Journal: Genomics Data

Article Title: Expression-profiling of apoptosis induced by ablation of the long ncRNA TRPM2-AS in prostate cancer cell

doi: 10.1016/j.gdata.2014.10.020

Figure Lengend Snippet: Validation of microarray data by qRT-PCR. Ten transcripts were quantified by qRT-PCR both in control and in TRPM2-AS KO PC3 cells. The resulting expression fold change is plotted against the expression fold change obtained from the Illumina HumanHT-12 V3.0 microarray data. A correlation coefficient of 0.97 was found between the two datasets.

Article Snippet: The human, androgen-independent prostate cancer cell line PC3 was obtained from ATCC (CRL-1435, ATCC).

Techniques: Biomarker Discovery, Microarray, Quantitative RT-PCR, Control, Expressing

Journal: Genomics Data

Article Title: Expression-profiling of apoptosis induced by ablation of the long ncRNA TRPM2-AS in prostate cancer cell

doi: 10.1016/j.gdata.2014.10.020

Figure Lengend Snippet:

Article Snippet: The human, androgen-independent prostate cancer cell line PC3 was obtained from ATCC (CRL-1435, ATCC).

Techniques: Expressing, Microarray, Gene Expression

Journal: PLoS ONE

Article Title: The Response of the Prostate to Circulating Cholesterol: Activating Transcription Factor 3 (ATF3) as a Prominent Node in a Cholesterol-Sensing Network

doi: 10.1371/journal.pone.0039448

Figure Lengend Snippet: ( A ) A provisional network was generated from integration of two microarray data sets. Node color represents increases (red), no significant changes (yellow), and decreases (green) in gene expression in murine prostate tissue after cholesterol alteration as ascertained by cDNA microarray. Changes in RNA expression levels of the corresponding nodes in LNCaP cells are shown as colored node boundaries (donut shape) and the color represents increases (red), no significant change (yellow), and decreases (green) in gene expression under CDM conditions compared to control. Arrows indicate direct activation, T-shaped lines direct repression, dashed arrows indirect activation, and lines physical interaction. ( B ) Gene expression under Normo and Hyper conditions ( in vivo ). To verify in vivo microarray data obtained from SCID experiments, mRNA levels of the indicated genes were determined. GAPDH expression was used to normalize gene expression. Error bars represent SD (n = 3). ( C ) Gene expression under Control and Cholesterol-depleted conditions ( in vitro ). LNCaP cells were incubated in CDM for 0, 3 or 16 h, and mRNA levels of the indicated genes were measured by RT-PCR analysis to validate cDNA microarray data. Error bars represent SD (n = 3). * p <0.05 (Student’s t-test).

Article Snippet: LNCaP human prostate tumor cells were purchased from American Type Culture Collection (ATCC, Manassas, VA) and maintained in RPMI1640 media (Invitrogen, Carlsbad, CA) supplemented with 10% FBS and 1% Penicillin/Streptomycin at 37°C under 5% CO 2 .

Techniques: Generated, Microarray, Gene Expression, RNA Expression, Control, Activation Assay, In Vivo, Expressing, In Vitro, Incubation, Reverse Transcription Polymerase Chain Reaction

Journal: PLoS ONE

Article Title: The Response of the Prostate to Circulating Cholesterol: Activating Transcription Factor 3 (ATF3) as a Prominent Node in a Cholesterol-Sensing Network

doi: 10.1371/journal.pone.0039448

Figure Lengend Snippet: ( A ) RT-PCR analysis in vivo . ATF3 levels are reduced in all prostatic lobes from Hyper mice, compared to those from the Normo group (AP = anterior prostate; VP = ventral prostate; DLP = dorsal prostate). ( B ) Immunoblot analysis. Immunoblot of PrEC lysates showed induction of ATF3 protein by CDM (left panel) and by β-cyclodextrin (right panel). MG132, a proteasome inhibitor, also increased ATF3 expression. ( C ) Immunofluorescence analysis. Induction of ATF3 protein by CDM in LNCaP cells as shown by IF. LNCaP cells were treated with CDM for 18 h, stained with anti-ATF3 antibody and nuclei were counterstained with DAPI (left panel: ATF3; middle panel: DAPI; right panel: overlay). ( D ) RT-PCR analysis. ATF3 mRNA levels in LNCaP cells treated with CDM were normalized to levels of GAPDH. RT-PCR analysis shows induction of ATF3 mRNA levels by CDM. ( E–F ) Promoter reporter analysis. A full-length ATF3 promoter was cloned into a luciferase reporter vector and transfected into LNCaP (D) or PrEC (E). Cells were then incubated in Control and CDM medium. ATF3 promoter activity was plotted as arbitrary units (± SD) after normalization with total protein concentration.

