL-4 and VCAM-1 triggered signals. These data demonstrate combined effects of three microenvironmental signals on the Bcl-2 family and illustrate the potential importance of such signalling pathways in drug resistance of tumour cells.

Animals↗
PubMed1998

Blood cells with reduced mitochondrial membrane potential and cytosolic cytochrome C can survive and maintain clonogenicity given appropriate signals to suppress apoptosis.

Reduction of mitochondrial membrane potential (Psim) and release of cytochrome c from mitochondria appear to be key events during apoptosis. Apoptosis was induced in IC.DP premast cells by the withdrawal of interleukin-3 (IL-3). Psim decreased by 12 hours and cytochrome c was detected in the cytosol at 18 hours. Despite these changes in the mitochondria after 18 hours of IL-3 deprivation, clonogenicity was unaffected when IL-3 was replenished at 18 hours. Activation of v-Abl tyrosine kinase (v-Abl TK) in IC.DP cells before IL-3 depletion led to increased levels of Bcl-XL, prevented reduction of Psim and the release of mitochondrial cytochrome c, and suppressed apoptosis. Activation of v-Abl TK 18 hours after withdrawal of IL-3 when </=10% of the cells had died restored Psim in the remaining cells. More than 40% of cells thus rescued by v-Abl TK between 18 and 42 hours could subsequently form colonies in the presence of IL-3. These data suggest that reduction in Psim precedes loss of mitochondrial cytochrome c in IC.DP cells; that v-Abl TK activation, probably via upregulation of Bcl-XL, prevents loss of Psim and blocks the release of cytochrome c from mitochondria; and that neither of these mitochondrial events is sufficient for commitment to apoptosis.

Animals↗
PubMed1998

Comparative effects of Bcl-2 over-expression and ZVAD.FMK treatment on dexamethasone and VP16-induced apoptosis in CEM cells.

It is becoming apparent that caspases are essential mediators of the execution phase of apoptosis. A decrease in mitochondrial membrane potential (PsiM) is also thought to be an early event in apoptosis. In this study, we compare the effects of Bcl-2 over-expression against N-benzyloxycabonyl-Val-Ala-Asp-fluoromethylketone (ZVAD. FMK)-sensitive caspase blockade on dexamethasone (DEX) and etoposide (VP16)-induced apoptosis in CEM T lymphoid cells. We assessed changes in nuclear chromatin, cell size, fragmentation, cell membrane permeability and PsiM. We found Bcl-2 over-expression and ZVAD.FMK-sensitive caspase inhibition were able to prevent chromatin condensation and cellular fragmentation. However, ZVAD.FMK was neither able to prevent loss of plasma membrane integrity nor PsiM depolarization which occur in both VP16 and DEX-induced apoptosis. In VP16-induced apoptosis, the increase in cell membrane permeability was actually potentiated by caspase inhibition. Interestingly, ZVAD.FMK did prevent VP16-induced but not DEX-induced cell shrinkage. These results suggest that not all the actions of Bcl-2 can be explained by its ability to prevent caspase activation. Rather Bcl-2 must have other targets of action which include functions associated with mitochondria.

Amino Acid Chloromethyl Ketones↗
PubMed1998

The rodent non-genotoxic hepatocarcinogen nafenopin suppresses apoptosis preferentially in non-cycling hepatocytes but also elevates CDK4, a cell cycle progression factor.

Rodent non-genotoxic hepatocarcinogens such as nafenopin suppress spontaneous and transforming growth factor beta1 (TGFbeta1)-induced rat hepatocyte apoptosis as well as inducing DNA synthesis. We wished to determine if these two processes are associated. In primary rat hepatocytes, nafenopin suppressed apoptosis from 1.9 to 0.63% but more apoptotic bodies were bromodeoxyuridine (BrdU)-labelled (0.35%) than predicted statistically from a random distribution of apoptosis within the cycling and non-cycling populations (0.10%). In contrast, TGFbeta1 induced hepatocyte apoptosis (7.8%) but fewer hepatocytes were BrdU-labelled (0.29%) than predicted (0.82%). Western blot analyses showed that nafenopin and TGFbeta1 had opposing effects on cyclin-dependent kinase 4 (CDK4) protein: nafenopin elevated CDK4 compared with controls, whereas TGFbeta1 caused a reduction. These data suggest that non-genotoxic hepatocarcinogens suppress apoptosis in the non-cycling population of hepatocytes and elevate CDK4 levels, possibly allowing potentially tumourigenic cells to enter the cell cycle.

