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N Kyprianou

Publications and source records attributed to N Kyprianou.

At least 73 records · Page 4Linked to original sources
PubMed1990

Effect of transforming growth factor-beta 1 on proliferation and death of rat prostatic cells.

The ability of transforming growth factor B1 (TGF beta 1) to inhibit proliferation and activate death of rat ventral prostatic glandular cells was tested both in vivo and in vitro. In vivo administration of 50 ng TGF beta 1/day directly to the regressed ventral prostate of previously castrated male rats had no effect on the proliferative regrowth of the prostatic glandular cells induced by exogeneous androgen replacement. In addition, androgen-stimulated ventral prostatic cell proliferation in vitro in organ culture was not affected by exposure to 0.1-20 ng/ml TGF beta 1. In contrast in vivo administration of 50 ng TGF beta 1/day directly to the ventral prostate of intact noncastrated male rats resulted in the death of about 25% of the prostatic glandular cells within 7 days of treatment. Such TGF beta 1 treatment did not lower serum testosterone, nor did it affect the size or DNA content of the seminal vesicles, demonstrating the local nature of the response. Likewise, in androgen-maintained ventral prostate organ cultures in vitro, there was a dose-response relationship between glandular cell death and TGF beta 1 concentration in the medium. These results demonstrate that TGF beta 1 can induce the death of androgen-dependent prostatic glandular cells even when physiological levels of androgen are present. Previous studies have demonstrated that both the receptor and the mRNA for TGF beta 1 increase rapidly in the ventral prostate after castration. Taken with the present data, these results suggest that TGF beta 1 may be a physiological intermediate in the programmed cell death of rat prostatic glandular cells activated after androgen ablation.

Animals↗
PubMed1989

"Thymineless" death in androgen-independent prostatic cancer cells.

The molecular mechanism of "thymineless" death induced by 5-fluorodeoxyuridine or trifluorothymidine, in androgen-independent rat prostatic adenocarcinoma AT-3 cells was investigated. Fragmentation of genomic DNA into discrete multiples of a nucleosomal unit (i.e. 180bp subunit) and induction of expression of TRPM-2, a programmed cell death-associated gene, temporally correlated with the activation of programmed cell death in this system. In contrast, killing of AT-3 cells by osmotic lysis, or membrane-targeted metabolic inhibitors results in neither the stereotypic DNA fragmentation into nucleosomal oligomers nor the elevation of TRPM-2 mRNA levels but to non-specific biochemical changes characteristic of necrosis. These results suggest that androgen-independent prostatic cancer cells retain a major portion of the programmed cell death cascade which can be activated by non-androgen ablative cytotoxic drugs that induce "thymineless" death.

Animals↗
PubMed1989

Relationship between DNA fragmentation and apoptosis in the programmed cell death in the rat prostate following castration.

Previous studies have demonstrated that the rapid involution of the rat ventral prostate following castration involves the death of the androgen-dependent epithelial cells present within the gland and that this death is the result of a series of discrete biochemical steps. The degradation of genomic DNA into nucleosomal-sized fragments is an early event in this process and is catalyzed by calcium magnesium-dependent endonuclease activity. The morphologic correlation of the involution process involves a series of structural changes which are collectively referred to as apoptosis. The apoptotic process describes the earliest apparent signs of morphologic change exhibited by the dying cells through their eventual complete destruction and deletion from the tissue. The temporal relationship between these recently described biochemical events and the morphologic changes of the apoptotic process were compared in the present study, in order to test the cause versus effect nature of DNA fragmentation in the programmed death of androgen dependent prostatic cells following castration. These studies demonstrated that the early elevation of the Ca+2 Mg+2-dependent endonuclease activity and the fragmentation of DNA into nucleosomal oligomers occurs within prostatic glandular epithelial cells and probably does not involve the direct participation of extraprostatic cells which may subsequently migrate into the gland. Once the DNA is initially cleaved into the nucleosomal oligomers, the subsequent participation of lysosomal enzymes act in a less restricted fashion to degrade both the nucleosomal DNA as well as the cytoplasmic elements and the cell becomes morphologically apoptotic. As the elevations in Ca+2 Mg+2-dependent endonuclease activity and DNA fragmentation are initiated at a time well before the cell is morphologically dead, as defined by apoptosis, these changes in DNA metabolism must not be the consequences of cell death but instead are early causal events in an active process of programmed cell death.

