RESUMEN
OBJECTIVE: To evaluate the short-term psychological consequences of gestational trophoblastic disease (GTD). DESIGN: A prospective observational multicentre cohort study. SETTING: Nationwide in the Netherlands. POPULATION: GTD patients. METHODS: Online questionnaires directly after diagnosis. MAIN OUTCOME MEASURES: Hospital Anxiety and Depression Scale (HADS), Distress Thermometer (DT), Impact of Event Scale (IES) and Reproductive Concerns Scale (RCS). RESULTS: Sixty GTD patients were included between 2017 and 2020. Anxious feelings (47%) were more commonly expressed than depressive feelings (27%). Patients experienced moderate to severe adaptation problems in 88%. Patients who already had children were less concerned about their reproductivity than were patients without children (mean score 10.4 versus 15.0, P = 0.031), and patients with children experienced lower distress levels (IES mean score 25.7 versus 34.7, P = 0.020). In addition, patients with previous pregnancy loss scored lower for distress compared with patients without pregnancy loss (IES mean score 21.1 versus 34.2, P = 0.002). DISCUSSION: We recommend that physicians monitor physical complaints and the course of psychological wellbeing over time in order to provide personalised supportive care in time for patients who have high levels of distress at baseline. CONCLUSIONS: GTD patients experience increased levels of distress, anxiety and depression, suggesting the diagnosis has a substantial effect on the psychological wellbeing of patients. The impact of GTD diagnosis on intrusion and avoidance seems to be ameliorated in patients who have children or who have experienced previous pregnancy loss. TWEETABLE ABSTRACT: Patients with gestational trophoblastic disease (GTD) experience short-term psychological consequences such as distress, anxiety and depression, suggesting that the diagnosis GTD has a substantial effect on the psychological wellbeing of patients. Various patient characteristics affect the impact of GTD diagnosis.
Asunto(s)
Ansiedad/psicología , Depresión/psicología , Enfermedad Trofoblástica Gestacional/psicología , Distrés Psicológico , Estrés Psicológico/psicología , Adulto , Ansiedad/etiología , Depresión/etiología , Femenino , Humanos , Masculino , Países Bajos , Embarazo , Estudios Prospectivos , Estrés Psicológico/etiología , Encuestas y CuestionariosRESUMEN
AIM: This study aimed to understand the role of miR-133a in progesterone actions, explore the regulative mechanism of the progesterone receptor, and investigate the effects of miR-133a on the progesterone-inhibited proliferation of mouse endometrial epithelial cells. METHODS: The expression of miR-133a induced by progesterone was detected by quantitative real-time PCR both in vivo and in vitro. Ishikawa subcell lines stably transfected with progesterone receptor subtypes were used to determine the receptor mechanism of progesterone inducing miR-133a. Specific miR-133a mimics or inhibitors were transfected into mouse uteri and primary cultured endometrial epithelial cells to overexpress or downregulate the miR-133a. The roles of miR-133a in the cell cycle and proliferation of endometrial epithelial cells were analysed by flow cytometry and Edu incorporation analysis. The protein levels of cyclinD2 in uterine tissue sections and primary cultured endometrial epithelial cells were determined by immunohistochemistry and Western blot analysis. RESULTS: Progesterone could induce miR-133a expression in a PRB-dependent manner in endometrial epithelial cells. miR-133a inhibited endometrial epithelial cell proliferation by arresting cell cycle at the G1 -S transition. Moreover, miR-133a acted as an inhibitor in downregulating cyclinD2 in endometrial epithelial cells. CONCLUSION: We showed for the first time that progesterone-induced miR-133a inhibited the proliferation of endometrial epithelial cells by downregulating cyclinD2. Our research indicated an important mechanism for progesterone inhibiting the proliferation of endometrial epithelial cells by inducing special miRNAs to inhibit positive regulatory proteins in the cell cycle.
