Journal: Oncotarget

Article Title: FOXO4 expression is related to stem cell-like properties and resistance to treatment in diffuse large B-cell lymphoma

doi: 10.18632/oncotarget.13690

Figure Lengend Snippet: ( A ) Study scheme of gene expression analyses using doxorubicin (Dox) and phenylbutyrate (PB)-treated surviving cells: Four different cell lines (BJAB, Raji, Toledo, and Daudi) are treated with doxorubicin (300 nM) or phenylbutyrate (8 mM) for 48 h, and cDNA microarray analysis is done to identify differentially expressed target genes between treatment-surviving cells and parental control cells. ( B ) FOXO4 mRNA level is significantly higher in phenylbutyrate-treated surviving (PB) cells of BJAB, Raji and Daudi than control cells. ( C ) Vorinostat-treated surviving cells show higher mRNA level of FOXO4 than control (Con) cells of BJAB and Raji. ( D ) Primary lymphoma cells from three patients with refractory diffuse large B-cell lymphoma (DLBCL) show increased expression of FOXO4 in phenylbutyrate-treated surviving cells (PB) compared to the corresponding control cells. ( E ) The FOXO4 mRNA level is significantly higher in primary cells from four patients with refractory DLBCL than that of a patient with DLBCL who achieved complete response. Data represent means ± SD from three independent experiments.

Article Snippet: After endogenous peroxidase blocking for 5 min, tissues were incubated with primary polyclonal antibody against FOXO4 (1:1000; GTX50500; GeneTex, Irvine, CA, USA) for 15 min using a BOND-MAX autoimmunostainer (Leica Biosystems, Melbourne, Australia) for 15 min.

Techniques: Gene Expression, Microarray, Control, Expressing

Journal: Oncotarget

Article Title: FOXO4 expression is related to stem cell-like properties and resistance to treatment in diffuse large B-cell lymphoma

doi: 10.18632/oncotarget.13690

Figure Lengend Snippet: ( A, B ) Phenylbutyrate-treated surviving cells (BJAB-PB and Raji-PB) increase the expression of FOXO4 and its transcriptional targets (p21, p27, and SOD) compared to control cells. ( C, D ) BJAB-PB and Raji-PB cells show decreased expression of cyclin D1, CDK4, and cyclin A compared to control cells.

Article Snippet: After endogenous peroxidase blocking for 5 min, tissues were incubated with primary polyclonal antibody against FOXO4 (1:1000; GTX50500; GeneTex, Irvine, CA, USA) for 15 min using a BOND-MAX autoimmunostainer (Leica Biosystems, Melbourne, Australia) for 15 min.

Techniques: Expressing, Control

Journal: Oncotarget

Article Title: FOXO4 expression is related to stem cell-like properties and resistance to treatment in diffuse large B-cell lymphoma

doi: 10.18632/oncotarget.13690

Figure Lengend Snippet: ( A ) Soft agar colony formation in FOXO4-transfected or siFOXO4-transfected BJAB cells shows the amplification of FOXO4 increase colony forming ability and the knockdown of FOXO4 decrease colony formation in BJAB cell line. ( B ) Decrease in mRNA levels of Nanog, Oct-4 and Sox-2 in siFOXO4-transfected (siFOXO4) is noted compared to siControl-transfected (siCon) BJAB cells. ( C ) Phosphorylated AKT level is decreased in BJAB-PB cells with FOXO4 overexpression whereas siFOXO4-transfected BJAB-PB cells show the reverse of phosphorylated AKT. ( D ) The western blot shows the expression of FOXO4 protein in a BJAB clone (c2) overexpressing FOXO4. ( E ) Tumor sphere formation is observed from a BJAB clone (c2) overexpressing FOXO4. ( F ) Immunohistochemical staining for FOXO4 in tumor tissue of DLBCL (× 200). ( G ) Kaplan-Meier curves shows superior progression-free survival and overall survival of FOXO4-high group with diffuse large B cell lymphoma than FOXO4-low group. The P value is calculated using the log-rank test.

Article Snippet: After endogenous peroxidase blocking for 5 min, tissues were incubated with primary polyclonal antibody against FOXO4 (1:1000; GTX50500; GeneTex, Irvine, CA, USA) for 15 min using a BOND-MAX autoimmunostainer (Leica Biosystems, Melbourne, Australia) for 15 min.

Techniques: Transfection, Amplification, Knockdown, Over Expression, Western Blot, Expressing, Immunohistochemical staining, Staining

Journal: Oncotarget

Article Title: FOXO4 expression is related to stem cell-like properties and resistance to treatment in diffuse large B-cell lymphoma

doi: 10.18632/oncotarget.13690

Figure Lengend Snippet: Characteristics of patients

Article Snippet: After endogenous peroxidase blocking for 5 min, tissues were incubated with primary polyclonal antibody against FOXO4 (1:1000; GTX50500; GeneTex, Irvine, CA, USA) for 15 min using a BOND-MAX autoimmunostainer (Leica Biosystems, Melbourne, Australia) for 15 min.

