Journal: The Journal of clinical endocrinology and metabolism

Article Title: HOXA10, Pbx2, and Meis1 protein expression in the human endometrium: formation of multimeric complexes on HOXA10 target genes.

doi: 10.1210/jc.2004-0817

Figure Lengend Snippet: FIG. 4. Meis1 is absent and Pbx2 is present in glandular epithelial Ishikawa cells. A, MEG-01 cell extract was used as a positive control for Meis1. Meis1 is absent in the Ishikawa cell extract. Goat IgG was used as a negative control antibody. B, BJAB cell extract was used as a positive control for Pbx2. Pbx2 is present in the Ishikawa cell extract. Rabbit IgG was used as a negative control antibody.

Article Snippet: BJAB cell lysate was used as a positive control for Pbx2 and megakaryoblastic cell lysate (Santa Cruz Biotechnology) was used as a positive control for Meis1.

Techniques: Positive Control, Negative Control

Journal: The Journal of Clinical Investigation

Article Title: Evolutionary conservation of human ketodeoxynonulosonic acid production is independent of sialoglycan biosynthesis

doi: 10.1172/JCI137681

Figure Lengend Snippet: A continuous cell viability assay (0–84 hours) was performed to evaluate mannose toxicity to various human and mouse cell lines including mutants of Kdn-associated metabolic enzymes. The mannose feeding concentration was set at 0–200 mM, and the same concentration of GlcNAc was used as a control for the hyperosmolarity effect. Results shown are for (A and B) HEK293A cells; (C and D) HUVECs; (E and F) BJAB K88 cells (GNE WT); (G and H) BJAB K20 cells (GNE-KO); (I and J) PMI WT cells; and (K and L) PMI-KO cells. (L) PMI-KO cells were always maintained in mannose-supplemented (25 μM) media for their survival during GlcNAC feeding. Data are representative of 2 independent experiments.

Article Snippet: HEK293A cells (Thermo Fisher Scientific) were cultivated in RPMI 1640 media with 10% FBS; HUVECs (American Type Culture Collection [ATCC]) in EBM2 media (Lonza); BJAB K20 and BJAB K88 cells (provided by Stephan Hinderlich and the late Werner Reutter, Beuth University of Applied Sciences, Berlin, Germany) in RPMI 1640 media with 10% FBS; PMI WT and PMI-KO cells in DMEM containing 1 g/L glucose and 10% FBS; and HK2 cells in keratinocyte serum-free media supplemented with bovine pituitary extract and recombinant EGF (all from Gibco, Thermo Fisher Scientific).

Techniques: Viability Assay, Concentration Assay, Control

Journal: The Journal of Clinical Investigation

Article Title: Evolutionary conservation of human ketodeoxynonulosonic acid production is independent of sialoglycan biosynthesis

doi: 10.1172/JCI137681

Figure Lengend Snippet: Levels of cytosolic free (and CMP-) Kdn (A–E) and CMP-Kdn (F–J) were measured by DMB-HPLC. Results are shown for (A and F) HEK293A cells; (B and G) BJAB K88 (GNE WT) cells; (C and H) BJAB K20 (GNE-KO) cells; (D and I) PMI WT mouse embryonic fibroblasts; and (E and J) PMI-KO cells. CMP-Sias were measured by DMB-HPLC after NaBH4 treatment. The mannose concentration was set at 0 or 15 mM, except for PMI-KO cells (0.025, 0.25, and 1 mM). Data are shown as the mean (SD). *P < 0.05, by unpaired t test.

Article Snippet: HEK293A cells (Thermo Fisher Scientific) were cultivated in RPMI 1640 media with 10% FBS; HUVECs (American Type Culture Collection [ATCC]) in EBM2 media (Lonza); BJAB K20 and BJAB K88 cells (provided by Stephan Hinderlich and the late Werner Reutter, Beuth University of Applied Sciences, Berlin, Germany) in RPMI 1640 media with 10% FBS; PMI WT and PMI-KO cells in DMEM containing 1 g/L glucose and 10% FBS; and HK2 cells in keratinocyte serum-free media supplemented with bovine pituitary extract and recombinant EGF (all from Gibco, Thermo Fisher Scientific).