Asunto(s)
Ciclina D2/biosíntesis , Endometrio/metabolismo , Regulación de la Expresión Génica/efectos de los fármacos , MicroARNs/metabolismo , Progesterona/farmacología , Animales , Western Blotting , Proliferación Celular/efectos de los fármacos , Proliferación Celular/fisiología , Células Epiteliales/metabolismo , Femenino , Citometría de Flujo , Inmunohistoquímica , Ratones , Reacción en Cadena en Tiempo Real de la PolimerasaRESUMEN
Under normal conditions, progesterone inhi-bits the estrogen-induced proliferation of endometrial epithelium. Our previous studies have shown that cyclin G1 was progesterone-dependent in mouse endometrial epithelium at peri-implantation, and exogenous cyclin G1 suppressed the proliferation of endometrial cancer cells. The objectives of this study are to determine whether cyclin G1, as a negative regulator of the cell cycle, is involved in the antiproliferative action of progesterone on endometrial epithelial cells, and to explore the possible molecular mechanism of cyclin G1 inhibition. The siRNA-mediated elimination of cyclin G1 attenuated the antiproliferative action of progesterone on endometrial epithelial cells. Immunoprecipitation showed that progesterone-induced cyclin G1 could interact with PP2A to mediate its phosphatase activity. The block of PP2A activity also attenuated the antiproliferative action of progesterone on endometrial epithelial cells and increased the phosphorylated Rb. In conclusion, progesterone-induced cyclin G1 mediates the inhibitory effect of progesterone on endometrial epithelial cell proliferation possibly through the recruitment of PP2A to dephosphorylate Rb.
Asunto(s)
Ciclina G1/metabolismo , Endometrio/citología , Células Epiteliales/metabolismo , Progesterona/farmacología , Animales , Proliferación Celular/efectos de los fármacos , Células Epiteliales/citología , Células Epiteliales/efectos de los fármacos , Femenino , Humanos , Ratones , Ácido Ocadaico/farmacología , Unión Proteica/efectos de los fármacos , Proteína Fosfatasa 2/metabolismo , ARN Interferente Pequeño/metabolismoRESUMEN
Tibolone, a tissue-selective compound with a combination of estrogenic, progestagenic, and androgenic properties, is used as an alternative for estrogen or estrogen plus progesterone hormone therapy for the treatment of symptoms associated with menopause and osteoporosis. The current study compares the endometrial gene expression profiles after short-term (21 days) treatment with tibolone to the profiles after treatment with estradiol-only (E(2)) and E(2) + medroxyprogesterone acetate (E(2) + MPA) in healthy postmenopausal women undergoing hysterectomy for endometrial prolapse. The impact of E(2) treatment on endometrial gene expression (799 genes) was much higher than the effect of tibolone (173 genes) or E(2) + MPA treatment (174 genes). Furthermore, endometrial gene expression profiles after tibolone treatment show a weak similarity to the profiles after E(2) treatment (overlap 72 genes) and even less profile similarity to E(2) + MPA treatment (overlap 17 genes). Interestingly, 95 tibolone-specific genes were identified. Translation of profile similarity into biological processes and pathways showed that ER-mediated downstream processes, such as cell cycle and cell proliferation, are not affected by E2 + MPA, slightly by tibolone, but are significantly affected by E(2). In conclusion, tibolone treatment results in a tibolone-specific gene expression profile in the human endometrium, which shares only limited resemblance to E(2) and even less resemblance to E2 + MPA induced profiles.
Asunto(s)
Endometrio/efectos de los fármacos , Estradiol/efectos adversos , Terapia de Reemplazo de Estrógeno/efectos adversos , Histerectomía Vaginal , Medroxiprogesterona/efectos adversos , Norpregnenos/efectos adversos , Transducción de Señal/efectos de los fármacos , Prolapso Uterino/tratamiento farmacológico , Análisis por Conglomerados , Quimioterapia Combinada , Endometrio/metabolismo , Endometrio/cirugía , Femenino , Expresión Génica/efectos de los fármacos , Perfilación de la Expresión Génica/métodos , Redes Reguladoras de Genes/efectos de los fármacos , Humanos , Análisis de Secuencia por Matrices de Oligonucleótidos , Reacción en Cadena de la Polimerasa , Posmenopausia , ARN Mensajero/metabolismo , Reproducibilidad de los Resultados , Globulina de Unión a Hormona Sexual/metabolismo , Transducción de Señal/genética , Prolapso Uterino/metabolismo , Prolapso Uterino/cirugíaRESUMEN
Tibolone, a steroidogenic compound with both estrogenic and progestagenic properties, is used as an alternative for estrogen or estrogen plus progesterone hormone therapy for the treatment of symptoms associated with menopause and osteoporosis. We have evaluated whether the effect of tibolone on a human endometrial cell line is similar to, or comparable with, the effect of estradiol (E(2)), medroxyprogesterone acetate (MPA) or E(2) + MPA treatment. Using stable transfection techniques, the estrogen receptor (ER) expressing human endometrial cancer cell line, ECC1, was altered to also express both progesterone receptors (PRs). These cells were then used to assess growth regulation and expression profiling (Affymetrix U133plus2) under the influence of E(2) (1 nM), MPA (1 nM), E(2) + MPA or tibolone (100 nM). Growth assessment and comparison of profiles indicate that tibolone behaves predominantly like MPA. Furthermore, regulation of prereplication complex genes, such as the minichromosome maintenance genes, could be involved in the observed strong inhibition of growth by tibolone as well as MPA. In addition, in total, 15 genes were found to be specific for tibolone treatment. These genes were predominantly involved in regulation of the cell cycle and differentiation.