Techniques:

Journal: Developmental cell

Article Title: Acute brain vascular regeneration occurs via lymphatic transdifferentiation.

doi: 10.1016/j.devcel.2021.09.005

Figure Lengend Snippet: Figure 2. The transdifferentiating iLVs switch molecular and structural identities and are restricted to stand-alone iLVs (A–C) The lyve1b+kdrl BLECs in the uninjured control (arrows), but not the transdifferentiating lyve1b+kdrl+ iLVs at 3 dpt after injury (arrowheads), were positive for anti-Prox1 (A) and fluorescence in situ hybridization (FISH)-vegfr3 (B). The statistics show the ratios of vessels positive for anti-Prox1 or FISH-vegfr3 among all the lyve1b+kdrl vessels and all the lyve1b+kdrl+ vessels. (C) (n = 6 larvae; two-way ANOVA by Dunnett’s multiple comparisons test; ***, p < 0.0001). Scale bar, 20 mm. (D and E) Positive control brain BVs in the uninjured larvae (D), the lineage-tracing system indicated that the vessels double positive for GFP and Dendra2 (E, arrow) and single positive for Dendra2 (D and E) expressed the blood-brain barrier marker glut1b, but vessels single positive for GFP (D, arrowhead) did not. Scale bar, 20 mm. (F–H) Single FIB-SEM image planes (right row) indicate cross sections of the vessels shown in the left row. Note that the mural of the DsRed+Dendra2+ vessel (G) is similar to the DsRed-Dendra2+ BV (H) but much thicker than the DsRed+Dendra2 iLV (F). Color rings mark the inner and outer surfaces of murals. Arrows indicate blood cells. Scale bars, 20 and 1 mm. (I–L) Live imaging shows BLECs (I), stand-alone iLVs (J, arrows), and track iLVs (K) at 2 and 4 dpt. Note that only the stand-alone iLVs express Dendra2 and recruit the GFP+ pericytes at 4 dpt (J, arrowheads). (L) The statistics show the ratios of transdifferentiation in the stand-alone iLVs and track iLVs at 4 dpt (n = 9 larvae; two-tailed unpaired t test; ***, p < 0.0001). Scale bar, 50 mm. (M–P) The TUNEL signals in the stand-alone iLVs (M) and track iLVs (N) at 7 dpt. The statistics show the ratios of TUNEL+ cells in stand-alone iLVs and in track iLVs (O) (n = 9 larvae; ***, p < 0.0001), and the ratios of iLVs undergoing transdifferentiation (Dendra2+GFP+) or undergoing apoptosis (GFP+TUNEL+) among all the iLVs at 7 dpt (P) (n = 30 larvae; two-tailed unpaired t test; ***, p < 0.0001). Scale bar, 50 mm. Data are represented as mean ± SD. See also Figure S2.

Article Snippet: Antibody staining, combination of FISH and antibody staining Whole-mount antibody staining and combination of FISH and antibody staining were performed as previously described (Chen et al., 2019; Liu et al., 2016; He et al., 2019, 2020) using antibodies against DsRed (1:200, Santa-Cruz), GFP or Venus (1:500, Abcam and Santa Cruz), Dendra2 (1:500, Antibody-online), Prox1 (1:1000, Abcam), and phospho-EphB4a (1:250, Signalway Antibody).

Techniques: Control, In Situ Hybridization, Positive Control, Marker, Imaging, Two Tailed Test, TUNEL Assay

Journal: Developmental cell

Article Title: Acute brain vascular regeneration occurs via lymphatic transdifferentiation.

doi: 10.1016/j.devcel.2021.09.005

Figure Lengend Snippet: Figure 4. Defective EphB4a leads to derepression of Notch in the track iLVs (A–L) Triple labeling of anti-Dendra2, anti-GFP, and FISH-dll4 (A–C)/notch1a (E–G)/hey1 (I–K). In the siblings, dll4/notch1a/hey1 were activated in the Dendra2+GFP+ stand-alone iLVs (A, E, and I) and Dendra2+GFP nascent BVs (B, F, and J), but not in the Dendra2GFP+ track iLVs (B, F, and J). By contrast, in the ephB4a mutant, the Dendra2GFP+ track iLVs at 3 dpt also exhibited dll4, notch1a, and hey1 expressions (C, G, and K). The statistics show the dll4/notch1a/ hey1 expression in the vessels of sibling and ephB4a mutant (D, H, and L) (n = 6 larvae; two-tailed unpaired t test; ***, p < 0.0001). (M–P) The Notch functional reporter tp1:Venus was activated in the Dendra2+DsRed+ stand-alone iLVs (M) and Dendra2+DsRed nascent BVs (N), but not in the Dendra2DsRed+ track iLVs (N). By contrast, in the ephB4a mutant, the Dendra2DsRed+ track iLVs at 3 dpt also exhibited Venus expression (O). The statistics show the tp1:Venus expression in the vessels of siblings and ephB4a mutants (P) (n = 6 larvae; two-tailed unpaired t test; ***, p < 0.0001). Scale bar, 20 mm. Data are represented as mean ± SD. See also Figures S6 and S7.

