ABSTRACT
The biological outcome of TMPRSS2:ERG chromosomal translocations in prostate cancer (PC) remains poorly understood. To address this, we compared the transcriptional effects of TMPRSS2:ERG expression in a transgenic mouse model with those of ERG knockdown in a TMPRSS2:ERG-positive PC cell line. This reveals that ERG represses the expression of a previously unreported set of androgen receptor (AR)-independent neuronal genes that are indicative of neuroendocrine (NE) cell differentiation-in addition to previously reported AR-regulated luminal genes. Cell sorting and proliferation assays performed after sustained ERG knockdown indicate that ERG drives proliferation and blocks the differentiation of prostate cells to both NE and luminal cell types. Inhibition of ERG expression in TMPRSS2:ERG-positive PC cells through blockade of AR signaling is tracked with increased NE gene expression. We also provide evidence that these NE cells are resistant to pharmacological AR inhibition and can revert to the phenotype of parental cells upon restoration of AR/ERG signaling. Our findings highlight an ERG-regulated mechanism capable of repopulating the parent tumor through the transient generation of an anti-androgen therapy-resistant cell population, suggesting that ERG may have a direct role in preventing resistance to anti-androgen therapy.
Subject(s)
Cell Differentiation/genetics , Cell Proliferation/genetics , Prostatic Neoplasms/genetics , Serine Endopeptidases/genetics , Trans-Activators/genetics , Animals , Antibiotics, Antineoplastic/pharmacology , Benzamides , Cell Differentiation/drug effects , Cell Line, Tumor , Cell Proliferation/drug effects , Disease Models, Animal , Doxorubicin/pharmacology , Epithelial Cells/metabolism , Gene Expression Profiling , Gene Expression Regulation, Neoplastic , Humans , Immunohistochemistry , Male , Mice, Transgenic , Microscopy, Confocal , Neuroendocrine Cells/metabolism , Nitriles , Oncogene Proteins, Fusion/genetics , Oncogene Proteins, Fusion/metabolism , Phenylthiohydantoin/analogs & derivatives , Phenylthiohydantoin/pharmacology , Prostatic Neoplasms/metabolism , Prostatic Neoplasms/pathology , RNA Interference , Receptors, Androgen/genetics , Receptors, Androgen/metabolism , Reverse Transcriptase Polymerase Chain Reaction , Serine Endopeptidases/metabolism , Trans-Activators/metabolism , Transcriptional Regulator ERG , Translocation, GeneticABSTRACT
Abscisic acid-responsive element binding protein (AREB1) is a basic domain/leucine zipper transcription factor that binds to the abscisic acid (ABA)-responsive element motif in the promoter region of ABA-inducible genes. Because AREB1 is not sufficient to direct the expression of downstream genes under non-stress conditions, an activated form of AREB1 (AREB1ΔQT) was created. Several reports claim that plants overexpressing AREB1 or AREB1ΔQT show improved drought tolerance. In our studies, soybean plants overexpressing AREB1ΔQT were characterized molecularly, and the phenotype and drought response of three lines were accessed under greenhouse conditions. Under conditions of water deficit, the transformed plants presented a higher survival rate (100%) than those of their isoline, cultivar BR 16 (40%). Moreover, the transformed plants displayed better water use efficiency and had a higher number of leaves than their isoline. Because the transgenic plants had higher stomatal conductance than its isoline under well-watered conditions, it was suggested that the enhanced drought response of AREB1ΔQT soybean plants might not be associated with altered transpiration rates mediated by ABA-dependent stomatal closure. However, it is possible that the smaller leaf area of the transgenic plants reduced their transpiration and water use, causing delayed stress onset. The difference in the degree of wilting and percentage of survival between the 35S-AREB1ΔQT and wildtype plants may also be related to the regulation of genes that protect against dehydration because metabolic impairment of photosynthesis, deduced by an increasing internal CO2 concentration, was not observed in the transgenic plants.
Subject(s)
Arabidopsis Proteins/genetics , Arabidopsis/genetics , Basic-Leucine Zipper Transcription Factors/genetics , Gene Expression Regulation, Plant , Glycine max/genetics , Plant Leaves/genetics , Water/metabolism , Abscisic Acid/metabolism , Arabidopsis/chemistry , Arabidopsis Proteins/metabolism , Basic-Leucine Zipper Transcription Factors/metabolism , Droughts , Plant Leaves/metabolism , Plants, Genetically Modified , Response Elements , Glycine max/metabolism , TransgenesABSTRACT
A slide hemagglutination test, here called SHAT, which is practical and economical for seroepidemiological surveys was standardized. This is an improved modification of the rapid hemagglutination test (RHA) which utilizes a short-lived reagent prepared with fresh blood cells. The reagent and conditions of the test were considerably modified and, most importantly, an alkaline-solubilized Trypanosoma cruzi epimastigote antigen reagent is proposed. The stability of the SHAT reagent was at least one year at 4 degrees C, in an appropriate liquid suspension. The SHAT was applied to 71 serum samples from patients with Chagas' disease and from 235 clinically healthy blood donors. Sensitivity, specificity and positive and negative predictive values for the selected cutoff titer corresponding to 1:4 dilution were 0.972 (0.903-0.992), 0.983 (0.957-0.993), 0.945 (0.867-0.979) and 0.991 (0.969-0.998), respectively. These values were comparable to those found for the RHA, immunofluorescence (IFT), indirect hemagglutination (IHAT) and complement fixation (CFT) tests. These data suggest that the SHAT should be useful for seroepidemiological surveys conducted at public health laboratories in developing countries.