Asunto(s)
Neoplasias Endometriales/genética , Neoplasias Endometriales/patología , Estrógenos/metabolismo , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Terapia de Reemplazo de Hormonas , Acetato de Medroxiprogesterona/farmacología , Norpregnenos/farmacología , Progesterona/metabolismo , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Neoplasias Endometriales/metabolismo , Estrógenos/farmacología , Femenino , Perfilación de la Expresión Génica , Humanos , Análisis de Secuencia por Matrices de Oligonucleótidos , Progesterona/farmacología , Transcripción Genética/efectos de los fármacos , Transcripción Genética/genéticaRESUMEN
Estrogen-stimulated growth of the malignant human endometrium can be balanced by the differentiating properties of progesterone. To study the molecular basis behind this, gene expression profiling was performed using complementary DNA microarray analysis. In this study, the human endometrial cancer cell lines ECC-1 and PRAB-36 were used as models. The ECC-1 cell line, which expresses high levels of estrogen receptor alpha and is stimulated in growth by estrogens, was used to study estrogen regulation of gene expression. The Ishikawa sub-cell line PRAB-36, expressing both PRA and PRB, progesterone receptor isoforms, and inhibited in growth by progestagens, was used to study progesterone regulation of gene expression. Using these two well-differentiated human endometrial cancer cell lines, 148 estrogen- and 148 progesterone-regulated genes were identified. After functional classification, the estrogen- and progesterone-regulated genes could be categorized in different biologically relevant groups. Within the group of "cell growth and/or maintenance," 81 genes were clustered, from which a number of genes could be involved in arranging the cross talk that exists between estrogen and progesterone signaling. On the basis of analysis of the current findings, it is hypothesized that cross talk between estrogen and progestagen signaling does not occur by counterregulation of single genes, but rather at the level of differential regulation of different genes within the same functional families.
Asunto(s)
Neoplasias Endometriales/patología , Estrógenos/farmacología , Progesterona/farmacología , Receptores de Progesterona/metabolismo , Western Blotting , Diferenciación Celular/genética , Línea Celular Tumoral , Proliferación Celular , Estrógenos/metabolismo , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Análisis de Secuencia por Matrices de Oligonucleótidos , Progesterona/metabolismo , Receptores de Progesterona/genética , Sensibilidad y EspecificidadRESUMEN
Tamoxifen treatment for breast cancer increases proliferation of the endometrium, resulting in an enhanced prevalence of endometrial pathologies, including endometrial cancer. An exploratory study was performed to begin to understand the molecular mechanism of tamoxifen action in the endometrium. Gene-expression profiles were generated of endometrial samples of tamoxifen users and compared with matched controls. The pathological classification of samples from both groups included atrophic/inactive endometrium and endometrial polyps. Unsupervised clustering revealed that samples of tamoxifen users were, irrespective of pathological classification, fairly similar and consequently form a subgroup distinct from the matched controls. Using SAM analysis (a statistical method to select genes differentially expressed between groups), 256 differentially expressed genes were selected between the tamoxifen and control groups. Upon comparing these genes with oestrogen-regulated genes, identified under similar circumstances, 95% of the differentially expressed genes turned out to be tamoxifen-specific. Finally, construction of a gene-expression network of the differentially expressed genes revealed that 69 genes centred around five well-known genes: TP53, RELA, MYC, epidermal growth factor receptor and beta-catenin. This could indicate that these well-known genes, and the pathways in which they function, are important for tamoxifen-controlled proliferation of the endometrium.
