mc3t3-e1 mouse calvaria-derived cells  (BioResource International Inc)

Journal: Heliyon

Article Title: Modification of TRPV4 activity by acetaminophen

doi: 10.1016/j.heliyon.2020.e03301

Figure Lengend Snippet: TRPV4-dependent [Ca 2+ ] i change in PC12 cells (A) Gene expression of TRP channels. Total RNA prepared from PC12, MC3T3-E1 (E1), and rat brain were subjected to reverse transcription followed by PCR using primers to assess mRNA expression of the indicated genes. Typical images of PCR products separated by agarose gel are shown. Intact images are shown in the supplementary Fig. S1 (B) PC12 cells were loaded with Fura-2-AM and stimulated with the TRPV4 agonist GSK1016790A (GSK). The change in the ratio of 510-nm fluorescence intensity excited at 340/380 nm was recorded and traces of the mean values from cells treated with the indicated concentrations of GSK are presented (C) Peak amplitudes in the GSK-induced [Ca 2+ ] i increase were measured and summarized in the bar graph. Each bar represents the means ± SEM of three independent experiments with approximately 10–20 cells in each experiment. ** P < 0.01 versus the corresponding value for cells stimulated with 0.1 μM GSK (D) Fura-2-loaded PC12 cells were first incubated with the indicated concentrations of HC-067047 (HC) or the vehicle alone, followed by stimulation with 3 μM GSK. Traces of the mean values from the cells are presented (E) Peak amplitudes in the GSK-induced [Ca 2+ ] i rise are summarized as in (C). ** P < 0.01 versus the corresponding value for cells treated with 3 μM GSK anole (0 μM HC-067407).

Article Snippet: The rat adrenal pheochromocytoma-derived cell line PC12, mouse calvaria-derived cell line MC3T3-E1, human cervical carcinoma HeLa cells, and other human-derived cell lines including Ca9-22, SAS, HaCaT, HSC-2, and HEK293 were obtained from RIKEN BioResource Research Center (Ibaraki, Japan) and maintained in the growth medium according to the instructions.

Techniques: Gene Expression, Reverse Transcription, Expressing, Agarose Gel Electrophoresis, Fluorescence, Incubation

Journal: Heliyon

Article Title: Modification of TRPV4 activity by acetaminophen

doi: 10.1016/j.heliyon.2020.e03301

Figure Lengend Snippet: Effect of APAP and AM 404 on [Ca 2+ ] i of PC12 cells (A) PC12 cells labeled with Fura-2-AM were treated with APAP (0.1 or 1 mM) or AM404 (10 or 100 μM) and the change in the ratio of 510-nm fluorescence intensity excited at 340/380 nm was recorded over time. Traces of the mean values from cells treated with the indicated concentrations of either APAP or AM404 are shown (B) Fura-2-loaded PC12 cells were first incubated with various concentrations of APAP, followed by adding 3 μM GSK1016790A (GSK). Traces of the mean values from the cells are shown (C) Summary of peak amplitudes in the GSK-induced increase in [Ca 2+ ] i . Each bar represents the means ± SEM of three independent experiments with approximately 10–20 cells in each experiment. ** P < 0.01 versus the corresponding value for cells treated with 3 μM GSK in the absence of APAP (0 mM APAP) (D) Fura-2-loaded PC12 cells were first incubated with various concentrations of AM404, followed by adding 3 μM GSK. Traces of mean values from the cells treated with different concentrations of AM404 are shown (E) Summary of peak amplitudes in the GSK-induced increase in [Ca 2+ ] i . Each bar represents the means ± SEM of three independent experiments with approximately 10–20 cells in each experiment. * P < 0.05, ** P < 0.01 versus the corresponding value for cells treated with 3 μM GSK in the absence of AM404 (0 μM AM404).

Article Snippet: The rat adrenal pheochromocytoma-derived cell line PC12, mouse calvaria-derived cell line MC3T3-E1, human cervical carcinoma HeLa cells, and other human-derived cell lines including Ca9-22, SAS, HaCaT, HSC-2, and HEK293 were obtained from RIKEN BioResource Research Center (Ibaraki, Japan) and maintained in the growth medium according to the instructions.

Techniques: Labeling, Fluorescence, Incubation

Journal: Heliyon

Article Title: Modification of TRPV4 activity by acetaminophen

doi: 10.1016/j.heliyon.2020.e03301

Figure Lengend Snippet: Expression of FAAH in PC12 and HeLa-mTRPV4 cells (A) Total RNA prepared from PC12 cells and rat brain was subjected to reverse transcription (RT) followed by PCR using primers for assessing mRNA expression of rat FAAH (rFAAH) or β-actin (rActb). Typical images of the PCR products separated on agarose gel are shown and the source images of agarose gel are shown in Supplementary Fig. S5 (B) RT-PCR was performed to detect mRNA expression of human FAAH (hFAAH) and β-actin (hActb) in HeLa-mTRPV4 cells and in SAS cells as a positive control expressing hFAAH. Typical images of the PCR products are shown and the source images of agarose gel are shown in Supplementary Fig. S5.

Article Snippet: The rat adrenal pheochromocytoma-derived cell line PC12, mouse calvaria-derived cell line MC3T3-E1, human cervical carcinoma HeLa cells, and other human-derived cell lines including Ca9-22, SAS, HaCaT, HSC-2, and HEK293 were obtained from RIKEN BioResource Research Center (Ibaraki, Japan) and maintained in the growth medium according to the instructions.

Techniques: Expressing, Reverse Transcription, Agarose Gel Electrophoresis, Reverse Transcription Polymerase Chain Reaction, Positive Control