Article Snippet: Antibody staining, combination of FISH and antibody staining Whole-mount antibody staining and combination of FISH and antibody staining were performed as previously described (Chen et al., 2019; Liu et al., 2016; He et al., 2019, 2020) using antibodies against DsRed (1:200, Santa-Cruz), GFP or Venus (1:500, Abcam and Santa Cruz), Dendra2 (1:500, Antibody-online), Prox1 (1:1000, Abcam), and phospho-EphB4a (1:250, Signalway Antibody).

Techniques: Labeling, Mutagenesis, Expressing, Two Tailed Test, Functional Assay

Journal: Developmental cell

Article Title: Acute brain vascular regeneration occurs via lymphatic transdifferentiation.

doi: 10.1016/j.devcel.2021.09.005

Figure Lengend Snippet: Figure 5. EphB4a represses track iLV transdifferentiation through suppression of Notch (A–C) The derepressed transdifferentiation of track iLV (Dendra2+DsRed+) in the ephB4a mutant (A, arrow) was rescued by treatment with DAPT, which returned to Dendra2DsRed+ (B, arrowheads). The statistics show the ratios of transdifferentiation among track iLVs in the ephB4a mutant with or without DAPT treatment (C) (n = 10 larvae; two-tailed unpaired t test; ***, p < 0.0001). (D–G) The derepressed transdifferentiation of track iLV (Dendra2+GFP+) in the ephB4a mutant without heat shock (D, arrow) or without dnMAML-Flag (E, arrow) was rescued by the heat-shock-induced, LEC-specific overexpression of dnMAML-Flag, which exhibited Dendra2GFP+ at 4 dpt (F, arrowhead). The statistics show the ratios of transdifferentiation among track iLVs in the ephB4a mutant groups (G) (n = 10 larvae; two-tailed unpaired t test; ***, p < 0.0001). Scale bar, 20 mm. Data are represented as mean ± SD. HS, heat shock. See also Figures S6 and S7.

Article Snippet: Antibody staining, combination of FISH and antibody staining Whole-mount antibody staining and combination of FISH and antibody staining were performed as previously described (Chen et al., 2019; Liu et al., 2016; He et al., 2019, 2020) using antibodies against DsRed (1:200, Santa-Cruz), GFP or Venus (1:500, Abcam and Santa Cruz), Dendra2 (1:500, Antibody-online), Prox1 (1:1000, Abcam), and phospho-EphB4a (1:250, Signalway Antibody).

Techniques: Mutagenesis, Two Tailed Test, Over Expression

Journal: Oncotarget

Article Title: p53 regulates the transcription of the anti-inflammatory molecule developmental endothelial locus-1 (Del-1).

doi: 10.18632/oncotarget.1318

Figure Lengend Snippet: Figure 1: Schematic representation of the region upstream of the Del-1 gene. Putative p53 response elements (p53REs) were predicted using the Genometrix program. CATG-containing p53REs in the upstream regions are indicated. The first nucleotide of CATG is numbered relative to the translation start site (TSS) of the Del-1 gene.

Article Snippet: Membranes were blocked and probed with antibody against p53 (DO-1, Santa Cruz) or β-actin (Cell Signaling).

Techniques:

Journal: Oncotarget

Article Title: p53 regulates the transcription of the anti-inflammatory molecule developmental endothelial locus-1 (Del-1).

doi: 10.18632/oncotarget.1318

Figure Lengend Snippet: Figure 2: Relative transcriptional activity is dependent on the presence of proximal p53 response elements in the upstream region of the Del-1 gene. (A) Schematic diagram of the Del-1 promoter constructs. Nucleotides are indicated relative to the TSS. Upstream fragments of the Del-1 gene were cloned into the pGL3 vector to generate four Del- 1 promoter deletion constructs containing multiple putative p53 response elements. (B) Relative luciferase activity of these constructs. The Del-1 promoter constructs shown in (A) were individually transfected into HEK293T cells. Luciferase activity was determined 24 h after transfection and is expressed as the fold activity over that of the empty pGL3 vector (EV). A Renilla luciferase vector was co-transfected for normalization of transfection efficiency. Values are the means ± standard deviations (SD) from triplicate transfections. Data represent four independent experiments. **, p < 0.01; ***, p < 0.001; n.s., non-significant vs. the Del-1_luc 2k construct (i.e., the construct exhibiting the highest activity).