Subject(s)
Chagas Disease/diagnosis , Hemagglutination Tests/methods , Humans , Predictive Value of Tests , Sensitivity and Specificity , Seroepidemiologic StudiesABSTRACT
A slide hemagglutination test, here called SHAT, which is practical and economical for seroepidemiological surveys was standardized. This is an improved modification of the rapid hemagglutination test (RHA) which utilizes a short-lived reagent prepared with fresh blood cells. The reagent and conditions of the test were considerably modified and, most importantly, an alkaline-solubilized Trypanosoma cruzi epimastigote antigen reagent is proposed. The stability of the SHAT reagent was at least one year at 4 degrees Celsius, in an appropriate liquid suspension. The SHAT was applied to 71 serum samples from patients with Chagas' disease and from 235 clinically healthy blood donors. Sensitivity, specificity and positive and negative predictive values for the selected cutoff titer corresponding to 1:4 dilution were 0.972 (0.903-0.992)9 O.983 (O.957-0.993) 0.945 (O.867-0.979) and O.991 (O.969-0.998), respectively. These values were comparable to those found for the RHA, immunofluorescence (IFT), indirect hemagglutination (IHAT) and complement fixation (CFT) tests. These data suggest that the SHAT should be useful for seroepidemiological surveys conducted at public health laboratories in developing countries.
Subject(s)
Humans , Chagas Disease/diagnosis , Hemagglutination Tests/methods , Predictive Value of Tests , Sensitivity and Specificity , Seroepidemiologic StudiesABSTRACT
In the present study we characterized the cytotoxicity of Wistar rat mononuclear cells from 21 animals which received 10(6) Walker 256 tumor cells by the subcutaneous route. All animals developed the tumor. Cytotoxicity was studied 15 days after inoculation using spleen, thymus and lymph node T lymphocytes as well as macrophages from the peritoneal cavity. A Walker 256 tumor cell suspension and tumor cells in culture (YAC-1) were labelled with 51Cr and used as target cells, according to the Herberman technique and a gamma counter was used for counting. Anti-Walker cell cytotoxicity was significantly decreased in T lymphocytes from the spleen (9.6% vs 51.1% for the control) and thymus (11.5% vs 38.2% for the control), whereas no difference was observed for lymph nodes (41.2% vs 49.5% for the control) or macrophages (43.4% vs 46.3% for the control). Anti-YAC-1 cytotoxicity was significantly decreased in T lymphocytes from all lymphoid organs compared to control: 23.6% vs 42.8% for the spleen, 22.6% vs 41.1% for the thymus, 26.6% vs 42.1% for lymph nodes, and 27.1% vs 46.3% for macrophages. No correlation was observed between tumor weight, and anti-Walker cytotoxicity or anti-YAC-1 cytotoxicity.
Subject(s)
Carcinoma 256, Walker/immunology , Killer Cells, Natural/immunology , T-Lymphocytes/immunology , Animals , Cytotoxicity, Immunologic , Rats , Rats, Wistar , Tumor Cells, CulturedABSTRACT
In the present study we characterized the cytotoxicity of Wistar rat mononuclear cells from 21 animals which received 10(6) Walker 256 tumor cells by the subcutaneous route. All animals developed the tumor. Cytotoxicity was studied 15 days after inoculation using spleen, thymus and lymph node T lymphocytes as well as macrophages from the peritoneal cavity. A Walker 256 tumor cell suspension and tumor cells in culture (YAC-1) were labelled with 51Cr and used as target cells, according to the Herberman technique and a gamma counter was used for counting. Anti-Walker cell cytotoxicity was significantly decreased in T lymphocytes from the spleen (9.6 vs 51.1 for the control) and thymus (11.5 vs 38.2 for the control), whereas no difference was observed for lymph nodes (41.2 vs 49.5 for the control) or macrophages (43.4 vs 46.3 for the control). Anti-YAC-1 cytotoxicity was significantly decreased in T lymphocytes from all lymphoid organs compared to control: 23.6 vs 42.8 for the spleen, 22.6 vs 41.1 for the thymus, 26.6 vs 42.1 for lymph nodes, and 27.1 vs 46.3 for macrophages. No correlation was observed between tumor weight, and anti-Walker cytotoxicity or anti-YAC-1 cytotoxicity.