Animals↗
PubMed1998

The non-genotoxic hepatocarcinogen nafenopin suppresses rodent hepatocyte apoptosis induced by TGFbeta1, DNA damage and Fas.

The suppression of apoptosis may contribute to the carcinogenicity of the peroxisome proliferators (PPs), a class of non-genotoxic rodent hepatocarcinogens. Our previous work demonstrated that the PP nafenopin suppressed both spontaneous and transforming growth factor beta1 (TGFbeta1)-induced hepatocyte apoptosis both in vivo and in vitro. Here, we extend these observations by demonstrating the ability of nafenopin to suppress apoptosis induced by other major candidates for the signalling of cell death in the liver. Treatment of rat or mouse hepatocyte monolayers with TGFbeta1 or the DNA damaging drugs etoposide or hydroxyurea induced high levels of apoptosis. Western blot analysis did not support a role for either p53 or p21waf1 in etoposide-induced apoptosis in rat hepatocytes. Treatment of mouse hepatocytes with an agonistic anti-Fas antibody also resulted in an induction of high levels of apoptosis. Pre-addition and continued exposure to nafenopin suppressed apoptosis induced by all three stimuli. Overall, our studies demonstrate that the ability of nafenopin to protect hepatocytes from apoptosis is not restricted to species or apoptotic stimulus. It is possible, therefore, that the PPs may suppress apoptosis by acting on diverse signalling pathways. However, it seems more likely that nafenopin suppresses hepatocyte apoptosis elicited by each death stimulus by impinging on a core apoptotic mechanism.

Animals↗
PubMed1998

Cisplatin- and paclitaxel-induced apoptosis of ovarian carcinoma cells and the relationship between bax and bak up-regulation and the functional status of p53.

We investigated the roles of p53 and Bcl-2 homologues in the induction of apoptosis by cisplatin and paclitaxel in wild-type p53-expressing human ovarian carcinoma cells and cisplatin-resistant derivatives that have lost p53 function. Cisplatin induced apoptosis in parental A2780 but not in cisplatin-resistant A2780/cp70 cells, whereas paclitaxel induced apoptosis in both cell lines. Immunoprecipitation of p53 using antibodies specific for p53 conformation (pAb 1620 and pAb 240) showed that there were no relative changes in p53 conformation before and after cisplatin treatment in either cell line. A2780/cp70 cells have lost p53 function, yet they have wild-type p53 gene sequence. However, A2780/cp70 cells constitutively express more p53 in a form detected by pAb 240, an antibody that also detects mutant conformations of p53 that are transcriptionally inactive. There were no changes in levels of Bcl-2, Bcl-XL, or 24-kDa Bax over 72 hr after exposure to cisplatin or paclitaxel, but each agent led to up-regulation of Bak and 21-kDa Bax in A2780 cells. Paclitaxel, but not cisplatin, increased Bak and 21-kDa Bax levels in A2780/cp70 cells. These data suggest that apoptosis in A2780 and A2780/cp70 is associated with an increased level of Bak and 21 kDa Bax after drug-induced damage and that functional p53 may be required for this effect after cisplatin but not after paclitaxel.

Antineoplastic Agents↗
PubMed1997

Germinal center-derived signals act with Bcl-2 to decrease apoptosis and increase clonogenicity of drug-treated human B lymphoma cells.

