Journal: Communications biology

Article Title: Chemosensitization of prostate cancer stem cells in mice by angiogenin and plexin-B2 inhibitors.

doi: 10.1038/s42003-020-0750-6

Figure Lengend Snippet: Fig. 9 3′-end fragment of 5S rRNA enhances stemness of P19 cells. a Small RNA in P19 cells treated with various concentration of ANG for 2 h. A total of 20 μg RNA was loaded on each lane. The intensity of small RNA bands was calculated by Image J and normalized to untreated control (n = 4). b Northern blot analysis of the 3′-end fragment of 5S rRNA in P19 cells treated with PBS or ANG (1 μg/ml) for 2 h. The intensity of the band around 45 nt was determined by Image J and normalized to PBS control. c In vitro proliferation of P19 cells transfected with scramble RNA, intact 5S rRNA, the 5′- or 3′-end fragment of 5S rRNA (n = 2). d Protein synthesis rate of stable P19 transfectants of various small RNA determined by 3[H] methionine incorporation (n = 3). e Sphere formation of stable P19 transfectants of various small RNA. Sphere numbers were counted from the entire well of 24-well plate containing 100,000 cells cultured in sphere-forming medium for 24 h (n = 2). Scale bar: 500 μm. f Immunoblots of Bcl-2, cleaved Parp-1, and AIF in stable P19 transfectants of various small RNA cultured in sphere-forming medium. Actin was used as loading control in each blot. *p < 0.05; **p < 0.01; ***p < 0.001; ns not significant.

Article Snippet: Human prostate cancer cell lines PC3 (male), DU145 (male), and LNCaP (male), mouse embryonal carcinoma cell line P19 (male) were purchased from ATCC.

Techniques: Concentration Assay, Control, Northern Blot, In Vitro, Transfection, Cell Culture, Western Blot

Journal: eLife

Article Title: KRAB-zinc finger protein gene expansion in response to active retrotransposons in the murine lineage

doi: 10.7554/eLife.56337

Figure Lengend Snippet: ( A ) Differential analysis of summative (all individual insertions combined) H3K9me3 enrichment at TE groups in Chr10-cl, Chr13.1-cl and Chr13.2-cl KO ES cells. TE groups targeted by one or several KRAB-ZFPs encoded within the deleted clusters are highlighted in orange (differential enrichment over the entire TE sequences) and red (differential enrichment at TE regions that overlap with KRAB-ZFP ChIP-seq peaks). ( B ) Top: Schematic view of the Cd59a / Cd59b locus with a 5’ truncated ETn insertion. ChIP-seq (Input subtracted from ChIP) data for overexpressed epitope-tagged Gm13051 (a Chr4-cl KRAB-ZFP) in F9 EC cells, and re-mapped KAP1 (GEO accession: GSM1406445) and H3K9me3 (GEO accession: GSM1327148) in WT ES cells are shown together with RNA-seq data from Chr4-cl WT and KO ES cells (mapped using Bowtie (-a -m 1 -- strata -v 2) to exclude reads that cannot be uniquely mapped). Bottom: Transcriptional activity of a 5 kb fragment with or without fragments of the ETn insertion was tested by luciferase reporter assay in Chr4-cl WT and KO ES cells.

Article Snippet: Cell line ( Mus musculus ) , F9 Embryonic carcinoma cells , ATCC , ATCC CRL-1720 , .

Techniques: ChIP-sequencing, RNA Sequencing, Activity Assay, Luciferase, Reporter Assay

Journal: eLife

Article Title: KRAB-zinc finger protein gene expansion in response to active retrotransposons in the murine lineage

doi: 10.7554/eLife.56337

Figure Lengend Snippet: ( A ) Differential gene expression in Chr2-cl and Chr4-cl KO ES cells. Significantly up- and downregulated genes ( adjusted p-value <0.05) are highlighted in red and green, respectively, KRAB-ZFP genes within the deleted clusters are shown in blue. ( B ) Correlation of TEs and gene deregulation. Plots show enrichment of TE groups within 100 kb of up- and downregulated genes relative to all genes. Significantly overrepresented LTR and LINE groups ( adjusted p-value <0.1) are highlighted in blue and red, respectively. ( C ) Schematic view of the downstream region of Chst1 where a 5’ truncated ETn insertion is located. ChIP-seq (Input subtracted from ChIP) data for overexpressed epitope-tagged Gm13051 (a Chr4-cl KRAB-ZFP) in F9 EC cells, and re-mapped KAP1 (GEO accession: GSM1406445) and H3K9me3 (GEO accession: GSM1327148) in WT ES cells are shown together with RNA-seq data from Chr4-cl WT and KO ES cells (mapped using Bowtie (-a -m 1 -- strata -v 2) to exclude reads that cannot be uniquely mapped). ( D ) RT-qPCR analysis of Chst1 mRNA expression in Chr4-cl WT and KO ES cells with or without the CRISPR/Cas9 deleted ETn insertion near Chst1 . Values represent mean expression (normalized to Gapdh) from three biological replicates per sample (each performed in three technical replicates) in arbitrary units. Error bars represent standard deviation and asterisks indicate significance (p<0.01, Student’s t-test). n.s.: not significant. ( E ) Mean coverage of ChIP-seq data (Input subtracted from ChIP) in Chr4-cl WT and KO ES cells over 127 full-length ETn insertions. The binding sites of the Chr4-cl KRAB-ZFPs Rex2 and Gm13051 are indicated by dashed lines.