Animals↗
PubMed1989

Expression of transforming growth factor-beta in the rat ventral prostate during castration-induced programmed cell death.

Castration-induced androgen deprivation leads to the activation of the programmed death of the androgen-dependent prostatic epithelial cells in the rat ventral prostate. In order to identify potential mediators of this programmed cell death, the expression of transforming growth factor-beta (TGF beta) in the rat ventral prostate was studied, after castration induced-androgen withdrawal. Steady state levels of TGF beta mRNA were determined by Northern blot analysis and compared with mRNA levels for prostatein C3, the major androgen-dependent secretory protein of ventral prostate and also with mRNA levels for TRPM-2, a gene that is specifically expressed during castration induced prostatic cell death. Within the first day after castration there was a dramatic increase in the levels of TGF beta mRNA in the ventral prostate (approximately 10-fold) and by 4 days after castration TGF beta mRNA was maximally expressed (approximately 40-fold increase), by which time the androgen-dependent C3 secretory protein mRNA transcripts have diminished to undetectable levels. Androgen administration to 4-day castrated rats led to a marked decrease in TGF beta mRNA to a level comparable to its constitutive expression obtained in the intact control animals, indicating that expression of TGF beta in the rat ventral prostate is under negative androgenic regulation. The transcript levels encoding TRPM-2 initially increased 10-fold within the first day after castration and by day 4 post castration there was a dramatic increase (approximately 50-fold) which correlated well with the maximal rate of cell death of the androgen-dependent prostatic epithelial cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Androgens↗
PubMed1988

Activation of a Ca2+-Mg2+-dependent endonuclease as an early event in castration-induced prostatic cell death.

Previous studies have demonstrated that castration-induced androgen withdrawal results in the fragmentation of prostatic DNA into nucleosomal oligomers, and this process comprises an early event in the activation of programed cell death in the rat ventral prostate. This DNA fragmentation could be due to changes in the chromatin conformation increasing its sensitivity to preexisting nucleases and/or to increases in the activity of the nucleases themselves. However, comparative kinetic analysis of in vitro DNA fragmentation induced by exogenous nucleases did not reveal any differences in the sensitivity of prostatic chromatin between intact and castrated rats. In contrast to these negative findings, using [3H] DNA as an exogenous substrate, it was shown that within the first day following castration there was a twofold increase in a Ca2+-Mg2+-dependent nuclease activity without a concomitant increase in other nuclear nucleases. This Ca2+-Mg2+-dependent nuclease activation occurred coincidental with the initial increase in nuclear DNA fragmentation following castration and preceded the enhanced appearance of morphological changes characteristic of dying cells (i.e., apoptosis), as well as the major increase in prostatic DNA loss. These results suggest that castration-induced androgen deprivation leads to a sequential activation of a Ca2+-Mg2+-dependent nuclease leading to the fragmentation of the genome into discrete nucleosomal-sized fragments of DNA, subsequently followed by the fragmentation of the nucleus itself (i.e., apoptosis) and eventually with the complete digestion of the nucleosomal oligomers into component nucleotides (i.e., DNA loss). Since the castration-induced nuclease is dependent upon calcium ions for maximal activity, a potential role of intracellular calcium in the early events activating prostatic cell death was investigated. Acute disturbances in intracellular calcium homeostasis within the ventral prostate by means of a potent calcium influx blocker, nifedipine, simultaneous with castration, resulted in a significant delay in the biochemical and morphological changes associated with prostatic cell death (i.e., prostatic weight loss, prostatic DNA loss, and DNA fragmentation). These results point to a potential role of intracellular calcium levels in the mechanism of activation of castration-induced death of the androgen-dependent epithelial cells in the ventral prostate.

