Sgc 996

The human cell lines GBC-SD and SGC-996 were purchased from the Shanghai Cell Institute Country Cell Bank. GBC-SD cells were cultured in high-glucose DMEM (Gibco, USA) at 37°C in a 5% CO₂ incubator, while SGC-996 cells were cultured in Roswell Park Memorial Institute (RPMI) 1640 medium. Both types of culture medium were supplemented with 10% fetal bovine serum (Gibco, USA), penicillin (100 U/mL) and streptomycin (100 U/mL) (Utah, HyClone, USA).

Three human gallbladder cancer cell lines were used: GBC-SD (Cell Bank of the Chinese Academy of Sciences, Shanghai, China); NOZ (Health Science Research Resources Bank, Osaka, Japan); and SGC-996 (Academy of Life Science, Tongji University, Shanghai, China). CV-1 (monkey kidney), NIH3T3 (murine fibroblast) and U251 (human giloma) cell lines were purchased from the Cell Bank of the Chinese Academy of Sciences. GBC-SD, NOZ, and NIH3T3 cells were cultured in DMEM (Gibco BRL, Carlsbad, CA, USA) containing 15% FBS (HyClone, Logan, UT, USA). SGC-996, CV-1 and U251 cells were cultured in RPMI medium 1640 (Gibco BRL) with 15% FBS at 37°C and 5% CO₂.

20 GBC tissues and the matching bile used in present study were obtained from the patients admitted to Fujian Medical University Union Hospital in China. The informed consents of agreement to use the samples for further study were signed pre-operation. The samples were collected according to the protocol approved by the Ethics Committee of the Medical Faculty of Fujian Medical University, according to the Declaration of Helsinki. The details of the patients including the age and sex of the patient, clinical stage, grade of the tumor and lymph node metastasis (LNM) had been described in [20 (link)]. The human GBC cell lines: NOZ (obtained from Health Science Research Resources Bank in Japan), GBC-SD (purchased from Shanghai Institutes for BiologicalSciences in China) and SGC-996 (provided by the Tumor Cytology Research Unit, Medical College, Tongji University, China) were maintained in Dulbecco’s Modified Eagle’s Medium (Gibco, USA) supplemented with 10 % fetal bovine serum (Gibco). Human dermal lymphatic endothelial cells (HDLECs, purchased from Sciencell, San Diego, California, USA) were incubated in endothelial cell medium (Sciencell). All of the cells were incubated at 37 °C under 95 % air and 5 % CO2.

NOZ cells were obtained from the Health Science Research Resources Bank (Osaka, Japan), GBC-SD, SGC-996, and EH-GB1 cells were purchased from the Cell Bank of Type Culture Collection of Chinese Academy of Sciences, and human embryonic kidney 293 T (HEK293T) cells were purchased from the American Type Culture Collection. NOZ and GBC-SD both were P53 WT genotype (P53^+/+). GBC-SD, SGC-996, EH-GB1, and HEK293T cells were cultured in Dulbecco’s modified Eagle’s medium (Gibco), and NOZ cells were cultured in William’s E medium (Gibco). All cell lines were supplemented with 10% fetal bovine serum (Gibco), penicillin (100 mg ml⁻¹) and streptomycin (100 mg ml⁻¹) and were incubated in a humidified chamber with 5% CO₂ at 37 °C. All cell cultures were ensured to be mycoplasma-negative cultures by monthly mycoplasma tests and were passaged with 0.25% trypsin containing 2.21 mm ethylenediaminetetraacetic acid (EDTA) in PBS when the cells reached 80~90% confluency. Gemcitabine (GEMZAR) was purchased from Eli Lilly. Cisplatin, cycloheximide, doxorubicin, and puromycin were purchased from MedChem Express.

The human GBC cell lines NOZ, EH-GB-1, EH-GB-2, SGC-996 and GBC-SD were purchased from the Cell Bank of Type Culture Collection of the Chinese Academy of Sciences (Shanghai, China). The OCUG-1 cell line was obtained from the Health Science Research Resources Bank (Osaka, Japan). The NOZ cell line was maintained in William’s medium (Gibco, Grand Island, NY, USA) supplemented with 10% fetal bovine serum (FBS; Gibco). The GBC-SD cells were maintained in DMEM medium (Gibco). The EH-GB-1, EH-GB-2, SGC-996 and OCUG-1 cells were cultured in RPMI 1640 (Gibco). DHA was purchased from Selleck Chemicals and dissolved in DMSO. Primary antibodies against TCTP, vinculin, paxillin and β-actin were purchased from Cell Signaling Technology (Beverly, MA, USA).

Two human GBC cell lines were used in the experiment. SGC-996 and GBC-SD were purchased from Cell Bank of the Chinese Academy of Science (Shanghai, China). SGC-996 and GBC-SD cell lines were cultured in Dulbecco’s modified Eagle’s medium (DMEM, Gibco BRL), containg 10% fetal bovine serum (FBS, HyClone) as well as 100U/ml penicillin and 100 μg/ml streptomycin. Cells were maintained at 37°C in 5% CO₂. Sesamin (>94% purity) was provided by Tianyi Lvbao Technology Co. (Wuhu, China) [11 (link)]. Sesamin was dissolved in DMSO as 0, 11, 33.3, 100 μM stock solution. Vehicle control consisted of DMSO equivalent to treatments.