Article Snippet: Cell line ( Mus musculus ) , F9 Embryonic carcinoma cells , ATCC , ATCC CRL-1720 , .

Techniques: Gene Expression, ChIP-sequencing, RNA Sequencing, Quantitative RT-PCR, Expressing, CRISPR, Standard Deviation, Binding Assay

Journal: eLife

Article Title: KRAB-zinc finger protein gene expansion in response to active retrotransposons in the murine lineage

doi: 10.7554/eLife.56337

Figure Lengend Snippet:

Article Snippet: Cell line ( Mus musculus ) , F9 Embryonic carcinoma cells , ATCC , ATCC CRL-1720 , .

Techniques: Recombinant, Expressing, Plasmid Preparation, CRISPR, Construct, Sequencing, Software

Journal: Toxicological sciences : an official journal of the Society of Toxicology

Article Title: Valproic acid-induced deregulation in vitro of genes associated in vivo with neural tube defects.

doi: 10.1093/toxsci/kfp002

Figure Lengend Snippet: FIG. 1. Effect of VPA on (A) proliferation and viability and (B) apoptosis in P19 embryonal carcinoma cells after 24 h of VPA exposure. (A) The dashed blue line represents the number of cells (760,000 ± 50,000 SD) present at the start of exposure. (B) The fold change of cleaved caspase-3 in P19 cells exposed to 1, 2.5, 5, and 10mM VPA, respectively, versus controls. Bars represent the data range (n ¼ 2).

Article Snippet: P19 mouse embryonal carcinoma cells (ATCC CRL-1825; American Type Culture Collection, Manassas, VA) were cultured at 37 C and 5% CO2 in Dulbecco’s Modified Eagle’s Medium (Swedish National Veterinary Institute, Uppsala, Sweden), supplemented with 10% fetal bovine serum (Swedish National Veterinary Institute), 2mM L-glutamine, 100 U/ml penicillin, and 100 lg/ml streptavidin.

Techniques:

Journal: Toxicological sciences : an official journal of the Society of Toxicology

Article Title: Valproic acid-induced deregulation in vitro of genes associated in vivo with neural tube defects.

doi: 10.1093/toxsci/kfp002

Figure Lengend Snippet: FIG. 2. Gene expression changes in P19 embryonal carcinoma cells after 1.5, 6, and 24-h exposure to 1 and 2.5mM VPA. (A–E) Venn diagrams showing the number of array elements (oligonucleotide probes) showing a significant VPA response in the F-test at p value p < 103 (A), p < 104 (B), p < 104 (C), p < 104 (D), and p < 103 (E), respectively. The total number of significant probes and their percentage among 15,308 array elements are given for each cut-off level. (F) Venn diagram showing a comparison between the number of probes showing a significant (p < 103) VPA response in the F-test and the analysis based on average fold change (Mav) across all arrays per time point (see ‘‘Materials and methods’’).

Article Snippet: P19 mouse embryonal carcinoma cells (ATCC CRL-1825; American Type Culture Collection, Manassas, VA) were cultured at 37 C and 5% CO2 in Dulbecco’s Modified Eagle’s Medium (Swedish National Veterinary Institute, Uppsala, Sweden), supplemented with 10% fetal bovine serum (Swedish National Veterinary Institute), 2mM L-glutamine, 100 U/ml penicillin, and 100 lg/ml streptavidin.

Techniques: Gene Expression, Comparison

Journal: Toxicological sciences : an official journal of the Society of Toxicology

Article Title: Valproic acid-induced deregulation in vitro of genes associated in vivo with neural tube defects.

doi: 10.1093/toxsci/kfp002

Figure Lengend Snippet: FIG. 5. Cross-platform comparison of microarray data. Log2 fold changes determined in NIA clone set cDNA microarrays for array elements previously (Kultima et al., 2004) found to show significant transcriptional response to VPA in (A) P19 cells exposed to VPA for 1mM for 24 h, and (B) embryos collected 6 h after ip injection of 600 mg/kg bodyweight on gestational day 8 in the NMRI mouse, are plotted against log2 fold changes determined in CodeLink Expression Bioarrays for P19 cells exposed to VPA for 1mM for 24 h.