Animals↗
PubMed1988

Human hepatocellular carcinoma tumor xenografts. Their androgen-receptor status and growth responses to castration.

Castrated or sham-operated male athymic mice were inoculated with cells from the human hepatocellular carcinoma cell line PLC/PRF/5. There were no significant differences between the two groups with respect to the number of animals developing tumors, the time to tumor development, or the subsequent rate of increase in either tumor base area or mouse serum alpha-fetoprotein concentration. Androgen receptors were assayed in nuclei obtained from three separate liver cancer cell lines and from normal adult human liver. Similar concentrations, ranging from 235 to 550 fmol/mg DNA, of nuclear androgen receptors were detected in all tissues. Low percentages of androgen receptors were retained on DNA-cellulose. Although the presence of receptors implies the potential for metabolic effects of androgens in normal and malignant liver, our in vivo studies suggest that castration does not alter significantly the growth of liver cancer xenografts in athymic mice.

Animals↗
PubMed1988

Activation of programmed cell death in the rat ventral prostate after castration.

The rapid involution of the rat ventral prostate after castration is an active process initiated by removal of the inhibitory effects of androgen on prostatic cell death. The present studies demonstrate that after castration-induced androgen deprivation a series of temporally discrete biochemical events are activated which result in the rapid programmed death of the subset of androgen-dependent cells within the rat ventral prostate. These biochemical steps involve 1) rapid loss of nuclear androgen receptor retention; by 12 h after castration, androgen receptors are no longer detectable in ventral prostatic nuclei; 2) an initial fragmentation of nuclear DNA into low mol wt (less than 1000 basepairs) nucleosomal oligomers which lack intranucleosomal break points; and 3) eventual complete digestion of these nucleosomal oligomers into component nucleotides. Additional studies demonstrate that activation of a Ca2+-Mg2+-dependent endonuclease is associated with this DNA fragmentation. By 4 days after castration, maximal DNA fragmentation is obtained, with 15% of the total nuclear DNA extractable as low mol wt fragments. Proteolytic enzymes are apparently not involved initially in this process, suggesting that DNA fragmentation is a discrete event in, rather than a result of, cell death. Flow cytometric analysis of nuclear DNA content demonstrated that each day after castration, a subpopulation of androgen-dependent cells in rat ventral prostate fragmented all of their genomic DNA, as opposed to the whole population of cells fragmenting an increasing portion of their DNA daily.

Animals↗
PubMed1988

Identification of a cellular receptor for transforming growth factor-beta in rat ventral prostate and its negative regulation by androgens.

Scatchard analyses of the binding of transforming growth factor-beta (TGF beta) to membranes from rat ventral prostate revealed the presence of high affinity (Kd = 140 pM) saturable binding sites for [125I]TGF beta. The binding of [125I]TGF beta to prostatic membranes, while displaced in the presence of excess unlabeled TGF beta, is unaffected by epidermal growth factor, nerve growth factor, fibroblast growth factor, or insulin, indicating the specificity of binding. Affinity labeling of these membrane receptors by covalent attachment to [125I]TGF beta with bis-(sulfosuccinimidyl)suberate and subsequent electrophoretic analysis of the labeled complexes revealed the specific binding of [125I]TGF beta to a macromolecule that predominantly migrates as a 260,000 mol wt band in 7.5% acrylamide gels. Castration-induced androgen deprivation produced a significant increase in [125I]TGF beta binding to prostatic membranes with no apparent change in the affinity of membrane receptors for TGF beta. TGF beta receptor levels per total gland increased approximately 2-fold by 3 days after castration, reached a peak value by day 4, and then declined during the subsequent 10 days. Androgen administration to 4-day castrated animals decreased the number of TGF beta receptors to a value similar to that in the intact controls. These results demonstrate the presence of specific binding sites for TGF beta in the rat ventral prostate. Furthermore, the TGF beta receptor levels seem to be under negative androgenic regulation, indicating a potential role for this growth factor in the mechanism of activation of castration-induced death of androgen-dependent epithelial cells in the ventral prostate.

