Journal: Biochemical and biophysical research communications

Article Title: Involvement of TRPV3 and TRPM8 ion channel proteins in induction of mammalian cold-inducible proteins.

doi: 10.1016/j.bbrc.2017.11.136

Figure Lengend Snippet: Fig. 1. TRPV4 is necessary for induction of CIP transcripts by hypothermia. (A) U-2 OS cells were transfected with plasmids expressing shRNAs against TRPV4 (V4) or vector alone (C) with (þ) or without (-) plasmids expressing rat TRPV4. Transfected cells were cultured at 37 C for 40 h, then transferred to the indicated temperatures, and protein levels were determined by western blot after further 8-h incubation. Relative band intensities after normalization to ACTIN expression are shown below the panels (representative of 2 in- dependent experiments). (B) U-2 OS cells were cultured at 37 C or 32 C in the presence of RN1734 for 10 h, and analyzed by western blot. Band intensities relative to those at 32 C were determined after normalization to ACTIN (left, representative results. Right, data indicate mean ± SEM; n ¼ 3). *, P < 0.05. **, P < 0.01. (C) U-2 OS cells were cultured at 37 C or 32 C in the presence of RN1734 for 6 h, and analyzed by RT-qPCR and expressed as relative to those at 32 C after normalization to 18S rRNA (data indicate mean ± SEM; n ¼ 3 per group).

Article Snippet: The sources of reagents were as follows: RN1734 (Tocris Bioscience, Bristol, UK), S408271 (Sigma-Aldrich), AMTB hydrochloride (Tocris Bioscience), 2-APB (Santa Cruz Biotechnology), camphor (Tokyo Chemical Industry, Tokyo, Japan), WS-12 (Tocris Bioscience), and Fura-2-AM (Molecular Probes).

Techniques: Transfection, Expressing, Plasmid Preparation, Cell Culture, Western Blot, Incubation, Quantitative RT-PCR

Journal: Biochemical and biophysical research communications

Article Title: Involvement of TRPV3 and TRPM8 ion channel proteins in induction of mammalian cold-inducible proteins.

doi: 10.1016/j.bbrc.2017.11.136

Figure Lengend Snippet: Fig. 2. Induction of CIPs by hypothermia in the absence of TRPV4. (A) Genotyping of cell lines derived from wild-type mouse (WT) and TRPV4-knockout (KO) mouse (cell lines 1 and 2) by PCR. (B) TRPV4-KO cells were cultured at 37 C or 32 C for 24 h, and analyzed by western blot. Band intensities relative to those at 32 C were determined after normalization to ACTIN (left, representative results. Right, data indicate mean ± SEM; n ¼ 3). *, P < 0.05. **, P < 0.01. (C, D) WT (C) and TRPV4-KO (D) cells were cultured in the presence of RN1734 and analyzed as in (B) (left, representative results. Right, data indicate mean ± SEM; n ¼ 3). ns, not significant. (E, F) TRPV4-KO cells were cultured in the presence of S408271 (E) or AMTB (F) and analyzed as in (B) (representative of 2 independent experiments).

Article Snippet: The sources of reagents were as follows: RN1734 (Tocris Bioscience, Bristol, UK), S408271 (Sigma-Aldrich), AMTB hydrochloride (Tocris Bioscience), 2-APB (Santa Cruz Biotechnology), camphor (Tokyo Chemical Industry, Tokyo, Japan), WS-12 (Tocris Bioscience), and Fura-2-AM (Molecular Probes).

Techniques: Derivative Assay, Knock-Out, Cell Culture, Western Blot

Journal: International Journal of Molecular Sciences

Article Title: Weak Ultraviolet B Enhances the Mislocalization of Claudin-1 Mediated by Nitric Oxide and Peroxynitrite Production in Human Keratinocyte-Derived HaCaT Cells

doi: 10.3390/ijms21197138

Figure Lengend Snippet: Involvement of transient receptor potential type vanilloid 1 (TRPV1) in the UVB-induced production of RNS. ( A,B ) After we exposed cells to weak UVB, the cells were incubated for 3 h in the absence (Veh) and presence of 10 μM AMG9810 (AMG), 10 μM RN1734 (RN), or 2 mM glycoletherdiaminetetraacetic acid (EGTA). ( C ) Cells were incubated for 3 h in the absence (Cont) and presence of olvanil (Olv). The cells were incubated with 5 μM Fluo-8 AM, 5 μM DAF-2DA, or 5 μM NiSPY-3 for 30 min. The fluorescence intensities of Fluo-8, DAF-2, and NiSPY-3 were measured using a plate reader. n = 8. ** p < 0.01 vs. Cont. ## p < 0.01 and NS p > 0.05 vs. Veh.

Article Snippet: AMG9810, olvanil, and RN1734 were from FUJIFILM Wako Pure Chemical Corporation (Tokyo, Japan).

Techniques: Incubation, Fluorescence