Article Snippet: Membranes were blocked and probed with antibody against p53 (DO-1, Santa Cruz) or β-actin (Cell Signaling).

Techniques: Activity Assay, Construct, Clone Assay, Plasmid Preparation, Luciferase, Transfection

Journal: Oncotarget

Article Title: p53 regulates the transcription of the anti-inflammatory molecule developmental endothelial locus-1 (Del-1).

doi: 10.18632/oncotarget.1318

Figure Lengend Snippet: Figure 3: Functional p53 response elements are required to enhance Del-1 transcription. (A) Schematic diagram of wild-type (WT) and mutant Del-1 promoter constructs. The Del-1_luc 2k construct was mutated at the consensus sequence (CATG→TATA) of either or both p53 response elements. (B) The WT and mutant Del-1_luc 2k constructs were independently transfected into HEK293T cells. Luciferase activity was determined 24 h after transfection and is expressed as the fold activity over that of the empty pGL3 vector (EV). Values are means ± SD from triplicate transfections. Data represent four independent experiments. *, p < 0.05; **, p < 0.01; n.s., non-significant vs. the WT Del-1_luc 2k construct.

Article Snippet: Membranes were blocked and probed with antibody against p53 (DO-1, Santa Cruz) or β-actin (Cell Signaling).

Techniques: Functional Assay, Mutagenesis, Construct, Sequencing, Transfection, Luciferase, Activity Assay, Plasmid Preparation

Journal: Oncotarget

Article Title: p53 regulates the transcription of the anti-inflammatory molecule developmental endothelial locus-1 (Del-1).

doi: 10.18632/oncotarget.1318

Figure Lengend Snippet: Figure 4: p53 positively regulates Del-1 transcription. (A) Mouse WT p53 or p53 with mutated DNA binding sites (G239A, R242A, and R243A) was transfected along with the 2k Del-1 promoter construct into HEK293T cells. Luciferase activity was determined 24 h after transfection and is expressed as the fold activity over that of the empty pGL3 vector. Values are means ± standard deviations from triplicate transfections. *, p < 0.05; ***, p < 0.001, vs. the 2k Del-1 promoter construct. (B) Mouse primary endothelial cells were treated with increasing concentrations of tenovin-1 and incubated for 24 h. The Del- 1 mRNA level was measured and is expressed as the fold increase over dimethyl sulfoxide (DMSO)-treated cells. Values are means ± SD from triplicate treatments. Data represent three independent experiments. *, p < 0.05; **, p < 0.01 vs. the DMSO-treated cells.

Article Snippet: Membranes were blocked and probed with antibody against p53 (DO-1, Santa Cruz) or β-actin (Cell Signaling).

Techniques: Binding Assay, Transfection, Construct, Luciferase, Activity Assay, Plasmid Preparation, Incubation

Journal: Oncotarget

Article Title: p53 regulates the transcription of the anti-inflammatory molecule developmental endothelial locus-1 (Del-1).

doi: 10.18632/oncotarget.1318

Figure Lengend Snippet: Figure 5: p53 directly binds to p53 response elements to enhance Del-1 transcription. (A) Representative ChIP analysis. HEK293T cells were transfected with a mouse p53- expressing plasmid, together with either the WT or mutant (mutated at both p53REs) 2k Del-1 promoter construct. Nuclear lysates were analyzed 48 h after transfection using a p53 antibody or control IgG and promoter regions containing the p53REs were amplified by PCR. (B) HEK293T cells were transfected with the plasmids in (A). Luciferase activity was determined 24 h after transfection and is expressed as the fold activity over that of the empty pGL3 vector. Values are means ± SD from triplicate transfections. Data represent three independent experiments. *, p < 0.05; ***, p < 0.001 vs. the cells co-transfected with the WT p53 and p53REwt Del-1 constructs.

Article Snippet: Membranes were blocked and probed with antibody against p53 (DO-1, Santa Cruz) or β-actin (Cell Signaling).

Techniques: Transfection, Expressing, Plasmid Preparation, Mutagenesis, Construct, Control, Amplification, Luciferase, Activity Assay

Journal: Oncotarget

Article Title: p53 regulates the transcription of the anti-inflammatory molecule developmental endothelial locus-1 (Del-1).

doi: 10.18632/oncotarget.1318

Figure Lengend Snippet: Figure 6: p53 levels determine Del-1 expression in mouse primary endothelial cells. Mouse p53 (A) and Del-1(B) mRNA levels were analyzed by real-time RT-PCR in primary endothelial cells from p53+/+, p53+/-, and p53-/- mice. The relative Del-1 mRNA levels in cells heterozygous or homozygous null for p53 were compared to those of p53+/+ mice. Values are means ± SD (n = 4–5 mice/group). Data represent three independent experiments. *, p < 0.05; **, p < 0.01; ***, p < 0.001.