Androgens↗
PubMed1987

Biological significance of measurable androgen levels in the rat ventral prostate following castration.

Within 12 hr after castration, there is a dramatic drop in the serum testosterone (T) levels to approximately 1.3% of the intact value (2.5 +/- 0.8 ng/ml). By 1 day following castration, the serum T levels are approximately 3.3% of the intact control level. In contrast, serum 5 alpha-dihydrotestosterone (DHT) levels decrease to only 50% of the intact value within 12 hr postcastration and remain at a value greater than 50% of the intact control level even following long-term castration for up to 20 weeks. Following castration, tissue T and DHT concentrations in rat ventral prostate (RVP) exhibited a similar sequence of changes. Within 12 hr after castration, there is a substantial decrease in T to 27% and DHT to 20% of their intact values; after a further transient decrease during the subsequent 7 days, these levels remain constant with RVP at approximately 40% for T and 20% for DHT of the intact control levels even following long-term castration. Thus castration induces only a partial withdrawal of the tissue androgens. The low but measurable androgen levels in RVP of castrated host are of adrenal origin, since following surgical adrenalectomy these remaining androgen levels become undetectable. Thus castration plus adrenalectomy produces a complete androgen withdrawal within the RVP. To determine the biological significance of the measurable androgen levels remaining following castration, the RVP cell number and the rate of prostatic DNA synthesis were compared in RVP following castration alone (ie, partial androgen withdrawal) or castration combined with surgical adrenalectomy (ie, complete androgen withdrawal). These results demonstrated that complete elimination of the remaining androgens in the RVP of long-term castrates, by means of surgical adrenalectomy, did not induce any further reduction in either of these prostatic growth parameters. Therefore, in the rat, DHT must be decreased to a critical threshold but does not have to be completely eliminated to decrease maximally androgen effect on the prostate.

Adrenal Glands↗
PubMed1987

Quantal relationship between prostatic dihydrotestosterone and prostatic cell content: critical threshold concept.

The shape of the prostatic dihydrotestosterone (DHT) dose vs prostatic cell number response curve can be used to determine if the androgen-induced increase in prostatic cell number occurs as a continuous graded process increasing with all levels of prostatic DHT, or whether the process is quantal, beginning to occur only when a critical threshold of prostatic DHT is reached. To make this determination, castrated-adrenalectomized rats, castrated rats given no treatment, and castrated rats given testosterone-filled silastic implants of varying lengths were used to construct such a prostatic DHT dose/prostatic cell number response curve ranging from undetectable to pharmacologically high levels of prostatic DHT. The shape of this dose-response curve was found not to be continuously hyperbolic, but instead, to be discontinuously sigmoidal. This demonstrates that, in the rat, androgen-induced increase in prostatic cell number occurs as a quantal process which can only begin when the concentration of prostatic DHT is above a critical threshold value (ie, 0.4 ng/10(8) cells for the rat ventral prostate). Thus, in order to completely prevent this androgenic stimulation of prostatic cell number, the prostatic DHT level has to be lowered to below this critical threshold but does not have to be completely eliminated.

Adrenalectomy↗
PubMed1987

Development of androgen-independent tumor cells and their implication for the treatment of prostatic cancer.