Techniques: Concentration Assay

Journal: The Journal of Clinical Investigation

Article Title: Evolutionary conservation of human ketodeoxynonulosonic acid production is independent of sialoglycan biosynthesis

doi: 10.1172/JCI137681

Figure Lengend Snippet: Mouse monoclonal anti–Kdn IgG antibody, with kdn3G recognizing the (Kdn) ganglioside GM3 and mouse monoclonal anti–Kdn IgM antibody, with kdn8kdn recognizing the α2,8-linked Kdn were used for flow cytometry to analyze cell-surface expression of Kdn. Results are shown for (A and B) BJAB K20 (GNE-KO) cells and (C and D) PMI-KO cells, with or without mannose feeding. (E) Empty vector, Cmas1, and Cmas2 were transiently expressed in HEK293A cells. Cells were harvested after 36 hours of mannose feeding (0 or 15 mM), and then membrane fractions were prepared to analyze the conjugated form of Sia by DMB-HPLC analysis (Kdn, Neu5Gc, and Neu5Ac). Representative HPLC peaks for empty vector (F and G) or Cmas2-transfected (H and I) membrane samples. (F and H) 0 mM and (G and I) 15 mM mannose feeding. Data are representative of 2 independent experiments. Ac, Neu5Ac; Gc, Neu5Gc; Man, mannose.

Article Snippet: HEK293A cells (Thermo Fisher Scientific) were cultivated in RPMI 1640 media with 10% FBS; HUVECs (American Type Culture Collection [ATCC]) in EBM2 media (Lonza); BJAB K20 and BJAB K88 cells (provided by Stephan Hinderlich and the late Werner Reutter, Beuth University of Applied Sciences, Berlin, Germany) in RPMI 1640 media with 10% FBS; PMI WT and PMI-KO cells in DMEM containing 1 g/L glucose and 10% FBS; and HK2 cells in keratinocyte serum-free media supplemented with bovine pituitary extract and recombinant EGF (all from Gibco, Thermo Fisher Scientific).

Techniques: Flow Cytometry, Expressing, Plasmid Preparation, Membrane, Transfection

Journal: Cancer Science

Article Title: Impairment of double‐strand breaks repair and aberrant splicing of ATM and MRE11 in leukemia–lymphoma cell lines with microsatellite instability

doi: 10.1111/j.1349-7006.2006.00165.x

Figure Lengend Snippet: Summary of microsatellite instability and mutation of intronic mononucleotide repeats of ATM and MRE11 in leukemia–lymphoma cell lines

Article Snippet: Cell lines A total of 50 leukemia–lymphoma cell lines derived from Burkitt lymphoma (BJAB, BL36, BL60, BL137, Daudi, Katata, NAB‐2, Namalwa, Raji and Ramos), diffuse large B‐cell lymphoma (OPL‐1, OPL‐2, OPL‐3, OPL‐4, OPL‐5, OPL‐7 and Pal‐1), ( 24 , 25 , 26 ) precursor B‐cell acute lymphoblastic leukemia (NALM‐6 and REH), T‐cell acute lymphoblastic leukemia (CCRF‐CEM, HPB‐ALL, MOLT‐4, MOLT‐14, Jurkat and Peer), adult T‐cell leukemia (MT‐1, MT‐2, MT‐4 and TL‐Om1), NK/NKT‐cell lymphoma (KHYG‐1, MOTN‐1, MTA, NK‐92, SNK‐6, SNT‐8 and YT), ( 27 ) Hodgkin lymphoma (HDLM‐2, HD‐My‐Z, KM‐H2, L‐428, L‐540 and L‐1236), non‐B non‐T acute lymphoblastic leukemia (P30/Ohkubo), pyothorax‐associated lymphoma (PAL) with dual B/T‐cell phenotype (Deglis), ( 28 ) acute myeloid leukemia (CTV‐1, P31/Fujioka, HEL, HL‐60 and KG‐1), and chronic myeloid leukemia (K‐562) were used in this study.

Techniques: Mutagenesis, Expressing, Control

Journal: Cancer Medicine

Article Title: miR‐155‐regulated mTOR and Toll‐like receptor 5 in gastric diffuse large B‐cell lymphoma