Article Snippet: LNCaP human prostate tumor cells were purchased from American Type Culture Collection (ATCC, Manassas, VA) and maintained in RPMI1640 media (Invitrogen, Carlsbad, CA) supplemented with 10% FBS and 1% Penicillin/Streptomycin at 37°C under 5% CO 2 .

Techniques: Reverse Transcription Polymerase Chain Reaction, In Vivo, Western Blot, Expressing, Immunofluorescence, Staining, Clone Assay, Luciferase, Plasmid Preparation, Transfection, Incubation, Control, Activity Assay, Protein Concentration

Journal: Oncotarget

Article Title: Isolation and genome-wide expression and methylation characterization of CD31 + cells from normal and malignant human prostate tissue

doi:

Figure Lengend Snippet: (A) Representative photomicrographs of hematoxylin and eosin stained frozen sections of benign (left) and malignant (right) prostatic tissues of macrodissected tissues used for ECs isolation. ECs are highlighted in the frozen sections by CD31 immunostaining (Original magnification, x200). (B) Representative immunofluorescent photomicrographs of CD31 (green) and vWF (red) expression and uptake of DiI-Ac-LDL (red) (original magnification, ×200) in CD31 + prostate TdECs and NdECs. Absence of pan-cytokeratin expression by immunofluorescence was observed in CD31 + prostate TdECs and NdECs. LNCaP was used as positive control for pan-cytokeratin immunofluorescence. Nuclei are stained with DAPI (blue). Endothelial tube network was formed by primary cultures of prostate TdECs and NdECs (original magnification, x100). (C) Representative reverse transcription-PCR analysis of RNA from primary cultures of TdECs and NdECs, HUVECs, or LNCaP cells using human-specific primers for human CD31, CD34, ICAM-1, VCAM-1, VEGFR1, VEGFR2, AR and PSA. GAPDH was used as a loading control.

Article Snippet: Primary cultures of human prostate endothelial cells (NdECs and TdECs) were cultured in endothelial growth medium [Endothelial Cell Growth Medium MV2 with Supplement Mix (PromoCell GmbH, Heidelberg, Germany) and 100 U/ml penicillin and 100 μg/ml streptomycin (Invitrogen)].

Techniques: Staining, Isolation, Immunostaining, Expressing, Immunofluorescence, Positive Control

Journal: Oncotarget

Article Title: Isolation and genome-wide expression and methylation characterization of CD31 + cells from normal and malignant human prostate tissue

doi:

Figure Lengend Snippet: Quantitative real-time PCR was used to validate gene expression of AREG , JMY , EPB41 , GMNN and FAM53C in endothelial cells derived from malignant lesions vs benign lesions in AA and CA patients with prostate cancer. Relative gene expression level for qRT-PCR was normalized to the reference gene GAPDH . Gene expression from microarray was plotted together with the qRT-PCR results. Results were shown as Mean ± SD (triplicate). “*” represents p value <0.05 by t-Test.

Article Snippet: Primary cultures of human prostate endothelial cells (NdECs and TdECs) were cultured in endothelial growth medium [Endothelial Cell Growth Medium MV2 with Supplement Mix (PromoCell GmbH, Heidelberg, Germany) and 100 U/ml penicillin and 100 μg/ml streptomycin (Invitrogen)].

Techniques: Real-time Polymerase Chain Reaction, Expressing, Derivative Assay, Quantitative RT-PCR, Microarray

Journal: Cell Death & Disease

Article Title: BTF3 confers oncogenic activity in prostate cancer through transcriptional upregulation of Replication Factor C