Article Snippet: Membranes were blocked and probed with antibody against p53 (DO-1, Santa Cruz) or β-actin (Cell Signaling).

Techniques: Expressing, Quantitative RT-PCR

Journal: Oncotarget

Article Title: p53 regulates the transcription of the anti-inflammatory molecule developmental endothelial locus-1 (Del-1).

doi: 10.18632/oncotarget.1318

Figure Lengend Snippet: Figure 7: Del-1 reciprocally regulates p53 expression in mouse primary endothelial cells. (A) Levels of p53 protein was analyzed by western blotting in primary endothelial cells from Del-1+/+ and Del-1-/- mice. Values are means ± SD (n = 3 mice/group). (B) Transcript levels of p53 were analyzed by real-time RT-PCR. WT primary endothelial cells were incubated in the absence or presence of recombinant Del-1 protein (10 nM) for 24 h. Values are means ± SD (n = 4-5 mice/group). *, p < 0.05.

Article Snippet: Membranes were blocked and probed with antibody against p53 (DO-1, Santa Cruz) or β-actin (Cell Signaling).

Techniques: Expressing, Western Blot, Quantitative RT-PCR, Incubation, Recombinant

Journal:

Article Title: Latency-Associated Nuclear Antigen of Kaposi's Sarcoma-Associated Herpesvirus Up-Regulates Transcription of Human Telomerase Reverse Transcriptase Promoter through Interaction with Transcription Factor Sp1

doi: 10.1128/JVI.78.19.10348-10359.2004

Figure Lengend Snippet: LANA synergistically activates Sp1-mediated transcription. (A) GAL4-Sp1 activates transcription. HEK293 cells were cotransfected with the GAL4 DBD fused in frame with Sp1 and the pFR reporter plasmid containing the 5XGAL4 response element. Approximately 107 cells were transfected, and at 24 h posttransfection, the cells were harvested and lysed for luciferase assays. Increasing amounts of GAL4-Sp1 showed proportional increases in luciferase activity. Fractions of the cell lysates were resolved by SDS-PAGE to demonstrate the increased expression of GAL4-Sp1 in samples with larger amounts of transfected DNA. (B) LANA modulates GAL4-Sp1-mediated transcription. HEK293 cells were transfected with increasing amounts of LANA to show the effect of LANA on GAL4-Sp1-mediated transcription. LANA showed a dose-dependent response of GAL4-Sp1-mediated luciferase activity, which was plotted in relative luciferase units. (C and D) LANA modulates GAL4-mediated transcription in HEK293T and BJAB cells. Approximately 107 cells were transfected in both cases; at 24 h posttransfection, the luciferase activity was measured as described above. All of these experiments were done independently three times in duplicate, and the average values are presented in the figure. The increased expression of LANA was detected by use of an anti-Myc monoclonal antibody because LANA has a Myc epitope at its C terminus.

Article Snippet: The membrane was blocked with 5% nonfat dried milk and then incubated with a mouse anti-GAL4 DBD antibody (Santa Cruz Biotechnology Inc.) and anti-Myc ascites 9E10 to detect LANA expression.

Techniques: Plasmid Preparation, Transfection, Luciferase, Activity Assay, SDS Page, Expressing

Journal:

Article Title: Latency-Associated Nuclear Antigen of Kaposi's Sarcoma-Associated Herpesvirus Up-Regulates Transcription of Human Telomerase Reverse Transcriptase Promoter through Interaction with Transcription Factor Sp1

doi: 10.1128/JVI.78.19.10348-10359.2004

Figure Lengend Snippet: Mapping of functional domains of LANA and Sp1. (A) The B domain of Sp1 is enough for transcriptional activation by LANA. HEK293 cells were cotransfected with full-length (FL) GAL4-Sp1 and a vector containing either the A or the A and B domains in the presence of LANA. The number of relative luciferase units indicated that domain B is sufficient for the LANA-mediated transcription of luciferase. The GAL4 DBD-Sp1 fusion proteins used in the assay are shown below the bar diagram. (B) Different deletion mutants of LANA, shown below the bar diagram, were cotransfected into HEK293 cells (107) in the presence of GAL4-Sp1 and the pFR Luc reporter plasmid, and the luciferase activity was measured as described earlier after 24 h posttransfection. The N terminus of LANA (aa 1 to 340) showed an up-regulation in GAL4-mediated luciferase activity, whereas the C terminus itself did not show any effect and the C terminus fused to the N terminus had enhanced activation of the GAL4-Sp1-mediated luciferase activity, plotted in terms of relative luciferase activity. The data shown here are representative of three independent experiments done in triplicate.

Article Snippet: The membrane was blocked with 5% nonfat dried milk and then incubated with a mouse anti-GAL4 DBD antibody (Santa Cruz Biotechnology Inc.) and anti-Myc ascites 9E10 to detect LANA expression.

