Additional file 1 of Cryptotanshinone chemosensitivity potentiation by TW-37 in human oral cancer cell lines by targeting STAT3–Mcl-1 signaling
Creators: name: In-Hyoung Yang
name: Hong, Seung-Hyun
name: Minjung Jung
name: Ahn, Chi-Hyun
name: Yoon, Hye-Jung
name: Hong, Seong Doo
name: Sung-Dae Cho
name: Shin, Ji-Ae
Publisher: name: figshare
Year: 2020
Subjects: subject: Biochemistry
subject: Cell Biology
subject: Genetics
subject: FOS: Biological sciences
subject: FOS: Biological sciences
subject: Molecular Biology
subject: 39999 Chemical Sciences not elsewhere classified
subject: FOS: Chemical sciences
subject: FOS: Chemical sciences
subject: Immunology
subject: FOS: Clinical medicine
subject: FOS: Clinical medicine
subject: Cancer
subject: 111714 Mental Health
subject: FOS: Health sciences
subject: FOS: Health sciences
subject: Hematology

description: Additional file 1: Figure S1. TW-37 induces apoptosis in human oral cancer cell lines by suppressing p-STAT3Tyr705. Ca9.22, HSC-4, and HN22 cell lines were treated with 2.5- and 5-μM TW-37 for 48 h. a The cell viability was measured using the trypan blue exclusion assay. Bar graphs represent the mean ± SD values of three independent experiments. *P &lt; 0.05. b Protein levels of cleaved caspase-3 and cleaved PARP were analyzed using Western blotting. Actin was used as a loading control. c All cell lines stained with DAPI were visualized by fluorescence microscopy (magnification, ×400). d The expressions of p-STAT3 (Y705) and STAT3 were detected using Western blotting. Actin was used as a loading control. Figure S2. TW-37-induced apoptosis depending on caspase activation in human oral cancer cell lines. HSC-3 cell lines were pretreated with 10-μM Z-VAD for 2 h with/without 5-μM TW-37 for 48 h. a Protein levels of cleaved PARP were analyzed using Western blotting analysis. Actin was used as a loading control. b Bar graphs represent the mean ± SD values of three independent experiments. *P &lt; 0.05; #P &lt; 0.05. Figure S3. TW-37 slightly decreases Mcl-1 mRNA levels at 24 and 48 h. The HSC-3 cell lines were treated with 5-μM of TW-37 for 24 or 48 h. Relative mRNA levels of Mcl-1 were measured with qPCR and normalized to GAPDH. Bar graphs represent the mean ± SD values of triplicate experiments. *P &lt; 0.05. Figure S4. STAT3 inactivation is sufficient to induce apoptosis via the inhibition of Mcl-1 expression. The HSC-3 cell lines were treated with 5-μM of cryptotanshinone or stattic for 24 h. a The cell viability was measured using the trypan blue exclusion assay. b Protein levels of p-STAT3 (Y705), STAT3, Mcl-1, and cleaved PARP were analyzed using Western blotting analysis. Actin was used as a loading control. c Relative mRNA levels of Mcl-1 were measured with qPCR and normalized to GAPDH. d The HSC-3 cell lines were transiently transfected with 0.5-μg pcDNA3.1 or pcDNA3.1-Mcl-1 for 6 h. Protein levels of p-STAT3 (Y705) and STAT3 were determined using Western blotting analysis. Actin was used as a loading control. All bar graphs represent the mean ± SD values of three independent experiments. *P &lt; 0.05. Figure S5. TW-37-induced apoptosis is mediated by the suppression of STAT3-Mcl-1 signaling unlike ABT-737. The HSC-3 cell lines were treated with the indicated concentrations of TW-37 or ABT-737 for 48 h. a Protein levels of p-STAT3 (Y705), STAT3, Mcl-1, and cleaved PARP were analyzed using Western blotting analysis. Actin was used as a loading control. b Bar graphs represent the mean ± SD values of three independent experiments. *P &lt; 0.05.