doi: 10.1038/s41419-020-03348-2

Figure Lengend Snippet: a BTF3 expression levels in prostate cancer and normal tissue. Transcriptome data of prostate tumors from TCGA and normal tissues from TCGA and GTEx samples were obtained from the UCSC Xena ( https://xena.ucsc.edu ). The gene expression is reported as log 2 (Transcripts per million (TPM) + 0.001). Each dot represents an individual sample ( n = 496 for prostate tumor; n = 149 for normal tissue. The black lines in each group indicate the mean ± S.D. **** p < 0.0001 (Mann–Whitney test). b Immunohistochemistry staining of tissue microarray from prostate tumors and adjacent noncancerous tissues using anti-BTF3 antibody. Representative images of BTF3 stained tumor and their corresponding noncancerous tissue sections are shown in weak, Intermediate, and strong staining, respectively. The scatter plot graph showing a statistical analysis of BTF3 expression (H score) in tumor and adjacent noncancerous tissues. Data are shown as mean ± S.D.; n = 82 for tumors and n = 55 for adjacent noncancerous tissues. Scale bar, 25 μm. ** p < 0.01 (Mann-Whitney test). c Relative cell growth of PC-3 and DU145 prostate cancer cells with or without BTF3-knockdown was measured by crystal violet assay. Data are shown as mean ± S.D. for three independent experiments. *** p < 0.001, **** p < 0.0001 (Student’s t -test). d , e DU145-Tet-On-shBTF3 xenografted mice were treated with or without doxycycline. The graph shows the fold change in tumor volume, with respect to the initial treatment at day 0 ( d ). The tumor volume and representative images of dissected tumors at the endpoint of treatment are shown ( e ). Dox, Doxycycline (2 mg/ml in drinking wat e r). The data are shown as mean ± S.E.M ( n = 6 for each group). **** p < 0.0001 (Student’s t -test).

Article Snippet: 55 adjacent noncancerous tissues and 82 human primary prostate cancer tissues were stained with primary antibody against BTF3 (Abcam, ab203517, Massachusetts, USA).

Techniques: Expressing, MANN-WHITNEY, Immunohistochemistry, Staining, Microarray, Crystal Violet Assay

Journal: Cell Death & Disease

Article Title: BTF3 confers oncogenic activity in prostate cancer through transcriptional upregulation of Replication Factor C

doi: 10.1038/s41419-020-03348-2

Figure Lengend Snippet: a Heat map expression plot of up-regulated and down-regulated genes in siBTF3 #1 transfected PC-3 cells versus control cells (siNC) as revealed by RNA sequencing. The gene expression was calculated according to the FPKM value. Genes with adjusted p -values less than 0.05 are shown. b KEGG pathway enrichment analysis of down-regulated gene sets upon BTF3 knockdown in PC-3 cells. Pathways with adjusted p -value less than 0.05 are shown. The threshold of adjusted p -value = 0.01 is shown as red dotted line. c GSEA analysis of the DNA damage repair gene signatures as indicated in siBTF3 transfected PC-3 cells versus control cells. Normalized enrichment score (NES), p -value and False Discovery Rate (FDR) q value of the correlation are shown. d BTF3 gene expression was plotted against the DNA replication and DNA damage repair signatures as indicated for TCGA prostate cancer datasets ( n = 496). Gene expression levels are reported as log 2 (TPM + 0.001). Each dot represents an individual sample of human prostate tumors. The linear regression Spearman’s correlation coefficient (r) and its p -values are indicated.

Article Snippet: 55 adjacent noncancerous tissues and 82 human primary prostate cancer tissues were stained with primary antibody against BTF3 (Abcam, ab203517, Massachusetts, USA).

Techniques: Expressing, Transfection, RNA Sequencing Assay

Journal: Cell Death & Disease

Article Title: BTF3 confers oncogenic activity in prostate cancer through transcriptional upregulation of Replication Factor C

doi: 10.1038/s41419-020-03348-2

Figure Lengend Snippet: a Flow cytometric analysis of BrdU incorporation was used to determine the S-phase entry in PC-3 and DU145 cells with or without BTF3-knockdown. Scatter plots of newly synthesized DNA (BrdU positive) versus total DNA (PI positive) are shown. The quantification of BrdU positive cells is shown as mean ± S.D. *** p < 0.001 (Student’s t -test). b Comet assay was performed to evaluate DNA damage in cells as in ( a ). Quantification of DNA in the tail from three independent experiments is shown as mean ± S.D. Scale bar, 100 μm. *** p < 0.001 (Student’s t -test). c Immunofluorescence staining of γH2AX (green) and DAPI (blue) in cells as in ( a ). Cells containing more than five foci were scored as positive. Mean ± S.D. for three independent experiments is shown. Scale bar, 20 μm. **** p < 0.0001 (Student’s t -test). d Flow cytometric analysis of the cell cycle for PC-3 and DU145 cells with or without BTF3-knockdown. Quantification of cells in each phase is shown as mean ± S.D. for three independent experiments. ** p < 0.01, **** p < 0.0001 (Student’s t -test).

Article Snippet: 55 adjacent noncancerous tissues and 82 human primary prostate cancer tissues were stained with primary antibody against BTF3 (Abcam, ab203517, Massachusetts, USA).