doi: 10.1002/cam4.4466

Figure Lengend Snippet: MiR‐155 inhibits DEPTOR with secondary activation of mTOR in B lymphoma cell lines. (A) A model for the interactions between miR‐155 and the 3'UTR of DEPTOR. The RNAhyb program predicts a binding site starting at 480 nucleotides after the stop codon. Upper: sequence of the target site at the 3'UTR of DEPTOR; Lower: sequence of miR‐155. The 5' and 3' ends are specified. (B) Confirmation of the binding site for miR‐155 with luciferase assay in U2932 cells. Y‐axis: relative luciferase activity. X‐axis, from left to right: U2932 cells transfected with miR‐155 and DEPTOR, with mt‐miR‐155 and DEPTOR, with miR‐155 and mt DEPTOR, and with mt miR‐155 and mt DEPTOR (wt: wild type; mt mutant). (C) Induction of phosphorylated mTOR (p‐mTOR) by miR‐155 in BJAB cells. An EGFP+ vector is used to over‐express miR‐155. After transfection and sorting, >90% of the cells are EGFP+. Overexpression of miR‐155 reduced the level of DEPTOR with secondary increase in the level of p‐mTOR. (D) A model for induction of p‐mTOR by miR‐155 in gastric diffuse large B‐cell lymphoma resistant to H. pylori eradication therapy. In the mTOR complex, DEPTOR together with either RAPTOR in mTORC1 or RICTOR in mTORC2 inhibit the activity of mTOR. In the presence of increased miR‐155 and decreased DEPTOR, the inactive complex dissociates into active form. D: DEPTOR and R: RAPTOR or RICTOR

Article Snippet: BJAB and U2932 cells derived from B‐cell lymphomas were obtained from Bioresource Collection and Research Center (BCRC, Hsinchu, Taiwan) and grown in RPMI160 medium.

Techniques: Activation Assay, Binding Assay, Sequencing, Luciferase, Activity Assay, Transfection, Mutagenesis, Plasmid Preparation, Over Expression

Journal: BMC Cancer

Article Title: Peripheral T-lymphocytes express WNT7A and its restoration in leukemia-derived lymphoblasts inhibits cell proliferation

doi: 10.1186/1471-2407-12-60

Figure Lengend Snippet: Information of the oligonucleotides used for the qRT-PCR analysis

Article Snippet: As may be seen in Figure , WNT7A - inducible expression in all three cell lines decreased strongly, especially in BJAB-Tet- WNT7A and in CEM-Tet-WNT7A cells, decreasing to 40.62 and 38.77%, respectively, after 48 h of growth in the presence of doxycycline.

Techniques: Sequencing

Journal: BMC Cancer

Article Title: Peripheral T-lymphocytes express WNT7A and its restoration in leukemia-derived lymphoblasts inhibits cell proliferation

doi: 10.1186/1471-2407-12-60

Figure Lengend Snippet: Expression of WNT7A in total peripheral blood cells, Peripheral blood mononuclear cells (PBMC), sorted CD3 + and CD19 + cells obtained from healthy volunteers' samples . (A) Dot blot graphics showing the selected region for sorting (upper left panel), CD3-Fluorescein isothiocyanate (FITC), and CD19-Phycoerythrin (PE) marked cells (upper right panel), isolated CD19 + cells (lower left panel), and isolated CD3 + cells (lower right panel). (B) Figure showing the average of the WNT7A relative expression ratio ± Standard deviation (SD) obtained from the different groups. Normalization was performed by setting CP values of PBMC as 1 and taking GAPDH , RPL32 , and RPS18 as reference genes. Experiments were carried out at least twice in all cases.

Article Snippet: As may be seen in Figure , WNT7A - inducible expression in all three cell lines decreased strongly, especially in BJAB-Tet- WNT7A and in CEM-Tet-WNT7A cells, decreasing to 40.62 and 38.77%, respectively, after 48 h of growth in the presence of doxycycline.

Techniques: Expressing, Dot Blot, Isolation, Standard Deviation

Journal: BMC Cancer

Article Title: Peripheral T-lymphocytes express WNT7A and its restoration in leukemia-derived lymphoblasts inhibits cell proliferation

doi: 10.1186/1471-2407-12-60

Figure Lengend Snippet: Relative expression of WNT7A in control cells and leukemia-derived cell lines . (A) Relative expression levels of WNT7A determined by qRT-PCR in resting and phytohemagglutinin-activated PBMC (activation was performed in PBMC from two volunteers and named TxPHA 1 and TxPHA 2). Values of the resting PBMCs (Control) of both volunteers were set as 1. (B) Relative expression levels of WNT7A determined by qRT-PCR in Jurkat, CEM, K562, HL60, and BJAB cell lines normalized to the healthy volunteers control group (setting as 1). In A, quantification was calculated using, Ribosomal Protein L32 ( RPL32 ) and Ribosomal Protein S18 ( RPS18 ) as reference genes. In B, quantification was calculated using additionally Glyceraldehyde-3-phosphate dehydrogenase ( GAPDH ). The graphics show means obtained with both reference genes and Standard deviations (SD). Experiments were carried out at least twice in duplicate.