Bcl-2 suppresses drug-induced apoptosis in vitro, although in many cases, this results only in a delayed onset of cell death. In vivo survival signals from the extracellular environment may also contribute to drug resistance and may act with Bcl-2 to promote long-term cell survival. Ligation of CD40 on B-lymphocytes in germinal centers (GCs) can suppress apoptosis induced by calcium ionophore or anti-IgM in vitro. We asked whether a combination of Bcl-2 expression and the provision of a culture environment that mimicked that of the GC [CD40 ligation and interleukin 4 (IL-4)] could increase the ability of B lymphoma cells to resist drug-induced apoptosis. A Burkitt lymphoma (BL) cell line transfected with either human bcl-2 (BL-bcl-2) or control plasmid (BL-Sv2) was used to examine the effects of Bcl-2 overexpression on the cellular response and long-term survival after treatment with the DNA-alkylating drug chlorambucil (CMB) in the presence or absence of CD40 ligation and IL-4. Administration of 20 microM CMB completely prevented cell proliferation. This was associated with an increase in p53 protein levels within 24 h, without an elevation in p21, Bax, or Mdm2 proteins. Analyses of cell cycle distribution and of cyclin B expression demonstrated that both cell lines arrested at G2/M, where they died. Fifty % of BL-Sv2 cells died within 2 days, whereas 50% cell death was not observed in the BL-bcl-2 cultures until 6 days had passed. Cross-linking of CD40 with a monoclonal antibody elevated Bcl-xL protein levels by 3 h and also provided a delay in CMB-induced death. Ninety-six h after the addition of 20 microM CMB, 78% of the BL-Sv2 cells were apoptotic, whereas ligation of CD40 on BL-Sv2 cells reduced the proportion of apoptotic cells to 38%. Overexpression of Bcl-2 (in BL-bcl-2 cells) reduced apoptosis to 41%. However, when the BL-bcl-2 cells were treated with CMB together with ligation of CD40, apoptosis was reduced further to only 17% at 96 h. The Bcl-2-mediated delay in the execution of CMB-induced apoptosis did not translate significantly to increased clonogenicity. In contrast, the provision of BL-Sv2 cells with an ability to interact with the adhesion molecule vascular cell adhesion molecule-1, CD40 ligation, and IL-4 significantly increased clonogenic survival, and this was improved in BL-bcl-2 cells exposed to these GC-derived signals. These data demonstrate that the kinetics of drug-induced apoptosis can be modulated by Bcl-2 as well as by IL-4, vascular cell adhesion molecule-1, and CD40 ligation, the latter possibly involving the function of Bcl-xL. That these factors appear to act together to enhance proliferative potential after DNA damage has important implications regarding the development of drug resistance in B-cell lymphomas and future strategies for improved chemotherapy.

Antineoplastic Agents, Alkylating↗
PubMed1997

Cell cycle specific induction of HL-60 cell differentiation and apoptosis by mycophenolic acid.

HL-60 cells undergo terminal differentiation and apoptosis in response to different types of sub-toxic and toxic perturbations respectively. The mechanism by which cells sense different amounts of perturbation to activate pathways that lead to the engagement of a relevant biological response is not known. The response of HL-60 cells to treatment with the immunosuppressant mycophenolic acid (MPA), a specific inhibitor of dGTP/GTP-synthesis, allowed quantitation of a metabolic perturbation which triggered a cellular response. 1.5 microM MPA induced 38% terminal differentiation to CD14 positive, early monocyte-like cells and 22% cell death by apoptosis, whereas 3 microM MPA induced 70% apoptosis but no differentiation. Despite the difference in biological outcomes, 72 h exposure to both 1.5 microM and 3 microM MPA caused a similar ( approximately 75%) depletion of total GTP levels. Cells synchronized by centrifugal elutriation were treated with MPA. Elutriated cells were overall less sensitive to the effects of MPA but 3 microM MPA induced significantly less apoptosis and more differentiation in an elutriation-enriched G1-population than in a population normally distributed in the cell cycle, suggesting that the effects of MPA in S-phase may subsequently lead to cell death. However, analysis of apoptosis by using a terminal deoxynucleotidyltransferase assay and measurement of bromodeoxyuridine incorporation showed that apoptosis was engaged in G1. Analysis of the phosphorylation status of the retinoblastoma protein demonstrated that Rb was hypophosphorylated prior to apoptosis and that in apoptotic cells, separated by flow cytometry, Rb protein was absent, presumably due to proteolysis. The loss of Rb protein did not appear to permit transit to S-phase, and was not accompanied by an expression of c-Myc. Surprisingly, therefore, an antimetabolite inducing a loss of GTP brought about cell death by apoptosis in the G1 phase of the cell cycle.

Journal Article↗
PubMed1997

v-Abl protein tyrosine kinase (PTK) mediated suppression of apoptosis is associated with the up-regulation of Bcl-XL.