Techniques: BrdU Incorporation Assay, Synthesized, Single Cell Gel Electrophoresis, Immunofluorescence, Staining

Journal: Cell Death & Disease

Article Title: BTF3 confers oncogenic activity in prostate cancer through transcriptional upregulation of Replication Factor C

doi: 10.1038/s41419-020-03348-2

Figure Lengend Snippet: a Heat map plot of differentially expressed genes involved in DNA replication, nucleotide excision repair and mismatch repair in siBTF3 transfected PC-3 cells or control cells. b Quantitative reverse transcription-PCR (qRT-PCR) analysis of the RFCs mRNA levels was conducted in cells as indicated. Data are shown as mean ± SD for three independent experiments (** p < 0.01, *** p < 0.001, **** p < 0.0001 by Student’s t -test). c qRT-PCR analysis of the RFCs mRNA levels was conducted in DU145-Tet-On-shBTF3 xenografted tumors treated with or without Dox for 22 days. ACTB was used as an endogenous control. Dox, Doxycycline, 2 mg/ml in drinking water. Data are shown as mean ± S.D. * p < 0.05, ** p < 0.01 (Student’s t -test). d The gene expression levels of BTF3 and RFC family genes were examined in the TCGA prostate cancer datasets ( n = 496). The gene expression levels are reported as log 2 (TPM + 0.001) and plotted as BTF3 gene expression over RFC family genes expression. Each dot represents an individual sample of human prostate tumor. Spearman’s correlation coefficient (r) and p -values were determined as indicated. e The RFC promoter luciferase reporter assay was perform e d in siNC or siBTF3 transfected HEK293T cells. pGL3-Basic, control vector; pGL3-RFC1/2/3/4/5, vectors carrying promoter sequence from RFC1, RFC2, RFC3, RFC4 and RFC5, respectively. Data are shown as mean ± S.D. n.s, not significant, ** p < 0.01, *** p < 0.001, **** p < 0.0001 (Student’s t -test). f , g Chromatin immunoprecipitation (ChIP) assay was performed by BTF3 chromatin immunoprecipitation followed by quantitative PCR for the RFC promoters in HEK 293T cells ( f ), PC-3 ( g , left panel) and DU145 ( g , right panel). Enrichment of RFC promoter region was normalized to the input. IgG was used as a negative control. Mean ± S.D. for three independent experiments is shown. * p < 0.05, ** p < 0.01, *** p < 0.001 (Student’s t -test).

Article Snippet: 55 adjacent noncancerous tissues and 82 human primary prostate cancer tissues were stained with primary antibody against BTF3 (Abcam, ab203517, Massachusetts, USA).

Techniques: Transfection, Quantitative RT-PCR, Expressing, Luciferase, Reporter Assay, Plasmid Preparation, Sequencing, Chromatin Immunoprecipitation, Real-time Polymerase Chain Reaction, Negative Control

Journal: The FASEB Journal

Article Title: 12-HETER1/GPR31, a high-affinity 12( S )-hydroxyeicosatetraenoic acid receptor, is significantly up-regulated in prostate cancer and plays a critical role in prostate cancer progression

doi: 10.1096/fj.201500076

Figure Lengend Snippet: 12-HETER1 expression positively correlates with aggressiveness and progression of prostate tumors. A) Expression of 12-HETER1 in prostate carcinoma cells. Immunocytochemistry on HEK293 cells expressing 12-HETER1. a, b) Staining with 12-HETER1 antibody was optimized on HEK293 cells transfected with 12-HETER1 (a) or pcDNA (b) vector. Strong brown staining was detected in HEK293/12-HETER1 cells (a) compared to negative control (b). c, d) Immunohistochemistry to detect 12-HETER1 in 2 cases of human prostate cancer tissue. c) Benign gland (N, left corner, arrow) with rare cells showing positive staining, and neoplastic glands (T, GS-3/4, arrowhead) showing intense brown staining (arrowhead); original magnification, ×10. d) Strong staining in area (upper left, arrow) with mixed GS-3/4 tumor area, weak staining in area (lower right, arrowhead) with GS-3 tumor; original magnification, ×10. Images are representatives from 7 samples. B) Array analysis of 12-HETER1 expression. a, b) Immunohistochemistry to assay 12-HETER1 expression on 2 human prostate tissue microarrays. Expression of 12-HETER1 is significantly up-regulated in disease stage II/III human prostate tumor specimens (n = 8, P = 0.007, Student’s t test. c, d) Representative images of normal (c) and prostate tumor (d) specimens.

Article Snippet: Next we used qPCR to determine 12-HETER1 expression in 2 commercially available human prostate tissue microarrays from OriGene Technologies, HPRT101 and HPRT102 ( ).

Techniques: Expressing, Immunocytochemistry, Staining, Transfection, Plasmid Preparation, Negative Control, Immunohistochemistry