Development of androgen-independent prostatic cancer cells from androgen-responsive cells can occur by a variety of mechanisms (e.g., environmental adaptation, multifocal origin, or genetic instability). Regardless of the mechanism of development, however, once androgen-independent cancer cells become present within prostatic cancer, the tumor is no longer homogeneous but is now heterogeneous. Once a prostatic cancer is heterogeneously composed of both androgen-dependent and -independent cancer cells, androgen withdrawal therapy, no matter how complete, cannot be curative. In order to produce cures of such heterogeneous prostatic cancers, hormonal therapy must be combined simultaneously with chemotherapy early in the course of the disease so that all the cancer populations (i.e., androgen-dependent and -independent) can be simultaneously affected within an individual patient.

Androgen Antagonists↗
PubMed1987

Intranuclear distribution of androgen receptors in human prostate carcinoma.

Androgen receptors in nuclei from human prostate carcinomas were characterized on the basis of their solubilization by, or resistance to, micrococcal nuclease. By this means, androgen receptors were assigned to three nuclear categories: those associated with nuclease-resistant structures, those associated with chromatin and those apparently uncommitted by association with either of these. Prostate carcinoma nuclei contained high concentrations (57-82% of total nuclear content) of nuclease-resistant androgen receptors. This was a different pattern from that observed previously for benign hypertrophic prostate epithelial nuclei which contained a variable high proportion of uncommitted androgen receptors. The differences could not be attributed to differential losses to cytosol, or to loss of functionality, as determined in vitro. The differences in distribution could reflect different responses of diseased cells to androgens, or the intervention of other factors more relevant to the disease process.

Animals↗
PubMed1986

Association states of androgen receptors in nuclei of human benign hypertrophic prostate.

Androgen receptors (AR) were quantified in nuclei purified from unfractionated benign hypertrophic prostate (bph) tissue and from separated epithelium and stroma from bph specimens. Both epithelial and stromal cell nuclei contained AR, although concentrations in epithelial cell nuclei were higher and more variable. Variations in AR levels in epithelial cell nuclei reflected variations in unfractionated-tissue nuclei. Nuclear AR were further characterized regarding extractability with or resistance to 0.6 mol/lKCl and micrococcal nuclease. Nuclei from unfractionated tissue, epithelium, and stroma contained populations of AR susceptible and refractory to solubilization with KC1 and nuclease. Nuclease- and salt-sensitive populations of AR were similar numerically. The observed variability in epithelial cell nuclear AR was attributable to a wide range of solubilizable AR. Nuclease-digestion profiles and sedimentation analyses revealed that this wide range was not due to AR associated with soluble chromatin oligomers but to AR not detectably associated with other nuclear components. In contrast, AR in stromal cell nuclei was predominantly resistant to KC1 and nuclease, and variability in total nuclear AR concentration was due to variation in the nonextractable population.

Cell Fractionation↗
PubMed1986

Evaluation of biopsy techniques for androgen receptor assay in human prostatic tissue.

Androgen receptors were assayed in nuclei prepared from human benign hypertrophic prostate tissue obtained at retropubic prostatectomy by transurethral resection (TURP) and by cold punch resection. Androgen receptor was not detected in 70% of the transurethrally resected samples, and in those cases where receptors were observed they showed no correlation with enucleated samples. However, androgen receptors were detected in all cold punch resected samples and enucleated tissue with similar concentrations evident in samples from the same gland. These results suggest that prostate specimens collected by TURP are not suitable for androgen receptor analysis, whereas the technique of cold punch resection yields prostate samples that are more reliable for a correct evaluation of androgen receptor content.

Aged↗
PubMedDate not supplied

Potent in vitro anticancer activities of ring-expanded ("fat") nucleosides containing the imidazo[4,5-e][1,3]diazepine ring system.

The ring-expanded ("fat") nucleoside, 4,8-diamino-6-imino-6H-1-beta-D-ribofuranosylimidazo[4,5-e][1,3]diazepine (1) and its 2',3',5'-tri-O-benzoyl derivative (2) exhibited potent broad spectrum anticancer activities in vitro against a wide variety of human tumor cell lines. The tribenzoyl derivative 2 was found to be considerably more active than the parent nucleoside 1. Further studies using human prostate cancer cells PC-3 and DU-145 suggest that the treatment of exponentially growing culture cells with 1 and 2 leads to marked loss of cell viability in a dose-dependent manner.