We demonstrated previously that the activation of v-Abl protein tyrosine kinase (PTK) in IC.DP murine pre-mast cells resulted in suppression of apoptosis after withdrawal of interleukin 3 (IL-3), that protein kinase C (PKC) translocated to the nucleus 6 h after v-Abl PTK activation and that inhibition of PKC restored apoptosis after IL-3 deprivation in the presence of v-Abl PTK activity. Here we demonstrate that v-Abl PTK activation is followed by an approximately twofold increase in mRNA level of Bcl-XL by 6 h and a corresponding increase in Bcl-XL protein level by 24 h. Bcl-xL RNA and protein decreased in IL-3 deprived cells in the absence of v-Abl PTK activity. Exposure of cells with v-Abl PTK active to the PKC inhbitor calphostin C (125 ng/ml) prevented the increase in Bcl-xL protein and resulted in apoptosis. No changes in Bax or Bcl-2 protein level were noted after IL-3 withdrawal and/or activation of v-Abl PTK. Bak was barely detectable and Bad protein level decreased in cells undergoing apoptosis. The data suggest that suppression of apoptosis by v-Abl PTK in the absence of IL-3 is associated with PKC signalling and the upregulation of Bcl-xL in IC.DP cells.

Animals↗
PubMed1997

Role of acid/base homeostasis in the suppression of apoptosis in haemopoietic cells by v-Abl protein tyrosine kinase.

Removal of interleukin-3 from murine IC.DP pre-mast cells results in irreversible commitment to apoptosis within 18 hours. To identify early events necessary for the engagement of apoptosis we examined the regulation of intracellular pH (pH(i)). IC.DP cells acidified 2 hours after removal of interleukin-3 (before discernible signs of apoptosis) and by 18 hours pH(i) had decreased by 0.15 units. The acidification was due to both an increase in an acid-loading process which only occurs when intracellular pH is above 6.8 and a slight reduction in H+ efflux via NA+/H+ exchange. Activation of a temperature sensitive mutant of v-Abl protein tyrosine kinase suppressed apoptosis of IC.DP cells in the absence of interleukin-3 but did not stimulate proliferation, and moreover prevented cellular acidification. Acidification of the cells by 0.2 units to pH 6.86 by complete inhibition of Na+/H+ exchange by 10 microM 5'-(N-methyl-N-isobutyl)-amiloride prevented the suppression of apoptosis by v-abl protein tyrosine kinase following IL 3 withdrawal. However in the presence of interleukin-3, addition of 10 microM 5'-(N-methyl-N-isobutyl)-amiloride only resulted in a fall of pH(i) to 7.17. Apoptosis did not occur and the cells continued to proliferate. Thus, in this model intracellular pH must fall below a critical value for apoptosis to occur. Together these data point to a step in cytokine deprivation induced apoptosis (at least in some haemopoietic cell types) which is either enhanced by or dependent upon an acidic intracellular environment which is the result of an increase in acid loading and inhibition of Na+/H+ exchange activity. One of the mechanisms by which activation of v-Abl protein tyrosine kinase suppresses apoptosis is by prevention of intracellular acidification.

Acid-Base Equilibrium↗
PubMed1997

Avoidance of apoptosis as a mechanism of drug resistance.

Inherent or acquired drug resistance is a major obstacle for the successful treatment of cancers. Many mechanisms of drug resistance have been described including a decreased drug uptake, an increase in DNA damage repair, enhanced drug detoxification, an altered level or mutation of the intracellular drug target or an increased drug efflux from the cell. Most of these mechanisms impinge upon the interaction of a drug with its cellular target or immediate consequences of such as interaction. For example, a decrease in the cellular levels of topoisomerase II thwarts the efficacy of certain topoisomerase II inhibitors, and enhanced levels of glutathione increase resistance to DNA alkylating agents. However, some tumours are inherently resistant to all chemotherapeutic agents, i.e. with different mechanisms of action. What is the mechanism(s) underlying this pleiotropic drug resistance? One possibility is that such drug-resistant tumour cells have an abnormally high threshold for the engagement of apoptosis (programmed cell death). The suppression of apoptosis as a mechanism for drug resistance is discussed in this article.

Apoptosis↗
PubMed1996

Characterization of cell volume loss in CEM-C7A cells during dexamethasone-induced apoptosis.