Article Snippet: P19 mouse embryonal carcinoma cells (ATCC CRL-1825; American Type Culture Collection, Manassas, VA) were cultured at 37 C and 5% CO2 in Dulbecco’s Modified Eagle’s Medium (Swedish National Veterinary Institute, Uppsala, Sweden), supplemented with 10% fetal bovine serum (Swedish National Veterinary Institute), 2mM L-glutamine, 100 U/ml penicillin, and 100 lg/ml streptavidin.

Techniques: Comparison, Microarray, Injection, Expressing

Journal: BMC Pharmacology & Toxicology

Article Title: Comparison of neurons derived from mouse P19, rat PC12 and human SH-SY5Y cells in the assessment of chemical- and toxin-induced neurotoxicity

doi: 10.1186/s40360-017-0151-8

Figure Lengend Snippet: Development of neurons derived from RA-treated P19 and SH-SY5Y cells, and NGF-stimulated PC12 cells up to 10 days in culture. The cells were plated at a density of 500 cells/mm 2 and immunostained against the neuron-specific protein βIII-tubulin. a Representative fluorescence microscopy images of neurons (20 × magnification). b Fluorescence of anti-βIII-tubulin antibodies measured in a microplate reader and expressed as relative fluorescence units (RFU). Data are means ± SEM of 3–4 independent experiments

Article Snippet: P19 mouse embryonal carcinoma cells (cat. no. ECACC 95102107), PC12 rat adrenal pheochromocytoma cells (cat. no. ECACC 88022401) and SH-SY5Y human neuroblastoma cells (cat. no. ECACC 94030304) were obtained from European Collection of Authenticated Cell Cultures (ECACC) (Porton Down, UK).

Techniques: Derivative Assay, Fluorescence, Microscopy

Journal: BMC Pharmacology & Toxicology

Article Title: Comparison of neurons derived from mouse P19, rat PC12 and human SH-SY5Y cells in the assessment of chemical- and toxin-induced neurotoxicity

doi: 10.1186/s40360-017-0151-8

Figure Lengend Snippet: Concentration-dependent effects of MeHg, okadaic acid and acrylamide on cell viability and expression of the neuron-specific protein βIII-tubulin in neuronally differentiated P19, PC12 and SH-SY5Y cells. The cells were plated at a density of 500 cells/mm 2 and cultured for 6 days in the differentiation media, followed by exposure to the test compounds for 48 h. Effects of MeHg ( a , b ), okadaic acid ( c , d ) and acrylamide ( e , f ) on the cell viability were assessed by using the calcein-AM assay ( a , c , e ), and the immunofluorescence of βIII-tubulin ( b , d , f ). The data are means ± SEM of n = 6 independent experiments ( n = 3 for okadaic acid in the βIII-tubulin assay; panel d ). The results are expressed as percentage of non-treated cells or cells treated with 0.1% DMSO (used as vehicle). Wells treated with 2% Triton X-100 for 30 min served as controls for maximal cell death. Statistical analysis was performed using repeated measures one-way ANOVA with post hoc Dunnett’s multiple comparisons test (* p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001) compared to corresponding controls

Article Snippet: P19 mouse embryonal carcinoma cells (cat. no. ECACC 95102107), PC12 rat adrenal pheochromocytoma cells (cat. no. ECACC 88022401) and SH-SY5Y human neuroblastoma cells (cat. no. ECACC 94030304) were obtained from European Collection of Authenticated Cell Cultures (ECACC) (Porton Down, UK).

Techniques: Concentration Assay, Expressing, Cell Culture, Calcein AM Assay, Immunofluorescence

Journal: BMC Pharmacology & Toxicology

Article Title: Comparison of neurons derived from mouse P19, rat PC12 and human SH-SY5Y cells in the assessment of chemical- and toxin-induced neurotoxicity

doi: 10.1186/s40360-017-0151-8

Figure Lengend Snippet: Representative fluorescence microscopy images of neuronally differentiated P19, PC12 and SH-SY5Y cells exposed to 1 μM methylmercury, 10 nM okadaic acid and 1 mM acrylamide. The cells were plated at a density of 500 cells/mm 2 and cultured for 6 days in the differentiation media, followed by exposure to the test compounds for 48 h. The cells were immunolabeled against the neuron-specific protein βIII-tubulin and the fluorescence microscopy images were obtained at 20 × magnification

Article Snippet: P19 mouse embryonal carcinoma cells (cat. no. ECACC 95102107), PC12 rat adrenal pheochromocytoma cells (cat. no. ECACC 88022401) and SH-SY5Y human neuroblastoma cells (cat. no. ECACC 94030304) were obtained from European Collection of Authenticated Cell Cultures (ECACC) (Porton Down, UK).