Techniques: Functional Assay, Activation Assay, Plasmid Preparation, Luciferase, Activity Assay

Journal: Carcinogenesis

Article Title: Vitamin D receptor status alters mammary gland morphology and tumorigenesis in MMTV-neu mice.

doi: 10.1093/carcin/bgh271

Figure Lengend Snippet: Fig. 1. VDR expression in mammary lesions and metastatic foci from MMTV-neu mice. Mammary tumors and lungs from MMTV-neu female mice (line N202, obtained from the Jackson Laboratory) were whole mounted to visualize preneoplastic lesions (A) (arrow) or formalin fixed, paraffin embedded and processed for H&E staining (C and E) and VDR immunostaining (brown staining) (B, D and F). Note VDR expression in early stage lesions (B), established tumors (D) and a metastatic focus (F) within the lung. Scale bars: (A), 200 mm; (B)--(D), 50 mm; (E) and (F), 100 mm. N, normal lung; M, metastatic focus. See online supplementary material for a colour version of this figure.

Article Snippet: Thoracic mammary glands or kidney (100 mg) were homogenized in Laemlli buffer containing phosphatase and protease inhibitors (34), separated by SDS--PAGE, transferred to nitrocellulose, blocked with 5% skimmed milk and immunoblotted with antibodies against VDR (clone C-20; Santa Cruz Biotechnology, Santa Cruz, CA) or c-neu (NCL-L-CBE-356; Vector Laboratories).

Techniques: Expressing, Formalin-fixed Paraffin-Embedded, Staining, Immunostaining

Journal: Carcinogenesis

Article Title: Vitamin D receptor status alters mammary gland morphology and tumorigenesis in MMTV-neu mice.

doi: 10.1093/carcin/bgh271

Figure Lengend Snippet: Fig. 2. Mammary gland morphology in 10.5-month-old MMTV-neu mice as a function of VDR genotype. Mammary ductal architecture (A, D and G), histopathology (B, E and H) and pre-neoplastic lesions (C, F and I) in 10.5-month-old MMTV-neu littermates wild-type for VDR (neu/VDRþ/þ, top panels) or with germline inactivation of one (neu/VDRþ/, middle panels) or two (neu/VDR/, bottom panels) alleles of the VDR gene. Inguinal glands from neu/VDRþ/þ

Article Snippet: Thoracic mammary glands or kidney (100 mg) were homogenized in Laemlli buffer containing phosphatase and protease inhibitors (34), separated by SDS--PAGE, transferred to nitrocellulose, blocked with 5% skimmed milk and immunoblotted with antibodies against VDR (clone C-20; Santa Cruz Biotechnology, Santa Cruz, CA) or c-neu (NCL-L-CBE-356; Vector Laboratories).

Techniques: Histopathology

Journal: Carcinogenesis

Article Title: Vitamin D receptor status alters mammary gland morphology and tumorigenesis in MMTV-neu mice.

doi: 10.1093/carcin/bgh271

Figure Lengend Snippet: Fig. 3. Effect of VDR status on health and survival of MMTV-neu mice. (A) Kaplan--Meier analysis of survival in MMTV-neu mice by VDR genotype. Female MMTV-neu littermates wild-type for VDR (neu/VDRþ/þ) or with germline inactivation of one (neu/VDRþ/) or two (neu/VDR/) alleles of the VDR gene were monitored for up to 18.5 months until they reached criteria for killing or died. Squares, neu/VDRþ/þ mice (n ¼ 63); circles, neu/VDRþ/ mice (n ¼ 144); triangles, neu/VDR/ mice (n ¼ 72). Arrows indicate mean survival for each group. Survival of neu/VDR/

Article Snippet: Thoracic mammary glands or kidney (100 mg) were homogenized in Laemlli buffer containing phosphatase and protease inhibitors (34), separated by SDS--PAGE, transferred to nitrocellulose, blocked with 5% skimmed milk and immunoblotted with antibodies against VDR (clone C-20; Santa Cruz Biotechnology, Santa Cruz, CA) or c-neu (NCL-L-CBE-356; Vector Laboratories).

Techniques:

Journal: Carcinogenesis

Article Title: Vitamin D receptor status alters mammary gland morphology and tumorigenesis in MMTV-neu mice.

doi: 10.1093/carcin/bgh271

Figure Lengend Snippet: Fig. 4. Pathology of mammary glands in MMTV-neu mice as a function of VDR. (A--C) Post-mortem examination of inguinal mammary gland. The mammary fat pad was photographed in neu/VDRþ/þ (A), neu/VDRþ/ (B) and neu/VDR/ (C) mice at 15 months of age. A white line is drawn around the perimeter of the fat pad and the location of the lymph node is indicated. Note atrophy of the mammary fat pad and abnormal skin in neu/VDR/ mice compared with neu/VDRþ/þ and neu/VDRþ/ littermates. (D--F) Ductal architecture. Representative micrographs of inguinal whole mounts from neu/VDRþ/þ (D), neu/VDRþ/

Article Snippet: Thoracic mammary glands or kidney (100 mg) were homogenized in Laemlli buffer containing phosphatase and protease inhibitors (34), separated by SDS--PAGE, transferred to nitrocellulose, blocked with 5% skimmed milk and immunoblotted with antibodies against VDR (clone C-20; Santa Cruz Biotechnology, Santa Cruz, CA) or c-neu (NCL-L-CBE-356; Vector Laboratories).