Asunto(s)
Antineoplásicos Hormonales/uso terapéutico , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/genética , Endometrio/metabolismo , Expresión Génica/efectos de los fármacos , Tamoxifeno/uso terapéutico , Antineoplásicos Hormonales/farmacología , Proliferación Celular/efectos de los fármacos , Endometrio/efectos de los fármacos , Endometrio/patología , Receptores ErbB/genética , Femenino , Perfilación de la Expresión Génica , Genes Relacionados con las Neoplasias , Genes myc/genética , Genes p53/genética , Humanos , Persona de Mediana Edad , Tamoxifeno/farmacología , Factor de Transcripción ReIA/genética , beta Catenina/genéticaRESUMEN
Tibolone is a synthetic steroid with estrogenic effects on brain, vagina, and bone without stimulating the endometrium. During tibolone treatment, it is thought that the progestagenic properties of tibolone stimulate cell differentiation, which effectively counterbalances the growth-stimulating effects of the estrogenic properties of tibolone. The objective of this study was to characterize the expression profile that reflects the endometrial responses to the separated estrogenic (growth-inducing) and progestagenic (growth-inhibiting) actions of tibolone, thus gaining insight into the counteracting effect of these properties of tibolone on the endometrium. The estrogenic action of tibolone was studied in the estrogen-responsive ECC1 cell line (expressing estrogen receptor alpha), and the progestagenic action was studied in the progesterone-responsive cell line Ishikawa PRAB-36 (expressing PRA and PRB). The data showed that the progestagenic and estrogenic effects of tibolone produce different expression profiles with a narrow overlap in genes; however, both properties modulate the same biological processes. The final genetic network analysis indicated that the estrogenic effect of tibolone is potentially counterbalanced by the progestagenic metabolite of tibolone via differential regulation of similar cellular processes. For example, both progestagenic and estrogenic properties stimulate proliferation, but they exert the opposite effect on apoptosis. The apoptosis network was stimulated by the progestagenic properties of tibolone; in contrast, the estrogenic effect of tibolone suppressed the apoptosis network. The current results indicate that this differential regulation is realized through modulation of a different group of genes and rarely via contraregulation of the same set of genes.
Asunto(s)
Moduladores de los Receptores de Estrógeno/farmacología , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Norpregnenos/farmacología , Progestinas/antagonistas & inhibidores , Adenocarcinoma/genética , Adenocarcinoma/patología , Diferenciación Celular/efectos de los fármacos , División Celular/efectos de los fármacos , Línea Celular Tumoral , Neoplasias Endometriales/genética , Neoplasias Endometriales/patología , Estradiol/farmacología , Femenino , Perfilación de la Expresión Génica , Humanos , Red Nerviosa , Transcripción Genética/efectos de los fármacosRESUMEN
OBJECTIVE: Recently, gene expression profiling techniques have been used on several human cancers to classify tumor subgroups with a specific biological behavior, which were previously undetected by the conventional histopathologic staging systems. In the current study, the clinical usefulness and prognostic value of gene expression profiling in human endometrial carcinomas were studied. METHODS: A macro cDNA array, containing cDNAs of 588 genes selected from different areas of cancer research, was used to generate gene expression profiles of tumor tissue samples. The gene expression profiles of 12 endometrial cancers, 3 benign (e.g. noncancer) endometrial tissue samples and 3 myometrial tissue samples, taken from human surgical specimen, were compared. RESULTS: The efficacy to generate a gene expression profile of these tissue samples was 77%. The RNA samples could be randomly taken from the tissue samples and were highly reproducible. Cluster analysis of gene expression profiles of the different samples showed that the benign endometrial and the myometrial samples clustered separately from the tumor samples, indicating that the gene expression profiles were tissue specific and not patient specific. Cluster analysis of the tumor samples revealed two distinct tumor clusters. Ranking of the tumors in the two clusters showed high similarity with the histopathologic classification [International Federation of Gynecology and Obstetrics (FIGO) grading]. CONCLUSION: Classification of endometrial tumors on basis of their gene expression profiles showed similarity with the FIGO grading system.