Techniques: Fluorescence, Microscopy, Cell Culture, Immunolabeling

Journal: BMC Pharmacology & Toxicology

Article Title: Comparison of neurons derived from mouse P19, rat PC12 and human SH-SY5Y cells in the assessment of chemical- and toxin-induced neurotoxicity

doi: 10.1186/s40360-017-0151-8

Figure Lengend Snippet: Effects of MeHg, BSO and GSH on the viability of RA-treated P19 cells ( a ) and SH-SY5Y cells ( b ). Cells cultured for 6 days in the differentiation media were pre-treated with 100 μM BSO for 17 h or 1 mM GSH for 1 h followed by exposure to 1 μM MeHg for 24 h. Cell viability was assessed with the PrestoBlue assay that measures cellular metabolic reduction, and extracellular LDH activity assay. Data are means ± SEM of n = 6 independent experiments. For the PrestoBlue assay, data are expressed as percentage of non-treated or 0.1% DMSO vehicle-treated cells. For the LDH assay, the data are presented as percentage of total cell death (cells treated with 2% Triton X-100). Statistical analysis was undertaken using one-way ANOVA with post hoc Bonferroni’s multiple comparisons test: * p < 0.05, ** p < 0.01, and **** p < 0.0001 (the effect of the treatment compared to the corresponding vehicle control, or the effect of the combination of MeHg + GSH or MeHg + BSO compared to MeHg per se ). ND = not determined

Article Snippet: P19 mouse embryonal carcinoma cells (cat. no. ECACC 95102107), PC12 rat adrenal pheochromocytoma cells (cat. no. ECACC 88022401) and SH-SY5Y human neuroblastoma cells (cat. no. ECACC 94030304) were obtained from European Collection of Authenticated Cell Cultures (ECACC) (Porton Down, UK).

Techniques: Cell Culture, Prestoblue Assay, Activity Assay, Lactate Dehydrogenase Assay, Control

Journal: BMC Pharmacology & Toxicology

Article Title: Comparison of neurons derived from mouse P19, rat PC12 and human SH-SY5Y cells in the assessment of chemical- and toxin-induced neurotoxicity

doi: 10.1186/s40360-017-0151-8

Figure Lengend Snippet: The effects of MeHg, GSH and BSO on TMRE fluorescence in RA-treated P19 ( a ) and SH-SY5Y cells ( b ). Cells cultured for 6 days in the differentiation media were pre-treated with 100 μM BSO for 17 h or with 1 mM GSH for 1 h followed by exposure to 1 μM MeHg for 24 h. Alterations in mitochondrial membrane potential were measured with the TMRE assay. Data are means ± SEM of n = 6 independent experiments, and expressed as percentage of non-treated or 0.1% DMSO vehicle-treated cells. Statistical analysis was undertaken using one-way ANOVA with post hoc Bonferroni’s multiple comparisons test: * p <0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001 (the effect of the treatment compared to the corresponding vehicle control, or the effect of the combination of MeHg + GSH or MeHg + BSO compared to MeHg per se )

Article Snippet: P19 mouse embryonal carcinoma cells (cat. no. ECACC 95102107), PC12 rat adrenal pheochromocytoma cells (cat. no. ECACC 88022401) and SH-SY5Y human neuroblastoma cells (cat. no. ECACC 94030304) were obtained from European Collection of Authenticated Cell Cultures (ECACC) (Porton Down, UK).

Techniques: Fluorescence, Cell Culture, Membrane, Control

Journal: Biomolecules

Article Title: Exogenous Selenoprotein V Induces Apoptosis in Murine Testicular Teratoma Cells via Mitochondrial Dysfunction and ROS Overproduction

doi: 10.3390/biom15121733

Figure Lengend Snippet: Effects of 24-h pre-incubation of F9 cells with protein solvent buffer or various concentrations of recombinant SELENOV on cell viability. ( A ) Viability cytogram of F9 cells following treatment with 100 μL/mL (equivalent to 50 μg/mL SELENOV application) or 200 μL/mL (equivalent to 100 μg/mL SELENOV application) of solvent buffer. The 10 μg/mL SELENOV group was compared to the untreated control, as the corresponding buffer volume was negligible. ( B ) Viability cytogram of F9 cells after treatment with different concentrations of recombinant SELENOV. ( C ) Effects of solvent buffer or recombinant SELENOV on cell viability and activation of cell death pathways. Data are presented as mean ± SEM. Statistical significance was determined by one-way ANOVA followed by Tukey’s post hoc test. *** p < 0.001 versus the Control group; ### p < 0.001 versus the corresponding Buffer group.

Article Snippet: The study utilized several established cell lines: F9 cells (a murine embryonal carcinoma model derived from testicular teratoma of C57BL/6 mice) and mouse fibroblasts (L-929), all obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA).

Techniques: Incubation, Solvent, Recombinant, Control, Activation Assay