Thus, we proposed the hypothesis that baicalin could suppress human cervical cancer by regulating PKC-associated pathways. In this study, the anti-cancer effect of baicalin was observed in 2 human cervical cancer cell lines (HeLa and SiHa). signal transducer and activator of transcription (STAT) 3 were reduced in baicalin administrated cervical cancer cells. Conclusions Baicalin exerted anti-cancer effects on human cervical cancer cells by targeting STAT3 regulated signaling pathways. which has been applied in Traditional Chinese Medicine (TCM) since ancient times. Modern pharmacological investigations revealed the biological activities of baicalin, such as anti-oxidant, anti-fibrosis, anti-bacterial, and anti-inflammatory effects [11]. Previous studies have indicated the anti-cancer activity of baicalin against multiple human cancers including hepatic cancer, lung cancer, and lymphoma [12C14]. There are very few studies investigating the anti-cancer effects of baicalin on human cervical cancer. Moreover, a previous study suggested the regulation effect of baicalin on PCK15. Thus, we proposed the hypothesis that baicalin could suppress human cervical cancer by regulating PKC-associated pathways. In this study, the anti-cancer effect of baicalin was observed in 2 human cervical cancer cell lines (HeLa and SiHa). The involvement of PKC/STAT3 signaling was also investigated as a possible molecular mechanism. We believe that results from Sunitinib this study could not only add more information concerning the mechanisms of pathogenesis of cervical cancer, but also contribute to accumulating evidence supporting potential application of baicalin as an anti-cancer agent in cervical cancer patients. Material and Methods Agents and antibodies Agents and antibodies included: baicalin (Sigma-Aldrich, Cat# 572667), TUNEL kit (Roche, Cat# 11684795910), PKC antibody Sunitinib (Cell Signaling Tech, Cat#9372, 1: 4000), STAT3 antibody (Cell Signaling Tech, Cat#4368, 1: 2000), phosphorylated STAT3 antibody (p-STAT3, Cell Signaling Tech, Cat#8119, 1: 2000), Survivin antibody (Abcam, Cat#ab76424, 1: 4000), matrix metalloproteinase (MMP)2 antibody (Abcam, Cat#ab37150, 1: 4000), MMP9 antibody (Abcam, Cat#ab38898, 1: 4000), Histone H3 antibody (Abcam, Cat#ab8580, 1: 4000), and GAPDH antibody (Sigma-Aldrich, Cat#G9545, 1: 6000). Cell lines and treatment Human cervical cancer HeLa and SiHa cells were purchased from China Center for Type Culture Collection (CCTCC). Cells were maintained in Dulbecco modified eagle medium (DMEM, Hyclone) supplemented with 10% fetal bovine serum (FBS, Hyclone), penicillin (100 U/mL, Sigma-Aldrich) and streptomycin (100 mg/mL, Sigma-Aldrich) in a humidified cell incubator providing 5% Sunitinib CO2 and 95% fresh air at 37C. Cells were exposed to baicalin for 48 hours at 10, 20, 30, and 40 mmol/L. Cell viability assessments The cell viability was assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5 diphenylterazolium bromide (MTT) assay in accordance with previous descriptions. Briefly, cells were seeded into a 96-well cell culture plate at density of 5103/well. Cells were treated with baicalin as described. Cells in each well were incubated with MTT (5 mg/mL, Sigma-Aldrich) at 37C for 4 hours. The resulted formazan crystals were dissolved by dimethyl sulfoxide (DMSO). The absorbance at 490 nm was determined by a plate reader. The cell viability was calculated by the formula: (ODtreatment/ODcontrol)100%. Cell apoptosis detection Cell apoptosis was detected by terminal transferase UTP nick end labeling assay (TUNEL) assay. Cultured cells were fixed with neutral buffered formaldehyde. A TUNEL kit was used to detect the apoptotic cells in according to the protocol provided by the manufacturer. An inverted fluorescent microscope was used to observe the cells and to capture the fluorescent images. Cervical cancer cell migration evaluation In this study, wound healing FANCH assay was used to evaluate the migration ability of cervical cancer cells. Briefly, cells were seeded and further cultured in 60 mm-culturing dishes and received above described treatment of baicalin accordingly. The wound was formed by using a 2-mm-wide razor and the edges were marked. After treatment, the cells were fixed by neutral buffered formaldehyde which was subjected to 4,6-diamidino-2-phenylindole, dihydrochloride (DAPI) fluorescent staining. An inverted fluorescent microscope was used to observe the cells and to capture the fluorescent images. Wound closure was measured by analyzing the captured images. Cervical cancer cell invasion assessment The cell.