Journal: PLoS ONE

Article Title: Khellin and Visnagin Differentially Modulate AHR Signaling and Downstream CYP1A Activity in Human Liver Cells

doi: 10.1371/journal.pone.0074917

Figure Lengend Snippet: A) HepG2 cells were treated with 10 µM visnagin, 10 µM khellin, and/or DMSO for 4 h, 8 h, 16 h, and 24 h. The results from PCR are shown as fold of DMSO-treated control cells. The data are mean from three independent experiments and were normalized to beta-actin transcription. * - value is significantly different from DMSO-treated cells ( p < 0.05). B) HepG2 cells were pre-treated with 20 µM MNF for 1 h and then exposed to 10 µM visnagin or 10 µM khellin for additional 16 h. The results from PCR are shown as fold of DMSO-treated control cells. The data are mean from three independent experiments and were normalized to beta-actin transcription. * - value is significantly different from DMSO-treated cells ( p < 0.05). # - value is significantly reduced in comparison to cells treated with VIS and KHEL, respectively; ( p < 0.05). C) HepG2 cells were treated with visnagin (VIS; 1 µM-20 µM), khellin (KHEL; 1 µM-20 µM), 1 µM 3MC, 5 nM TCDD, and/or vehicle (DMSO; 0.1% v/v) for 48 h. Thereafter, western blotting analyses for detection of CYP1A1 and actin were performed as described in Materials and Methods section. The representative western blot analysis of two independent experiments (passages) is presented. D) HepG2 cells were treated with visnagin (VIS; 1 µM-20 µM), khellin (KHEL; 1 µM-20 µM), 1 µM 3MC, 5 nM TCDD, and/or vehicle (DMSO; 0.1% v/v) for either 16 h (upper panel) or 48 h (lower panel). EROD activity was determined as described in Materials and Method section. Analyses were performed in three independent experiments and are shown as fold induction over untreated cells. * - value is significantly different from DMSO-treated cells ( p < 0.05). E) HepG2 cells were treated with TCDD (5 nM) for 48 h. Thereafter, substrate mixture was supplemented with increasing doses of visnagin (VIS 1 nM -20 µM) or khellin (KHEL; 1 nM -20 µM) and EROD activity was determined as described in Materials and Methods section. Data are mean from three independent experiments and are expressed as percentage (%) of TCDD-mediated induction (i.e. induction by TCDD = 100%). * - value is significantly different from TCDD-treated cells ( p < 0.005).

Article Snippet: Blots were probed with primary antibodies against CYP1A1 (goat polyclonal, sc-9828, G-18, diluted 1:500 – for detection in human hepatocytes; rabbit polyclonal, sc-20772, H-70, diluted 1:500 – for detection in HepG2 cells), CYP1B1 (mouse monoclonal, sc-374228, G-4, 1:1000), actin (goat polyclonal; sc-1616, 1-19, diluted 1:2000), all purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA) or GAPDH (rabbit monoclonal, 2118, 14C10, diluted 1:1000) purchased from Cell Signaling Technology, overnight at 4°C.

Techniques: Control, Comparison, Western Blot, Activity Assay

Journal: PLoS ONE

Article Title: Khellin and Visnagin Differentially Modulate AHR Signaling and Downstream CYP1A Activity in Human Liver Cells

doi: 10.1371/journal.pone.0074917

Figure Lengend Snippet: Influence of visnagin and khellin treatment on CYP1A1 mRNA expression in human primary hepatocytes.

Article Snippet: Blots were probed with primary antibodies against CYP1A1 (goat polyclonal, sc-9828, G-18, diluted 1:500 – for detection in human hepatocytes; rabbit polyclonal, sc-20772, H-70, diluted 1:500 – for detection in HepG2 cells), CYP1B1 (mouse monoclonal, sc-374228, G-4, 1:1000), actin (goat polyclonal; sc-1616, 1-19, diluted 1:2000), all purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA) or GAPDH (rabbit monoclonal, 2118, 14C10, diluted 1:1000) purchased from Cell Signaling Technology, overnight at 4°C.

Techniques: Expressing

Journal: Journal of Cellular and Molecular Medicine

Article Title: The role of ATP signalling in response to mechanical stimulation studied in T24 cells using new microphysiological tools

doi: 10.1111/jcmm.13520

Figure Lengend Snippet: Investigations on purinergic P2X and P2Y receptors involved in the stretch‐related Ca 2+ responses in T24 cells using pharmacological reagents. A, ATP 3‐dose‐response from 10 −6 mol L −1 to 10 −4 mol L −1 in media with and without calcium. B, ATP 3‐dose‐response from 10 −6 mol L −1 to 10 −4 mol L −1 in the presence and absence of non‐selective P2X antagonist PPADS 10 −6 mol L −1 and 10 −5 mol L −1 . C, 3‐dose‐response from 10 −6 mol L −1 to 10 −4 mol L −1 generated by ATP and different P2Y receptor agonists UTP, ADP. Data were collected from several cells in different experiments and presented as mean ± SEM

Article Snippet: Slides were then incubated with primary antibodies; guinea pig anti‐P2X3 (1:250; Novus Cat# NB100‐1658, RRID:AB_10001676, rabbit anti‐P2Y 6 (1:50; H‐70; Santa Cruz Biotechnology Cat# sc‐20127, RRID: AB_2156250), diluted in PBS containing 0.05% Triton X‐100 and 0.2% BSA for 1 hour in a moist chamber at room temperature.

Techniques: Generated

Journal: Journal of Cellular and Molecular Medicine

Article Title: The role of ATP signalling in response to mechanical stimulation studied in T24 cells using new microphysiological tools

doi: 10.1111/jcmm.13520

Figure Lengend Snippet: Fluorescence micrographs depicting immunoreactivity to P2 receptors in T24 cells. (A) P2X3, (B) P2Y 6 . P2 receptor subunit (red), cytoskeletal actin fibres (green) and cell nuclei (blue); scale bar = 15 μm

Article Snippet: Slides were then incubated with primary antibodies; guinea pig anti‐P2X3 (1:250; Novus Cat# NB100‐1658, RRID:AB_10001676, rabbit anti‐P2Y 6 (1:50; H‐70; Santa Cruz Biotechnology Cat# sc‐20127, RRID: AB_2156250), diluted in PBS containing 0.05% Triton X‐100 and 0.2% BSA for 1 hour in a moist chamber at room temperature.

Techniques: Fluorescence