Article Snippet: As may be seen in Figure , WNT7A - inducible expression in all three cell lines decreased strongly, especially in BJAB-Tet- WNT7A and in CEM-Tet-WNT7A cells, decreasing to 40.62 and 38.77%, respectively, after 48 h of growth in the presence of doxycycline.

Techniques: Expressing, Control, Derivative Assay, Quantitative RT-PCR, Activation Assay

Journal: BMC Cancer

Article Title: Peripheral T-lymphocytes express WNT7A and its restoration in leukemia-derived lymphoblasts inhibits cell proliferation

doi: 10.1186/1471-2407-12-60

Figure Lengend Snippet: WNT7A expression in immature cells from patients with Acute lymphoblastic leukemia (ALL) . (A) Box plot graphics showing ΔCP values taking GAPDH (left panel) or RPL32 (right panel) as reference genes. The graphics display median (dark lines), 25-75th percentile (boxes), interquartile ranges (whiskers), and outliers (small, dark circles) from 14 patients with ALL. Peripheral blood cells from 19 controls were included for comparison. Statistical significances are shown between both groups. (B) Graphic showing an average of the relative expression levels of WNT7A from the control and from the group with patients with ALL. The value obtained in the healthy volunteers control group was set as 1 and normalized either with GAPDH or RPL32 as reference genes.

Article Snippet: As may be seen in Figure , WNT7A - inducible expression in all three cell lines decreased strongly, especially in BJAB-Tet- WNT7A and in CEM-Tet-WNT7A cells, decreasing to 40.62 and 38.77%, respectively, after 48 h of growth in the presence of doxycycline.

Techniques: Expressing, Comparison, Control

Journal: BMC Cancer

Article Title: Peripheral T-lymphocytes express WNT7A and its restoration in leukemia-derived lymphoblasts inhibits cell proliferation

doi: 10.1186/1471-2407-12-60

Figure Lengend Snippet: WNT7A overexpression in Jurkat cells . Overexpression of WNT7A in Jurkat cells using the lentiviral system was confirmed by qRT-PCR assays. (A) Graphic showing WNT7A relative expression taking GAPDH , RPL32 and RPS18 as constitutive genes in cells infected with pLVX (vector alone) or with pLVX-WNT7A. Infection was carried out in duplicate (pLVX-WNT7A clone 1 and 2). (B) Amplification products of WNT7A and RPS18 were run in 2.0% agarose gels and photodocumented. (C) Western blot assays (from 50 μg total protein) showing the presence of WNT7a protein only in pLVX-WNT7A-infected cells. The magic mark (2 μl) was used as protein marker.

Article Snippet: As may be seen in Figure , WNT7A - inducible expression in all three cell lines decreased strongly, especially in BJAB-Tet- WNT7A and in CEM-Tet-WNT7A cells, decreasing to 40.62 and 38.77%, respectively, after 48 h of growth in the presence of doxycycline.

Techniques: Over Expression, Quantitative RT-PCR, Expressing, Infection, Plasmid Preparation, Amplification, Western Blot, Marker

Journal: BMC Cancer

Article Title: Peripheral T-lymphocytes express WNT7A and its restoration in leukemia-derived lymphoblasts inhibits cell proliferation

doi: 10.1186/1471-2407-12-60

Figure Lengend Snippet: WNT7A inhibits cell proliferation in Jurkat cells . (A) Percentage of cell proliferation measured in Jurkat cells, Jurkat-pLVX (stable transduced with empty-vector), or Jurkat-pLVX-WNT7A cells after 48 h of culture. Percentages of cell proliferation were obtained by taking Optical density (OD) values of the Jurkat cells as 100% cell proliferation. (B) Leukemia-derived Jurkat cells and PBMC were exposed for 48 h to 3 μg/mL of recombinant human WNT7a (+rhWNT7a). Afterward, the percentage of cell proliferation was measured by employing WST-1 and setting the value of non-treated control cells (Jurkat and PBMC for their respective assays) at 440 nm as 100% of cell proliferation.

Article Snippet: As may be seen in Figure , WNT7A - inducible expression in all three cell lines decreased strongly, especially in BJAB-Tet- WNT7A and in CEM-Tet-WNT7A cells, decreasing to 40.62 and 38.77%, respectively, after 48 h of growth in the presence of doxycycline.

