Journal: Cancer medicine

Article Title: Biliary tumorigenic effect on hypopharyngeal cells is significantly enhanced by pH reduction.

doi: 10.1002/cam4.2194

Figure Lengend Snippet: FIGURE 1 pH 4.0 optimally enhances bile‐induced nuclear translocation of phospho‐NF‐κB and activation of STAT3 in HHPC. Immunofluorescence staining for phospho‐NF‐κB (p‐p65 S536) reveals that primary bile acids with DCA at strongly acidic pH (4.0) induced (A) p‐NF‐κB nuclear translocation, demonstrating increased p‐NF‐κB nuclear staining [green: p‐p65 (S536); blue: DAPI for nuclear staining] and (B) activation of STAT3, demonstrating elevated phospho‐STAT3(Tyr707) staining [green: p‐STAT3 (Tyr707); blue: DAPI for nuclear staining], compared to HHPC exposed to bile at weakly acidic (5.5) or neutral pH (7.0) and controls at pH 4.0. 5.5 and 7.0. DCA, deoxycholic acid; HHPC, human hypopharyngeal primary cells

Article Snippet: HHPC were provided by Celprogen Inc., Torrance, CA and cultured as previously described.17,19 In detail, The HHPC were plated in noncoated flasks and were grown in Human Hypopharyngeal Normal Cell Culture Media with Serum (Celprogen Inc. CA), at 37°C in humidified air and 5% CO2.

Techniques: Translocation Assay, Activation Assay, Immunofluorescence, Staining

Journal: Cancer medicine

Article Title: Biliary tumorigenic effect on hypopharyngeal cells is significantly enhanced by pH reduction.

doi: 10.1002/cam4.2194

Figure Lengend Snippet: FIGURE 3 Bile‐induced NF‐κB transcriptional activity is optimized at pH 4.0 in HHPC. A, Columns represent luciferase NF‐κB relative transcriptional activity (mean ± SE of two independent experiments) in HHPC transfected with control luciferase reporter (luc2P) and NF‐ κB luciferase responsive element (NF‐κB‐Luc2P). B, Columns represent ratios of luciferase NF‐κB relative transcriptional activity between experimental groups (primary bile acids with DCA) and controls at pH 4.0, 5.5 and 7.0 (NF‐κB‐Luc2P/Luc2P: NF‐κB luciferase responsive element/control luciferase reporter). DCA, deoxycholic acid; HHPC, human hypopharyngeal primary cells

Article Snippet: HHPC were provided by Celprogen Inc., Torrance, CA and cultured as previously described.17,19 In detail, The HHPC were plated in noncoated flasks and were grown in Human Hypopharyngeal Normal Cell Culture Media with Serum (Celprogen Inc. CA), at 37°C in humidified air and 5% CO2.

Techniques: Activity Assay, Luciferase, Transfection, Control

Journal: Cancer medicine

Article Title: Biliary tumorigenic effect on hypopharyngeal cells is significantly enhanced by pH reduction.

doi: 10.1002/cam4.2194

Figure Lengend Snippet: FIGURE 4 Bile‐induced transcriptional activation of cancer‐related genes is optimized at pH 4.0 in HHPC. A, The highest transcriptional levels of the analyzed NF‐κB‐related genes with oncogenic function, are depicted in HHPC exposed to primary bile acids with DCA at strongly acidic pH (4.0) relative to HHPC exposed to bile with DCA at weakly acidic (5.5) or neutral pH (7.0) and controls at pH 4.0, 5.5 and 7.0. Weakly acidic pH (5.5) also induced higher transcriptional level of the analyzed genes compared to corresponding control at pH 5.5. Graphs, created by GraphPad Prism 7 software (transcriptional levels of the analyzed genes are normalized to hGAPDH by real‐time qPCR analysis) (one‐way ANOVA, Freidman test). B, Graphs represent transcriptional levels of each analyzed gene, RELA(p65), TNF‐α, STAT3, bcl‐2, WNT5A, and EGFR (relative to hGAPDH reference gene), in experimental and control‐treated HHPC. The data are derived from real‐time qPCR analysis. C, Table shows overexpression ratios between bile (primary bile acids) with DCA at pH 4.0, 5.5 and 7.0 and corresponding controls (data are derived from three independent experiments. Graphs, created by GraphPad Prism 7 software; by t‐test; multiple comparisons by Holm‐Sidak). DCA, deoxycholic acid; HHPC, human hypopharyngeal primary cells

Article Snippet: HHPC were provided by Celprogen Inc., Torrance, CA and cultured as previously described.17,19 In detail, The HHPC were plated in noncoated flasks and were grown in Human Hypopharyngeal Normal Cell Culture Media with Serum (Celprogen Inc. CA), at 37°C in humidified air and 5% CO2.

Techniques: Activation Assay, Control, Software, Derivative Assay, Over Expression