Asunto(s)
Neoplasias Endometriales/genética , Adulto , Anciano , Anciano de 80 o más Años , Neoplasias Endometriales/diagnóstico , Neoplasias Endometriales/metabolismo , Femenino , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Humanos , Persona de Mediana Edad , Familia de Multigenes , Análisis de Secuencia por Matrices de Oligonucleótidos , Pronóstico , Sensibilidad y EspecificidadRESUMEN
Tibolone, a synthetic steroid acting in a tissue-specific manner and used in hormone replacement therapy, is converted into three active metabolites: a Delta(4) isomer (exerting progestogenic and androgenic effects) and two hydroxy metabolites, 3 alpha-hydroxytibolone (3 alpha-OH-tibolone) and 3beta-OH-tibolone (exerting estrogenic effects). In the present study an endometrial carcinoma cell line (Ishikawa PRAB-36) was used to investigate the progestogenic properties of tibolone and its metabolites. This cell line contains progesterone receptors A and B, but lacks estrogen and androgen receptors. When tibolone was added to the cells, complete conversion into the progestogenic/androgenic Delta(4) isomer was observed within 6 d. Furthermore, when cells were cultured with tibolone or when the Delta(4) isomer or the established progestagen medroxyprogesterone acetate was added to the medium, marked inhibition of growth was observed. Interestingly, 3 beta-OH-tibolone also induces some inhibition of growth. These growth inhibitions were not observed in progesterone receptor-negative parental Ishikawa cells, and progestagen-induced growth inhibition of PRAB-36 cells could readily be reversed using the antiprogestagen Org-31489. Upon measuring the expression of two progesterone-regulated genes (fibronectin and IGF-binding protein-3), tibolone, the Delta(4) isomer and medroxyprogesterone acetate showed similar gene expression regulation. These results indicate that tibolone, the Delta(4) metabolite, and to some extent 3 beta-OH-tibolone exert progestogenic effects. Tibolone and most likely 3 beta-OH-tibolone are converted into the Delta(4) metabolite.
Asunto(s)
Neoplasias Endometriales/metabolismo , Norpregnenos/farmacología , Progestinas/metabolismo , División Celular/efectos de los fármacos , Neoplasias Endometriales/química , Neoplasias Endometriales/patología , Femenino , Fibronectinas/genética , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Proteína 3 de Unión a Factor de Crecimiento Similar a la Insulina/genética , Acetato de Medroxiprogesterona/farmacología , Norpregnenos/metabolismo , Progesterona/farmacología , Progestinas/antagonistas & inhibidores , Receptores de Progesterona/análisis , Células Tumorales CultivadasRESUMEN
Identification of genes involved in the transition from androgen-dependent to androgen-independent prostate cancer is important to extend our current knowledge of the disease. Using differential display RT-PCR analysis between androgen-dependent and androgen-independent prostate cancer cells, we have identified a novel gene, designated GC109. GC109 harbours a putative Cys-His cluster, a nuclear localisation signal, a leucine zipper and a ret finger protein (rfp)-like domain. GC109 mRNA expression in normal human tissues was found not to be restricted to the prostate. However, using a variety of 15 human cancer cell lines, GC109 mRNA was preferentially expressed in androgen-dependent LNCaP-FGC, compared with androgen-independent LNCaP-LNO, DU145 and PC3 human prostate cancer cells. Finally, the GC109 gene was mapped on human chromosome 2p24. Based on its protein domain structure and chromosomal localisation, we hypothesise that GC109 may be involved in chromosomal rearrangements in prostate cancer.
Asunto(s)
Andrógenos/fisiología , Cromosomas Humanos Par 2/genética , Regulación Neoplásica de la Expresión Génica , Neoplasias Hormono-Dependientes/genética , Neoplasias de la Próstata/genética , Secuencia de Bases , Northern Blotting , Mapeo Cromosómico , Humanos , Masculino , Datos de Secuencia Molecular , Neoplasias Hormono-Dependientes/metabolismo , Neoplasias de la Próstata/metabolismo , ARN Mensajero/genética , ARN Neoplásico/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Células Tumorales CultivadasRESUMEN
BACKGROUND: The transition from androgen-dependent to androgen-independent prostate cancer is not fully understood but appears to involve multiple genetic changes. We have identified a gene, GC79, that is more highly expressed in androgen-dependent LNCaP-FGC human prostate cancer cells than in androgen-independent LNCaP-LNO human prostate cancer cells. Physiologic levels (0.1 nM:) of androgens repress expression of GC79 messenger RNA (mRNA) in LNCaP-FGC cells. To determine the role of GC79, we cloned its complementary DNA (cDNA) and functionally characterized its product. METHODS: The differentially expressed GC79 gene was cloned from human prostate cDNA libraries, sequenced, and transfected into mammalian cells to study its function. Expression of GC79 was analyzed in various adult and fetal human tissues and in prostate glands of castrated rats. The association of GC79 expression and apoptosis was investigated in COS-1 and LNCaP cells transfected with GC79 cDNA. All statistical tests are two-sided. RESULTS: Sequence analysis indicates that GC79 encodes a large, complex, multitype zinc-finger protein, containing nine C(2)H(2)-type zinc-finger domains, a cysteine-rich region, and a GATA C(4)-type zinc-finger domain. Castration-induced androgen withdrawal increased the expression of GC79 mRNA in the regressing rat ventral prostate, suggesting that the expression of GC79 mRNA is associated with the process of apoptotic cell death in the rat ventral prostate. Transfection and induction of GC79 cDNA in both COS-1 and LNCaP prostate cancer cells led to an apoptotic index that was eightfold higher (P:<.001, two-sided Student's t test) than that observed in uninduced transfected cells. CONCLUSIONS: We have cloned an androgen-repressible gene, GC79, that is potentially involved in apoptosis. This finding may have implications for the development of androgen-independent prostate cancer and, ultimately, for the treatment of prostate cancer.