Techniques:

Journal: Carcinogenesis

Article Title: Vitamin D receptor status alters mammary gland morphology and tumorigenesis in MMTV-neu mice.

doi: 10.1093/carcin/bgh271

Figure Lengend Snippet: Fig. 5. Serum estrogen in MMTV-neu mice as a function of VDR genotype. 17b-Estradiol was measured by radioimmunoassay in serum from neu/ VDRþ/þ, neu/VDRþ/ and neu/VDR/ mice at the indicated ages. a, P 5 0.05, neu/VDRþ/þ versus neu/VDR/; b, P 5 0.05, neu/VDRþ/þ or neu/VDRþ/ versus neu/VDR/. No differences in serum estradiol were detected between neu/VDRþ/þ mice and neu/VDRþ/ mice. Data are means SE of 6--8 mice per genotype.

Article Snippet: Thoracic mammary glands or kidney (100 mg) were homogenized in Laemlli buffer containing phosphatase and protease inhibitors (34), separated by SDS--PAGE, transferred to nitrocellulose, blocked with 5% skimmed milk and immunoblotted with antibodies against VDR (clone C-20; Santa Cruz Biotechnology, Santa Cruz, CA) or c-neu (NCL-L-CBE-356; Vector Laboratories).

Techniques: RIA Assay

Journal: Carcinogenesis

Article Title: Vitamin D receptor status alters mammary gland morphology and tumorigenesis in MMTV-neu mice.

doi: 10.1093/carcin/bgh271

Figure Lengend Snippet: Fig. 6. Kinetics of mammary tumor appearance in neu/VDRþ/þ and neu/ VDRþ/ mice. Kaplan--Meier analysis of tumor development in nulliparous female MMTV-neu littermates wild-type for VDR (neu/VDRþ/þ) or with germline inactivation of one (neu/VDRþ/) allele of the VDR gene. Mice were monitored for mammary tumor development by palpation for up to 18.5 months. Squares, neu/VDRþ/þ mice (n ¼ 63); circles, neu/VDRþ/

Article Snippet: Thoracic mammary glands or kidney (100 mg) were homogenized in Laemlli buffer containing phosphatase and protease inhibitors (34), separated by SDS--PAGE, transferred to nitrocellulose, blocked with 5% skimmed milk and immunoblotted with antibodies against VDR (clone C-20; Santa Cruz Biotechnology, Santa Cruz, CA) or c-neu (NCL-L-CBE-356; Vector Laboratories).

Techniques:

Journal: Carcinogenesis

Article Title: Vitamin D receptor status alters mammary gland morphology and tumorigenesis in MMTV-neu mice.

doi: 10.1093/carcin/bgh271

Figure Lengend Snippet: Fig. 7. Histopathology and neu expression in primary tumors from MMTV-neu mice as a function of VDR status. (Left) Representative H&E stained sections of primary mammary tumors from neu/VDRþ/þ (A), neu/VDRþ/ (C) and neu/VDR/ (E) littermates. (Right) Immunohistochemistry for c-neu in mammary tumors from neu/VDRþ/þ (B), neu/VDRþ/ (D) and neu/VDR/ (F) mice. No differences in morphology or c-neu staining were detected as a function of VDR status. Scale bar: 50 mM. (G) Western blot for c-neu in tumors from neu/VDRþ/þ, neu/VDRþ/ and neu/VDR/ mice. Tumor homogenates were separated by SDS--PAGE, transferred to nitrocellulose and incubated with antibody against c-neu. The blot shows two independent tumor homogenates from both neu/VDRþ/þ

Article Snippet: Thoracic mammary glands or kidney (100 mg) were homogenized in Laemlli buffer containing phosphatase and protease inhibitors (34), separated by SDS--PAGE, transferred to nitrocellulose, blocked with 5% skimmed milk and immunoblotted with antibodies against VDR (clone C-20; Santa Cruz Biotechnology, Santa Cruz, CA) or c-neu (NCL-L-CBE-356; Vector Laboratories).

