Journal: Nature communications

Article Title: A red-shifted fluorescent substrate for aldehyde dehydrogenase

doi: 10.1038/ncomms4662

Figure Lengend Snippet: AldeRed 588-A is a specific substrate for ALDH. ( a ) Fluorescent candidates and the ALDEFLUOR™ reagent were tested with K562 and L1210/cpa cells. The x-axis represents selected detection filters of the LSRII FACS system. ( b ) AldeRed 588-A and the ALDEFLUOR™ reagent tested with L1210/cpa and L1210 cells. ( c ) Co-staining of AldeRed 588-A and the ALDEFLUOR™ reagent with K562 and L1210/cpa cells. DEAB: diethylaminobenzaldehyde.

Article Snippet: The murine leukemia cell line L1210 (ALDH low ) and L1210/cpa (ALDH hi ) were provided by Dr. Richard J. Jones (Johns Hopkins University) and maintained in suspension in RPMI 1640 supplemented with 10% FBS.

Techniques: Staining

Journal: General physiology and biophysics

Article Title: The expression of P-gp in leukemia cells is associated with cross-resistance to protein N-glycosylation inhibitor tunicamycin.

doi: 10.4149/gpb_2016039

Figure Lengend Snippet: Figure 2. Resistance of P-gp-positive cells to TNM. A. Expression of MDR1 and MRP1 genes in leukemia cells. Upper part: Detec tion of transcripts by RT PCR. The data shown are representative of three independ ent measurements. Lower part: Quantifica tion of PCR products. The data are expressed as the mean values ± S.E.M. from three independent measurements; * levels differ significantly from those of S or C (p < 0.01). B. Typical record of apoptosis/necrosis detec tion by the FAV and PI apoptosis necrosis kit. Data shown were obtained for SKM-1 cells passaged twice in the absence (left part) or presence of TNM (right part). C. Cell vi ability, detected in the lower-left panels of the apoptosis/necrosis cytogram (see panel B) for P-gp-negative and P-gp-positive L1210, SKM-1 and MOLM-13 cells passaged 1, 2 or 3 times in the presence of TNM. The data are expressed as the mean values ± S.E.M. from three independent measurements. Differences between P-gp-positive and P- gp-negative cells were always significant (p < 0.01) after the second passage in the pres ence of TNM. S, L1210 cells; R, L1210/R; T, L1210/T cells; C, SKM-1 and MOLM-13 cells; V, SKM-1/VCR and MOLM-13/VCR cells; L, SKM-1/LEN and MOLM-13/LEN cells; FAV, fluorescein isothiocyanate-labeled annexin V; PI, propidium iodide; TNM, tunicamycin.

Article Snippet: The following cell variants were used in this study: P-gp-negative cells obtained from Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH, (DSMZ Germany): • Murine lymphocytic leukemia L1210 (ACC 123); • Human acute myeloid leukemia SKM-1 (ACC 547); • Human acute myeloid leukemia MOLM-13 (ACC 554); • MOLM-13/LEN, obtained by culturing in media with stepwise increasing concentrations of LEN (Celgene, USA) (Imrichova et al. 2014, 2015) P-gp-positive cell variants: • L1210/R and L1210/T, obtained either by cultivation in stepwise-increasing concentrations of VCR (Polekova et al. 1992) or by transfection with a retrovirus encoding human P-gp (Sulova et al. 2010); the retrovirus was pHaMDRwt (Addgene product 10957 USA), which contains the full-length P-gp cDNA (Pastan et al. 1988); • SKM-1/VCR and SKM-1/LEN, obtained by culturing in media with stepwise-increasing concentrations of VCR (Sigma Aldrich, USA) and LEN (Imrichova et al. 2014, 2015); • MOLM-13/VCR, obtained by culturing in media with stepwise-increasing concentrations of VCR (Imrichova et al. 2014, 2015).

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Labeling

Journal: General physiology and biophysics

Article Title: The expression of P-gp in leukemia cells is associated with cross-resistance to protein N-glycosylation inhibitor tunicamycin.

doi: 10.4149/gpb_2016039

Figure Lengend Snippet: Figure 3. Structural characteristics of P-gp-positive and P-gp- negative leukemia cells. A. Light microscopy (magnification ×100). Cells were passaged one (1st), two (2nd) or three (3rd) times in the presence of tunicamycin and compared with control untreated cells (C). SKM-1/V and SKM-1/L represent SKM-1/VCR and SKM-1/ LEN cells; MOLM-13/V and MOLM-13/L represent MOLM-13/ VCR and MOLM-13/LEN cells. The data shown are representa tive of three independent measurements. B. Electron microgram of L1210 and L1210/R cells passaged in the absence or presence of tunicamycin.

Article Snippet: The following cell variants were used in this study: P-gp-negative cells obtained from Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH, (DSMZ Germany): • Murine lymphocytic leukemia L1210 (ACC 123); • Human acute myeloid leukemia SKM-1 (ACC 547); • Human acute myeloid leukemia MOLM-13 (ACC 554); • MOLM-13/LEN, obtained by culturing in media with stepwise increasing concentrations of LEN (Celgene, USA) (Imrichova et al. 2014, 2015) P-gp-positive cell variants: • L1210/R and L1210/T, obtained either by cultivation in stepwise-increasing concentrations of VCR (Polekova et al. 1992) or by transfection with a retrovirus encoding human P-gp (Sulova et al. 2010); the retrovirus was pHaMDRwt (Addgene product 10957 USA), which contains the full-length P-gp cDNA (Pastan et al. 1988); • SKM-1/VCR and SKM-1/LEN, obtained by culturing in media with stepwise-increasing concentrations of VCR (Sigma Aldrich, USA) and LEN (Imrichova et al. 2014, 2015); • MOLM-13/VCR, obtained by culturing in media with stepwise-increasing concentrations of VCR (Imrichova et al. 2014, 2015).