Asunto(s)
Andrógenos/metabolismo , Proteínas de Unión al ADN , Proteínas de Neoplasias/genética , Neoplasias de la Próstata/genética , Neoplasias de la Próstata/patología , Dedos de Zinc/genética , Secuencia de Aminoácidos , Animales , Apoptosis , Secuencia de Bases , Northern Blotting , Clonación Molecular , Regulación Neoplásica de la Expresión Génica , Humanos , Masculino , Datos de Secuencia Molecular , Neoplasias de la Próstata/metabolismo , ARN Mensajero/análisis , ARN Neoplásico/análisis , Proteínas Represoras , Factores de Transcripción , Células Tumorales CultivadasRESUMEN
The recently developed cDNA expression array technique can be used to generate gene-expression fingerprints of tumour specimens. To gain insight into molecular mechanisms involved in the development and progression of cancer, this cDNA expression array technique could be a useful tool, however, no established methods for interpreting the results are yet available. We used the Atlas cancer cDNA expression array (Clontech, USA) for analysing total RNA isolated from four human endometrial carcinoma samples (two cell-lines and two tissue samples), one benign endometrial tissue sample and a human breast cancer cell-line, in order to develop a method for analysing the array data. The obtained gene-expression profiles were highly reproducible. XY-scatterplots and regression analysis of the logarithmic transformed data provided a practical method to analyse the data without the need of preceding normalization. Three genes (Decorin, TIMP3 and Cyclin D1) were identified to be differentially expressed between the benign endometrial tissue sample and the endometrial carcinoma samples (tissue and cell-lines). These three genes may potentially be involved in cancer progression. A higher degree of similarity in gene-expression profile was found between the endometrial samples (tissue and cell-lines) than between the endometrial samples and the breast cancer cell-line, which is indicative for an endometrial tissue-specific gene-expression profile.
Asunto(s)
Neoplasias Endometriales/genética , Perfilación de la Expresión Génica , Análisis de Secuencia por Matrices de Oligonucleótidos/métodos , ADN Complementario/análisis , Estudios de Evaluación como Asunto , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Reproducibilidad de los Resultados , Células Tumorales CultivadasRESUMEN
Using differential-display RT-PCR analysis between androgen-dependent LNCaP-FGC and androgen-independent LNCaP-LNO human prostate-cancer cells, we have identified a gene not previously described as being expressed in prostate. The gene is more highly expressed in androgen-independent than in androgen-dependent LNCaP prostate-cancer cells. Sequence analysis showed that the gene has already been cloned as a transcript present in embryonic brain, with unknown functions. Expression of the gene was found not to be restricted to the prostate, and not regulated by androgens in androgen-independent prostate-cancer cells. In concert with the cell-culture system, Northern-blot analysis of gene expression in vivo, using a panel of human prostate-cancer xenografts, demonstrated that the gene is more highly expressed in androgen-independent than in androgen-dependent prostate-cancer xenografts. The gene could be mapped on human chromosome 8q11. The 8q arm is known to be frequently amplified during prostate-cancer progression and harbors several proto-oncogenes potentially involved in cancer development. Since expression of the gene is positively correlated with prostate-cancer progression and its 8q11 chromosomal localization, we hypothesize that the gene may be involved in the development and progression of prostate cancer.