Techniques: Histopathology, Expressing, Staining, Immunohistochemistry, Western Blot, SDS Page, Incubation

Journal: Carcinogenesis

Article Title: Vitamin D receptor status alters mammary gland morphology and tumorigenesis in MMTV-neu mice.

doi: 10.1093/carcin/bgh271

Figure Lengend Snippet: Fig. 8. VDR expression in tumors from neu/VDRþ/þ, neu/VDRþ/ and neu/VDR/ littermates. Tumors were evaluated for VDR expression by quantitative real time PCR (A), western blotting (B) and immunohistochemistry (C). Data in (A) are expressed as means SE of 6 values per genotype. ND, not detectable. In (B) tumor and kidney homogenates were separated by SDS--PAGE and immunoblotted with a polyclonal antibody directed against VDR (clone C-20). The blot shows two independent tumor homogenates from both neu/VDRþ/þ and neu/VDRþ/ mice and one homogenate from a neu/VDR/

Article Snippet: Thoracic mammary glands or kidney (100 mg) were homogenized in Laemlli buffer containing phosphatase and protease inhibitors (34), separated by SDS--PAGE, transferred to nitrocellulose, blocked with 5% skimmed milk and immunoblotted with antibodies against VDR (clone C-20; Santa Cruz Biotechnology, Santa Cruz, CA) or c-neu (NCL-L-CBE-356; Vector Laboratories).

Techniques: Expressing, Real-time Polymerase Chain Reaction, Western Blot, Immunohistochemistry, SDS Page

Journal: Molecular & Cellular Proteomics : MCP

Article Title: Parvulin (Par14), a Peptidyl-Prolyl cis - trans Isomerase, Is a Novel rRNA Processing Factor That Evolved in the Metazoan Lineage *

doi: 10.1074/mcp.M900147-MCP200

Figure Lengend Snippet: Par14-associated trans-acting factors putatively involved in ribosome biogenesis Probable trans-acting factors identified in Par14-associated pre-rRNP complexes are shown. Trans-acting factors involved in ribosome biogenesis are classified into functional groups. For proteins having human and yeast orthologs, the gene names are indicated (obtained by Blink analysis of the NCBI database). Proteins were identified by either LC-MS/MS or MALDI-TOF/MS combined with LC-MS/MS as described in supplemental Tables 1 and 4 and Ref. 12 . NCBI accession numbers (GI no.) are shown. Involvement of yeast orthologs in the preribosomal complexes is shown (27). snoRNP, small nucleolar ribonucleoprotein; Brix, biogenesis of ribosome in Xenopus ; TGF, transforming growth factor; FHA, forkhead-associated; Chr, chromosome; BRCT, BRCA1 carboxyl terminus; RNPs, ribonucleoproteins.

Article Snippet: Antibodies against B23, fibrillarin, and nucleolin were obtained from Santa Cruz Biotechnology (Santa Cruz, CA).

Techniques: Functional Assay, RNA modification, Blocking Assay, RNA Binding Assay

Journal: Scientific Reports

Article Title: A Recombinant Secondary Antibody Mimic as a Target-specific Signal Amplifier and an Antibody Immobilizer in Immunoassays

doi: 10.1038/srep24159

Figure Lengend Snippet: ( A ) Scheme showing different combinations of primary antibodies with either HRP-conjugated secondary antibodies or HRP-GST-ABD. ( B – E ) BSA ( B , C ) or EpCAM ( D , E ) are immobilized on the surface of the ELISA plates and various concentrations of either rabbit anti-BSA ( B ) or mouse anti-BSA primary antibodies ( C ) or either rabbit anti-EpCAM ( D ) or mouse anti-EpCAM primary antibodies ( E ) are applied. Linear responses of each measurement are plotted as insets of each graph.

Article Snippet: After blocking, the solutions of primary antibodies against BSA (rabbit antibodies purchased from Santa Cruz Biotech and mouse antibodies from Abcam) or EpCAM (rabbit and mouse antibodies purchased from Sino Biological Inc.) were prepared with the PBS buffer by using serial dilutions.

Techniques: Enzyme-linked Immunosorbent Assay

Journal: Scientific Reports

Article Title: A Recombinant Secondary Antibody Mimic as a Target-specific Signal Amplifier and an Antibody Immobilizer in Immunoassays

doi: 10.1038/srep24159

Figure Lengend Snippet: ( A ) Scheme showing the adaptation of GST-ABD as an anchoring adaptor for antigen-capturing antibodies. ( B , C ) GST-ABDs are spread on the surface of GSH-coated plates and saturated with capturing antibodies, either anti-BSA rabbit IgGs ( B ) or anti-EpCAM rabbit IgGs ( C ). Various amounts of BSA ( B ) or EpCAM ( C ) are added. Linear responses for each measurement are plotted as insets of each graph.

Article Snippet: After blocking, the solutions of primary antibodies against BSA (rabbit antibodies purchased from Santa Cruz Biotech and mouse antibodies from Abcam) or EpCAM (rabbit and mouse antibodies purchased from Sino Biological Inc.) were prepared with the PBS buffer by using serial dilutions.

Techniques: