human small cell carcinoma adrenal gland cortex sw13 cells  (ATCC)

Journal: PLoS ONE

Article Title: Comprehensive Suppression of All Apoptosis-Induced Proliferation Pathways as a Proposed Approach to Colorectal Cancer Prevention and Therapy

doi: 10.1371/journal.pone.0115068

Figure Lengend Snippet: A. HCT116 cells were exposed for 24 h to mock (M), 1 mg/ml roasted coffee extract (R), 5 mM butyrate and 100 µg/ml propolis (BP), or butyrate/propolis and increasing concentrations of coffee extract: 0.25, 0.5 or 1.0 mg/ml. Apoptosis was measured by flow cytometry as described in . B. HCT-116 cells were treated as in (A) for 17 hours and total cell lysates were analyzed by western blotting. C. HCT-116 and LT 97 cells were exposed for 20 h, and HCT-R cells were exposed for 42 h to mock (M), 0.5 µM AZD6244 (A), the combination; 5 mM butyrate, 100 µg/ml propolis, 1 mg/ml roasted coffee extract (BPR), or BPR and 0.5 µM AZD6244 (BPR-A). Apoptosis was measured by flow cytometry, as described in . Statistically significant differences in apoptotic levels are noted by asterisks (P<0.05). D. Clonal growth assays. Percent clonal growth was calculated by dividing the number of cell colonies by the number of plated cells, and multiplying by 100. The ratios of clonal growth were calculated by dividing the percent clonal growth in mock-treated samples by the percent clonal growth in agent-treated samples. Experiments were repeated four to six times with triplicate samples per experiment. Statistically significant differences among cell lines in their mean ratios were determined by one-way ANOVA. For cells exposed to butyrate, F(6,31) = 10.26, P<0.0001; for cells exposed to propolis and coffee extract, F(6,28) = 5.493, P = 0.0007, and for cells exposed to butyrate, propolis and coffee extract, F(6,26) = 10.56, P<0.0001. The post-test calculations used the Bonferroni correction to adjust for multiple comparisons with 95% confidence. Among the cells exposed to butyrate, statistically significant differences (P<0.05, CI 95%) were detected in HCT116 vs. CCL92, HCT116 vs. HEK293, HCT116 vs. SW13, HCT116 vs. LA1-5s, HCT116 vs. HCT-R, CCD841CoN vs. HEK293, and CCD841CoN vs. HCT-R cells. Among the cell lines exposed to propolis and coffee extract, statistically significant differences (P<0.05, CI 95%) were detected in HCT116 vs. SW13, HCT-R vs. CCL92, CCD841CoN vs. HCT-R cells, HEK293 vs. HCT-R, and SW13 vs. HCT-R cells. Among cell lines exposed to butyrate, propolis, and coffee extract statistically significant differences (P<0.05, CI 95%) were detected in HEK293 vs. SW13 cells, and HCT116 vs. all other cell lines except for HCT-R cell line.

Article Snippet: The following cell lines were cultured in alpha-MEM with 10% FBS: human small cell carcinoma adrenal gland/cortex SW13 cells (minus phenotype, isolated by dilution cloning from a heterogeneous sample ATCC CCL105 in the laboratory of Dr. Elizabeth Hull, Midwestern University, AZ), human neural crest-derived non-neuronal progenitor LA1-5s cells (provided by Dr. R. Ross, Fordham University, NY), normal human fetal colonic cells CCD841CoN ((ATCC CRL-1790), human embryonic kidney epithelial HEK293T cells (ATCC CRL-1573), and mouse embryonic fibroblasts CCL92 (ATCC CCL-92).

Techniques: Flow Cytometry, Western Blot

Journal:

Article Title: Identification of Novel Principles of Keratin Filament Network Turnover in Living Cells V⃞

doi: 10.1091/mbc.E03-09-0707

Figure Lengend Snippet: Details of KF network formation in living SW13 cells synthesizing HK8-CFP and HK18-YFP fluorescent hybrids. (A) Epifluorescence microscopy (detection of HK18-YFP) of a characteristic peripheral segment (position of plasma membrane indicated at 3.5 min) highlighting intermediate steps of KF network production. Two KF precursors are marked by color and followed until integration into the filament system. Note the formation of small granules below the cell surface that elongate, fuse, and become part of the KF network. The corresponding movie 5 shows further details. (B) Size changes of a single particle that was tracked from its first detection until integration into the network. For clarity's sake only the pixels associated with that particular particle are shown. Sampling frequency, 30 s. (C) The graph is derived from B, showing a surface view of the altering particle dimensions demonstrating a continuous, nonincremental particle elongation (l) before fusion with the KF network over time (t). (D) 3D reconstruction (voltex presentation) of the HK18-YFP fluorescence as recorded by confocal laser scanning microscopy of a peripheral region revealing spatial characteristics of various intermediates of filament assembly in relationship to intracellular topology (position of cell border demarcated). The volume of the top segment has the dimensions 20 × 20 × 3.5 μm. The bottom cube is a cross section to specifically highlight the increase in diameter of KF precursors only after integration into the peripheral KF network. (E) Plot summarizing the quantitative analysis of 10 time-space diagrams (see, e.g., C) depicting the increase in length (top graph) and diameter (bottom graph). Note the continuous elongation of particles during the entire 20 min in contrast to the increase in diameter that is limited to the first 5 min. Bars, SD. Bars, 1 μm in A and D.

Article Snippet: Cell Lines and cDNA Transfection The following cell lines were used and cultured according to the specifications given by ATCC or the respective references: rat kangaroo kidney PtK2 cells (ATCC CCL-56), Madin-Darby canine kidney (MDCK) cells (clone 20; ATCC CCL-34), bovine kidney-derived MDBK cells (ATCC CCL-22), human mammary adenocarcinoma-derived MCF7 cells (ATCC HTB-22), human hepatocellular carcinoma-derived PLC cells (ATCC CRL8024), human colon carcinoma CaCo-2 cells (ATCC HTB-37), human immortalized HaCaT keratinocytes ( Boukamp et al., 1988 ), and human small-cell carcinoma SW13 cells derived from the adrenal cortex (ATCC CCL-105).

Techniques: Epifluorescence Microscopy, Clinical Proteomics, Membrane, Single Particle, Sampling, Derivative Assay, Fluorescence, Confocal Laser Scanning Microscopy