Techniques: Light Microscopy, Control

Journal: General physiology and biophysics

Article Title: The expression of P-gp in leukemia cells is associated with cross-resistance to protein N-glycosylation inhibitor tunicamycin.

doi: 10.4149/gpb_2016039

Figure Lengend Snippet: Figure 5. Proportions of cells in the different phases of the cell cycle after their cultivation in the presence or absence of TNM. A. Phase of cell cycle detected by staining of DNA with PI (L1210 cells). B. Quantification of cells in the subG1 phase (apoptotic cells) after 24- or 48-hour cultivation in the presence of TNM. S, L1210 cells; R, L1210/R; T, L1210/T cells; C, SKM-1 and MOLM-13 cells; L, SKM-1/LEN and MOLM- 13/LEN cells. Data are expressed as the mean ± S.E.M. from three independent measurements and are expressed in percentage of control experiment in the absence of TNM. * p < 0.01, + p < 0.05 vs. control. C and D. Quantification of P-gp-positive and P-gp-negative cells in the different phases of the cell cycle after cultivation of cells in the presence (black column) or absence (white column) of TNM for 24 or 48 hours. The data are expressed as the mean values ± S.E.M. from three independent measurements. * p < 0.01 vs. control in the absence of TNM.

Article Snippet: The following cell variants were used in this study: P-gp-negative cells obtained from Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH, (DSMZ Germany): • Murine lymphocytic leukemia L1210 (ACC 123); • Human acute myeloid leukemia SKM-1 (ACC 547); • Human acute myeloid leukemia MOLM-13 (ACC 554); • MOLM-13/LEN, obtained by culturing in media with stepwise increasing concentrations of LEN (Celgene, USA) (Imrichova et al. 2014, 2015) P-gp-positive cell variants: • L1210/R and L1210/T, obtained either by cultivation in stepwise-increasing concentrations of VCR (Polekova et al. 1992) or by transfection with a retrovirus encoding human P-gp (Sulova et al. 2010); the retrovirus was pHaMDRwt (Addgene product 10957 USA), which contains the full-length P-gp cDNA (Pastan et al. 1988); • SKM-1/VCR and SKM-1/LEN, obtained by culturing in media with stepwise-increasing concentrations of VCR (Sigma Aldrich, USA) and LEN (Imrichova et al. 2014, 2015); • MOLM-13/VCR, obtained by culturing in media with stepwise-increasing concentrations of VCR (Imrichova et al. 2014, 2015).

Techniques: Staining, Control

Journal: General physiology and biophysics

Article Title: The expression of P-gp in leukemia cells is associated with cross-resistance to protein N-glycosylation inhibitor tunicamycin.

doi: 10.4149/gpb_2016039

Figure Lengend Snippet: Figure 6. Concanavalin A (ConA) and Sambucus nigra agglutinin (SNA) binding to crude membrane fractions (detected by lectin-based microarray assay) and wheat germ agglutinin (WGA) binding to the cell surface (detected by flow cytometry) of P- gp-negative and P-gp-positive leukemia cells. The data are expressed as the mean values ± S.E.M. from three independent measure ments. * p < 0.02 vs. control C, + p < 0.02 vs. control in the absence of TNM. S, L1210 cells; R, L1210/R; T, L1210/T cells; C, SKM- 1 and MOLM-13 cells; V, SKM-1/VCR and MOLM-13/VCR cells; L, SKM-1/LEN and MOLM-13/LEN cells.

Article Snippet: The following cell variants were used in this study: P-gp-negative cells obtained from Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH, (DSMZ Germany): • Murine lymphocytic leukemia L1210 (ACC 123); • Human acute myeloid leukemia SKM-1 (ACC 547); • Human acute myeloid leukemia MOLM-13 (ACC 554); • MOLM-13/LEN, obtained by culturing in media with stepwise increasing concentrations of LEN (Celgene, USA) (Imrichova et al. 2014, 2015) P-gp-positive cell variants: • L1210/R and L1210/T, obtained either by cultivation in stepwise-increasing concentrations of VCR (Polekova et al. 1992) or by transfection with a retrovirus encoding human P-gp (Sulova et al. 2010); the retrovirus was pHaMDRwt (Addgene product 10957 USA), which contains the full-length P-gp cDNA (Pastan et al. 1988); • SKM-1/VCR and SKM-1/LEN, obtained by culturing in media with stepwise-increasing concentrations of VCR (Sigma Aldrich, USA) and LEN (Imrichova et al. 2014, 2015); • MOLM-13/VCR, obtained by culturing in media with stepwise-increasing concentrations of VCR (Imrichova et al. 2014, 2015).

Techniques: Binding Assay, Membrane, Microarray, Flow Cytometry, Control