A reduction in cell volume is a fundamental feature of apoptosis. We have characterized changes in cell volume, together with nuclear changes, occurring in dexamethasone-induced apoptosis in CEM-C7A lymphoblastoid cells. Cell volume was measured by electronic cell sizing and flow cytometry, and two distinct phases of volume loss were observed. The first phase began 12 h after addition of dexamethasone (5 microM) and progressed until 36 h when chromatin condensation was detected in intact cells. Removal of dexamethasone before 36 h (the precommitment period) resulted in reversal of the volume decrease and prevented the appearance of nuclear changes. Cell shrinkage in the first 24 h of dexamethasone exposure was associated with a net loss of potassium but no change in cellular buoyant density. There were no significant differences in the rates of volume recovery after either hypertonic or hypotonic stimuli. These observations favor a mechanism of cell shrinkage involving loss of the entire cytoplasmic contents, possibly following proteolysis, rather than loss of only osmolytes and water. The second phase of volume loss was coincident with chromatin condensation and was associated with cellular fragmentation and a reduction in cellular density. We conclude that volume loss in this model of apoptosis is mediated by multiple mechanisms that are both dependent and independent of cellular fragmentation.

Apoptosis↗
PubMed1996

Requirement of basement membrane for the suppression of programmed cell death in mammary epithelium.

Apoptosis is an active mechanism of cell death required for normal tissue homeostasis. Cells require survival signals to avoid the engagement of apoptosis. In the mammary gland, secretory epithelial cells are removed by apoptosis during involution. This cell loss coincides with matrix metalloproteinase activation and basement membrane degradation. In this paper we describe studies that confer a new role for basement membrane in the regulation of cell phenotype. We demonstrate that the first passage epithelial cells isolated from pregnant mouse mammary gland die by apoptosis in culture, but that cell death is suppressed by basement membrane. The correct type of extracellular matrix was required, since only a basement membrane, not plastic or a collagen I matrix, lowered the rate of apoptosis. Attachment to a matrix per se was not sufficient for survival, since apoptotic cells were observed when still attached to a collagen I substratum. Experiments with individually isolated cells confirmed the requirement of basement membrane for survival, and demonstrated that survival is enhanced by cell-cell contact. A function-blocking anti-beta1 integrin antibody doubled the rate of apoptosis in single cells cultured with basement membrane, indicating that integrin-mediated signals contributed to survival. We examined the cell death-associated genes bcl-2 and bax in mammary epithelia, and found that although the expression of Bcl-2 did not correlate with cell survival, increased levels of Bax were associated with apoptosis. We propose that basement membrane provides a survival stimulus for epithelial cells in vivo, and that loss of interaction with this type of matrix acts as a control point for cell deletions that occur at specific times during development, such as in mammary gland involution.

Animals↗
PubMed1996

DNA-damaging agents induce both p53-dependent and p53-independent apoptosis in immature thymocytes.

Apoptosis in the immature thymus can be induced by both p53-dependent and -independent pathways, the former being activated by exposure to DNA-damaging agents and the latter being induced by glucocorticoids [Nature (Lond.) 362:847-849; Nature (Lond.) 362:849-852 (1993)]. We report that the DNA-damaging agents etoposide and gamma-radiation induced similar levels of apoptosis in both proliferatively enriched and quiescent immature rat thymocytes, as assessed by flow cytometry and the formation of both kilobase-pair and 180-bp integer fragments of DNA. However, a marked stabilization of p53 occurred exclusively in the proliferatively enriched population, which was also enriched for immature CD4- CD8- and mature CD4+ CD8-/CD4- CD8+ cells. In contrast, DNA damage-induced apoptosis in quiescent mature peripheral T cells was associated with an accumulation of p53. Our studies suggest that stabilization of p53 in thymocytes in response to DNA damage may be developmentally regulated. In immature thymocytes obtained from p53-null mice, DNA-damaging agents induced apoptosis at significantly lower levels and at later times than that seen in cells from p53 wild-type animals. These data support the hypothesis that DNA-damaging agents induce apoptosis primarily via a p53-dependent pathway in immature thymocytes as previously reported. We report here that DNA damage can also induce apoptosis by a p53-independent pathway in a particular subpopulation of immature thymocytes.

Amino Acid Sequence↗
PubMed1996

Cisplatin, camptothecin, and taxol sensitivities of cells with p53-associated multidrug resistance.