Antineoplastic Agents↗
PubMedDate not supplied

Radiation-induced apoptosis: predictive and therapeutic significance in radiotherapy of prostate cancer (review).

Current therapy for advanced prostate cancer is hampered by the propensity of the disease to progress from an androgen-dependent state to an androgen-independent state. Current treatment for advanced disease is palliative. Therefore, the therapeutic goal for prostate cancer treatment today is to arrest the disease at an early state when it is still localized to the gland. The standard treatment for clinically localized disease is radical prostatectomy or radiation therapy by way of external beam irradiation or local radioactive seed implants (brachytherapy). In advanced disease, the use of radiation therapy is limited to palliation of pain secondary to bone metastases and for spinal cord compression. Tracking residual disease and predicting outcome is limited to following the level of prostate specific antigen (PSA) production, evaluating for bone or solid organ metastasis, and analyzing their preoperative clinical stage, PSA and Gleason's score. Apoptosis as a molecular process of genetically regulated cell death has a critical endpoint that coincides with the goal of successful treatment of human malignancies. Since in cancer treatment the therapeutic goal is to trigger tumor-selective cell death, activation of the apoptotic pathway in prostatic tumor cells offers attractive and potentially effective therapeutic targets. As our understanding of the vital role of apoptosis in the development and growth of the prostate gland has expanded, numerous genes that encode apoptotic regulators have been identified that are severely impaired in prostate tumors. Human prostate cancer cells undergo apoptosis in response to androgen ablation, chemotherapeutic agents and ionizing irradiation. The expression of apoptotic modulators within individual prostate tumors appears to correlate with the cancer cell's sensitivity to traditional therapeutic modalities, including radiotherapy. No strict correlation between radiation-induced apoptosis and longevity of prostate cancer patients has emerged, possibly because the ability to achieve an initial remission alone does not adequately predict long-term outcome and patient survival. In this review we summarize the current understanding of the effects of radiation therapy on prostatic tumor cells within the context of the therapeutic significance of radiation-induced apoptosis in the effective elimination of androgen independent prostate cancer cells. As we enter a new millenium, identification of distinct molecular markers predictive of therapeutic response of prostatic tumors to radiation therapy may afford alternative prognostic indicators in optimizing our treatment protocols for advanced disease.

Apoptosis↗
PubMedDate not supplied

Androgen regulation of programmed death of normal and malignant prostatic cells.

Androgen-dependent normal prostatic glandular cells and androgen-dependent prostatic cancer cells can be induced to undergo cell death after androgen ablation. This death does not require the cells to proliferate and occurs as an energy-dependent process collectively referred to as "programmed cell death" in which the cells actively commit "suicide." Associated with this programmed cell death pathway is the enhanced expression of a series of genes and the fragmentation of the genomic DNA into nucleosomal oligomers. This genomic DNA fragmentation is the irreversible commitment step in the death of the cell and results from activation of Ca2+/Mg(2+)-dependent endonuclease activity within the cell nucleus. This activation is due to sustained elevation of intracellular free Ca2+ (Cai) induced after androgen ablation. Metastatic prostatic cancer within an individual patient is heterogeneous, including both androgen-dependent and -independent cancer cells. Thus, androgen ablation is rarely curative since it only induces the programmed death of the androgen-dependent cancer cells without activating this pathway in the androgen-independent cancer cells within the patient. Androgen-independent prostatic cancer cells do not activate this death process after androgen ablation, since this does not induce a sustained increase in Cai. A new approach to treat androgen-independent prostatic cancer cells has focused on the use of chemotherapeutic agents to induce a sustained increase in Cai. These studies demonstrate that if such a sustained elevation in Cai is maintained, even androgen-independent prostatic cancer cells undergo programmed cell death.

Androgens↗