Techniques: Transduction, Plasmid Preparation, Derivative Assay, Recombinant, Control

Journal: BMC Cancer

Article Title: Peripheral T-lymphocytes express WNT7A and its restoration in leukemia-derived lymphoblasts inhibits cell proliferation

doi: 10.1186/1471-2407-12-60

Figure Lengend Snippet: Restoration of WNT7A does not induce the canonical β-catenin pathway in Jurkat cells . (A) Relative expression levels of AXIN2 , MYC , JUN , and FRA-1 , determined by qRT-PCR in Jurkat-pLVX or Jurkat-pLVX-WNT7A clone-1 and -2 infected cells. Quantification was calculated by normalizing with Jurkat-pLVX (setting as 1) and utilizing GAPDH and RPL32 as reference genes. The graphic shows medians obtained with both reference genes ± Standard deviations (SD). (B) Relative expression levels of AXIN2 , MYC , JUN , and FRA-1 , determined by qRT-PCR in Jurkat-pLVX or Jurkat-pLVX-WNT7A clone-1 in the absence or presence of LiCl. Quantification was calculated by normalizing LiCl-treated cells with their corresponding Jurkat-pLVX or Jurkat-pLVX-WNT7A non-treated cells (setting each as 1) and utilizing GAPDH and RPL32 as reference genes. The graphic shows medians obtained with both reference genes ± SD. (C) Western blot analysis detecting β-catenin and β-actin in the presence or absence of LiCl and WNT7a.

Article Snippet: As may be seen in Figure , WNT7A - inducible expression in all three cell lines decreased strongly, especially in BJAB-Tet- WNT7A and in CEM-Tet-WNT7A cells, decreasing to 40.62 and 38.77%, respectively, after 48 h of growth in the presence of doxycycline.

Techniques: Expressing, Quantitative RT-PCR, Infection, Western Blot

Journal: BMC Cancer

Article Title: Peripheral T-lymphocytes express WNT7A and its restoration in leukemia-derived lymphoblasts inhibits cell proliferation

doi: 10.1186/1471-2407-12-60

Figure Lengend Snippet: WNT7A overexpression in K562-, BJAB- and CEM-Tet-WNT7A cells . Overexpression of WNT7A in these cell lines using the doxycycline-inducible lentiviral expression system was confirmed by qRT-PCR and Western blot assays. (A) Graphics showing the amplification curves obtained for WNT7A and RPL32 in cells treated or not with Doxycycline (Doxy) for 4 h. The WNT7A normalized relative ratio of the cells treated with Doxy was calculated utilizing the non-treated cells as calibrator and taking GAPDH, RPL32 and RPS18 as reference genes in two independent experiments. (B) Western blot assays (from 50 μg total protein) showing the presence of WNT7a protein after a 38-h incubation with Doxy. Photo-documentation of the Coomassie stained gel is included as protein loading control.

Article Snippet: As may be seen in Figure , WNT7A - inducible expression in all three cell lines decreased strongly, especially in BJAB-Tet- WNT7A and in CEM-Tet-WNT7A cells, decreasing to 40.62 and 38.77%, respectively, after 48 h of growth in the presence of doxycycline.

Techniques: Over Expression, Expressing, Quantitative RT-PCR, Western Blot, Amplification, Incubation, Staining, Control

Journal: BMC Cancer

Article Title: Peripheral T-lymphocytes express WNT7A and its restoration in leukemia-derived lymphoblasts inhibits cell proliferation

doi: 10.1186/1471-2407-12-60

Figure Lengend Snippet: Restoration of WNT7A in K562-, BJAB-, and CEM-Tet-WNT7A cells inhibits their cell proliferation . Graphs of the left column depict the percentage of cell proliferation measured in K562-, BJAB-, and CEM-Tet-WNT7A cells after 24-h and 48-h incubation with Doxycycline (Doxy). The empty-vector cell lines (K562-, BJAB-, and CEM-Tet) were also included as controls. The percentage of cell proliferation was measured by adding WST-1 to the cell cultures for 2 h and by reading the absorbance of treated and untreated cells at 440 nm. The value in each cell line of non-treated cells after 24 or 48 h was setting as 100% of cell proliferation. Right column graphs illustrate the histograms of the flow-cytometric analysis done for apoptosis detection utilizing Annexin-V-FLUOS as markers in non-treated (filled curve) and 48-h Doxy-treated cells (open curve). Ordinate: number of cells; Abscissa: fluorescence intensities.

Article Snippet: As may be seen in Figure , WNT7A - inducible expression in all three cell lines decreased strongly, especially in BJAB-Tet- WNT7A and in CEM-Tet-WNT7A cells, decreasing to 40.62 and 38.77%, respectively, after 48 h of growth in the presence of doxycycline.

Techniques: Incubation, Plasmid Preparation, Fluorescence