With the use of clonogenic survival assays, we show that wild-type p53-expressing A2780 human ovarian cell lines transfected with a dominant negative mutant p53 gene (codon 143, valine to alanine) acquired cross-resistance to ionizing radiation, cisplatin, doxorubicin, and 1-beta-D-arabinofuranosylcytosine. However, these mutant p53-transfected cell lines retained sensitivity to taxol and camptothecin. We also show that immature thymocytes from mice with the p53 gene genetically inactivated showed reduced ability to undergo apoptosis after treatment with ionizing radiation and cisplatin compared with wild-type mice. However, taxol-induced apoptosis in thymocytes does not seem to be dependent on p53 status. Camptothecin also induced apoptosis in a p53-independent manner in thymocytes at low doses but in a p53-dependent manner at high doses. These data suggest that taxoids and camptothecin analogs could have activity in tumors that have aberrant p53 function and provide a rationale for the clinical observations of responsiveness of refractory ovarian cancer to these drugs.

Animals↗
PubMed1995

Further characterisation of the in situ terminal deoxynucleotidyl transferase (TdT) assay for the flow cytometric analysis of apoptosis in drug resistant and drug sensitive leukaemic cells.

Apoptosis, originally defined by specific morphological changes, is characterised biochemically by non-random cleavage of DNA. Depending on cell type, this DNA cleavage proceeds from 300 and 50kbp fragments prior to, concomitantly with, or in the absence of 180bp integer fragmentation. Incorporation into fragmented DNA of biotin-labelled nucleotides by terminal deoxynucleotidyl transferase (TdT) has recently become a standard flow cytometric assay for the identification and quantitation of apoptosis. Nucleotide incorporation is visualized using avidin-tagged fluorescein isothiocyanate (FITC) (Gorczyca et al.: Cancer Res 53:1945-1951, 1993; Jonker et al.: Cytometry (Suppl 13):Abstr 99A, 1993). Here, we characterise this assay further in three different haemopoietic cell lines. Drug-induced DNA damage is not identified by the TdT assay unless it is coupled to the apoptotic response. This was demonstrated using cells in which activation of the oncogenic Abelson-encoded protein tyrosine kinase suppressed drug-induced apoptosis, but did not inhibit drug-induced DNA damage (by melphalan, hydroxyurea, or etoposide). Furthermore, the TdT assay identifies DNA fragments formed during apoptosis induced by etoposide and N-methylformamide in HL60 and MOLT-4 cells, including those high molecular weight DNA fragments formed in MOLT-4 cells which were not further cleaved to 180-200bp integer fragments. Our results support the use of flow cytometry and the TdT assay to reliably measure apoptotic cells in heterogeneous cell samples.

Abelson murine leukemia virus↗
PubMed1995

MCF-7 human mammary adenocarcinoma cell death in vitro in response to hormone-withdrawal and DNA damage.

Anti-oestrogens exert a tumoristatic effect on estrogen-receptor-positive breast carcinomas in vivo. At a cellular level this may reflect inhibition of cell proliferation and/or cell death counterbalanced by continued proliferation of a cell subpopulation. We evaluated the MCF-7 human mammary adenocarcinoma cell line as an in vitro model to study the effects of the novel oestrogen antagonist ICI 182,720 on cell population dynamics (cell gain vs. cell loss). After oestrogen-withdrawal monolayer cell number declined over 10 days, accompanied by cell detachment. This decrease in viable cell number was elevated 2-fold by ICI 182,780. Detached cells exhibited DNA fragments of 50 and 300 kbp, typical of apoptotic cells. However, internucleosomal cleavage to 180 bp integer fragments was not seen, and these detached cells exhibited a morphology which was not consistent with apoptosis. The remaining attached monolayer cells were morphologically viable (> 99%) with regard to both nuclear morphology and plasma membrane integrity. There was no difference in cell cycle phase distribution between oestrogen-withdrawn and ICI 182,780-treated cells; both induced accumulation in G1 phase. MCF-7 cells were also exposed to a variety of DNA damaging agents known to induce apoptosis in other cell types. We could demonstrate only limited induction of morphologically recognisable apoptosis in MCF-7 cells treated with methyl methanesulphonate. Our results add to the controversy surrounding the ability of the MCF-7 cell line to undergo apoptosis in vitro in response to anti-oestrogen therapies.

Adenocarcinoma↗