Asunto(s)
Carcinoma/genética , Cromosomas Humanos Par 8/genética , Regulación Neoplásica de la Expresión Génica , Genes Relacionados con las Neoplasias , Neoplasias de la Próstata/genética , Animales , Secuencia de Bases , Northern Blotting , Mapeo Cromosómico , Progresión de la Enfermedad , Humanos , Masculino , Ratones , Ratones Desnudos , Datos de Secuencia Molecular , Trasplante de Neoplasias , Neoplasias Hormono-Dependientes/genética , ARN Mensajero/biosíntesis , Células Tumorales CultivadasRESUMEN
The beta 1-subunit of Na+,K+-ATPase was isolated and identified as an androgen down-regulated gene. Expression was observed at high levels in androgen-independent as compared to androgen-dependent (responsive) human prostate cancer cell lines and xenografts when grown in the presence of androgens. Down-regulation of the beta 1-subunit was initiated at concentrations between 0.01 nM and 0.03 nM of the synthetic androgen R1881 after relatively long incubation times (> 24 h). Using polyclonal antibodies, the concentration of beta 1-subunit protein, but not of the alpha 1-subunit protein, was markedly reduced in androgen-dependent human prostate cancer cells (LNCaP-FGC) cultured in the presence of androgens. In line with these observations it was found that the protein expression of total Na+,K+-ATPase in the membrane (measured by 3H-ouabain binding) was also markedly decreased. The main function of Na+,K+-ATPase is to maintain sodium and potassium homeostasis in animal cells. The resulting electrochemical gradient is facilitative for transport of several compounds over the cell membrane (for example cisplatin, a chemotherapeutic agent experimentally used in the treatment of hormone-refractory prostate cancer). Here we observed that a ouabain-induced decrease of Na+,K+-ATPase activity in LNCaP-FGC cells results in reduced sensitivity of these cells to cisplatin-treatment. Surprisingly, androgen-induced decrease of Na+,K+-ATPase expression, did not result in significant protection against the chemotherapeutic agent.
Asunto(s)
Andrógenos/fisiología , Neoplasias Hormono-Dependientes/enzimología , Neoplasias de la Próstata/enzimología , ATPasa Intercambiadora de Sodio-Potasio/biosíntesis , Animales , Antineoplásicos/antagonistas & inhibidores , Antineoplásicos/farmacología , Muerte Celular/efectos de los fármacos , Cisplatino/antagonistas & inhibidores , Cisplatino/farmacología , Regulación hacia Abajo/efectos de los fármacos , Regulación Enzimológica de la Expresión Génica/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Sustancias Macromoleculares , Masculino , Metribolona/farmacología , Ratones , Ratones Desnudos , Trasplante de Neoplasias , Neoplasias Hormono-Dependientes/tratamiento farmacológico , Neoplasias Hormono-Dependientes/genética , Neoplasias Hormono-Dependientes/patología , Reacción en Cadena de la Polimerasa , Neoplasias de la Próstata/tratamiento farmacológico , Neoplasias de la Próstata/genética , Neoplasias de la Próstata/patología , ATPasa Intercambiadora de Sodio-Potasio/genética , ATPasa Intercambiadora de Sodio-Potasio/metabolismo , Congéneres de la Testosterona/farmacología , Trasplante Heterólogo , Células Tumorales CultivadasRESUMEN
Androgens play a crucial role in several stages of male development and in the maintenance of the male phenotype. Androgens act in their target cells via an interaction with the androgen receptor, resulting in direct regulation of gene expression. The androgen receptor is a phosphoprotein and modulation of the phosphorylation status of the receptor influences ligand-binding and consequently transcription activation of androgen responsive genes. Androgen binding induces a conformational change in the ligand-binding domain, accompanied by additional receptor phosphorylation. Subsequently the liganded androgen receptor interacts with specific androgen response elements in the regulatory regions of androgen target genes, resulting in stimulation of gene expression. Anti-androgens induce a different conformational change of the ligand-binding domain, which does not or only partially result in stimulation of transactivation. Interestingly, different anti-androgens can induce different inactive conformations of the androgen receptor ligand-binding domain. Recent evidence strongly supports a ligand dependent functional interaction between the ligand-binding domain and the NH2-terminal transactivating domain of the androgen receptor. Two regions in the NH2-terminal domain are involved in this interaction, whereas in the ligand-binding domain the AF-2 AD core region is involved.
Asunto(s)
Receptores Androgénicos/metabolismo , Andrógenos/metabolismo , Humanos , Ligandos , Fosforilación , Conformación Proteica , Receptores Androgénicos/química , Receptores Androgénicos/genética , Activación TranscripcionalRESUMEN
High doses of progesterone are used in the treatment of advanced and recurrent endometrial cancer. Unfortunately the response rate is relatively low: 10-30%. The mechanisms involved in the development of insensitivity to progesterone treatment of endometrial cancer tissue are largely unknown. As tumour development is thought to be associated with a cascade of genetic alterations, it can be expected that genetic changes are involved in the development of progesterone insensitivity in endometrial carcinomas. We therefore started an investigation to identify, isolate and characterise progesterone-regulated genes involved in progesterone-induced growth inhibition in endometrial carcinoma cells. Using differential display PCR eight progesterone-regulated cDNA clones were identified in endometrial carcinoma cell lines. Four of these progesterone-regulated cDNA clones were regulated in the for growth progesterone-sensitive cell line IK-3H12 and not regulated in the for growth-insensitive cell line ECC-1. This indicates that these four cDNA clones represent potentially important genes, which could be involved in inhibition of growth of endometrial carcinoma tissue by progesterone.
Asunto(s)
Antineoplásicos/uso terapéutico , Neoplasias Endometriales/tratamiento farmacológico , Neoplasias Endometriales/metabolismo , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Progesterona/uso terapéutico , Antineoplásicos/farmacología , División Celular/efectos de los fármacos , Resistencia a Antineoplásicos , Femenino , Humanos , Progesterona/farmacología , Células Tumorales CultivadasRESUMEN
The androgen receptor (AR) protein is an important transacting factor that is necessary for mediating gene expression of androgen-responsive genes. The expression of the AR gene is regulated by androgens and agents that utilize the calcium, protein kinase A, and protein kinase C pathways. Although the role of the calcium and protein kinase A pathways in the regulation of the AR gene has been investigated, the mechanism of regulation of AR through the protein kinase C pathway is not known. We have isolated the 5'-flanking region of the mouse AR gene and identified a consensus TPA (12-O-tetradecanoylphorbol 13-acetate)-response element (TRE). Transient transfection assays indicate that the TRE sequence is sufficient to confer TPA responsiveness to cells treated with TPA. Gel retardation assays and DNA footprint analysis demonstrated specific binding of the TRE and protection of the TRE sequence. Thus, these results describe a TRE in the 5'-flanking region of the AR gene and demonstrate that the TRE is responsive to TPA treatment.
Asunto(s)
Receptores Androgénicos/genética , Secuencias Reguladoras de Ácidos Nucleicos , Animales , Línea Celular , Cloranfenicol O-Acetiltransferasa/genética , Huella de ADN , Regulación de la Expresión Génica , Ratones , ARN Mensajero/metabolismo , Acetato de Tetradecanoilforbol/farmacología , TransfecciónRESUMEN
When androgen receptor containing cells are cultured in the presence of the PKA stimulator forskolin, a rapid dephosphorylation of the androgen receptor occurs resulting in a decrease in the amount of 112 kDa androgen receptor isoform and an increase in 110 kDa androgen receptor isoform on SDS-PAGE. To establish which amino acid residues in the androgen receptor were phosphorylated in control and forskolin-treated cells, trypsin-digested androgen receptors were subjected to RP-HPLC analysis and subsequently to Edman degradation. It was observed that serine residues 506, 641, and 653 were potentially phosphorylated in control cells, while after forskolin treatment strong evidence was obtained that phosphorylation of serines 641 and 653 was significantly reduced. When the dephosphorylated androgen receptor was analyzed for its transcription activation capacity, it was observed that androgen-induced transcriptional regulation of two endogenous genes (PSA) and beta 1-subunit of Na,K-ATPase), in cells cultured in the presence of forskolin, was inhibited as compared to the control situation. The observation that the dephosphorylated androgen receptor was transcriptionally less active was further strengthened by the finding that the dephosphorylated androgen receptor was markedly impaired in ligand binding (Bmax was found to be reduced by approximately 40%). The current investigations show for the first time a clear function for the rapid phosphorylation which occurs directly after synthesis of the androgen receptor, namely, effective ligand binding.