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. 2016 Jun 29;11(1):79–83. doi: 10.1080/19336950.2016.1207026

Figure 3.

Figure 3.

CIM0216 and pregnenolone sulfate are TRPM3 ligands. (A) HEK293 cells containing a tetracycline-inducible TRPM3 transcription unit were infected with recombinant lentiviruses encoding the collagenase promoter/luciferase reporter gene. The cells were serum-starved for 24 hours in the presence of tetracycline (1 μg/ml) and then stimulated with either CIM0216 (20 μM) or pregnenolone sulfate (PregS, 20 μM) in the presence or absence of mefenamic acid (Mef, 30 μM) for 24 hours. Cell extracts were prepared and analyzed for luciferase activities. Luciferase activity was normalized to the protein concentration. Data shown are mean +/− SD of 3 experiments performed in quadruplicate; (☆☆☆, P < 0.001). (B) HEK293 cells containing a tetracycline-inducible TRPM3 transcription unit were infected with a lentivirus encoding the collagenase promoter/luciferase reporter gene (Coll.luc). In addition, cells were infected with a lentivirus encoding a TRPM3-specific shRNA. As a control, cells were infected with a lentivirus generated with the lentiviral transfer vector pLL3.7 (mock). The cells were serum-starved for 24 hours in the presence of tetracycline (1 μg/ml) and then stimulated with either CIM0216 (20 μM) or pregnenolone sulfate (PregS, 20 μM) for 24 hours as indicated. Cell extracts were prepared and analyzed for luciferase activities. Luciferase activity was normalized to the protein concentration. Data shown are mean +/− SD of 3 experiments performed in quadruplicate (☆☆☆, P < 0.001).