Caspase 9 p35 p10

Human colon cancer cell lines (HCT116 and DLD1) were purchased from Procell Company (Wuhan, China) and human colon mucosal epithelial cell line (NCM460) were purchased from Fenghui Biotechnology (Hunan, China). All cells were cultured in DMEM containing 10% fetal bovine serum (FBS) and 100 U/mL mixture of penicillin and streptomycin. The TDA is provided by the School of Pharmacy, Chongqing University. Cell counting kit 8 was purchased from MCE. Annexin V-FITC/PI Apoptosis Detection kit was purchased from CWBIO (Beijing, China). Primary antibodies for Western blotting, immunofluorescence and immunohistochemistry against target proteins are shown as follows: Caspase9/P35/P10 (Proteintech, Chicago, IL, USA, 66169), Caspase3 (Zen Bioscience, Chengdu, China, 300968), cleaved-Caspase3 (Zen Bioscience, 380169), PARP1 (Zen Bioscience, 380451), LC3 (Sangon Biotech, Shanghai, China, D163557), ATG5 (Sangon Biotech, D121650),SQSTM1/p62 (Beyotime, Shanghai, China, AF5312), β-actin (Santa Cruz, sc-47778), cyclinD1 (Cell Signaling Technology, USA, 2978S), cyclinB1 (Cell Signaling Technology, 4135S), PERK (Cell Signaling Technology, 3192S), IRE1 (Zen Bioscience, 220399), XBP1(Zen Bioscience, 381710), CHOP (Zen Bioscience, 381679) and HSPA5 (Sangon Biotech, D260466).

The human LC cell HepG2, Hep3B, and Huh-7 cell lines were purchased from China Infrastructure of Cell Line Resources (School of Basic Medicine Peking Union Medical College, China) and cultured in Dulbecco’s modified Eagle medium containing 10% (v/v) FBS and 100 U/ml streptomycin/penicillin at 37°C and 5% CO₂. Cell Counting Kit-8 (CCK-8) reagent was purchased from Dojindo (Japan). Hoechst 33,342 reagent was purchased from Beyotime (China). An Annexin-V FITC apoptosis detection kit (#556547) and BrdU cycle detection kit Part A (#559619) were purchased from BD Biosciences Pharmingen (United States). Antibodies against Bcl-2, Bax, Caspase 9/p35/p10, cyclin D1, and beta-actin were purchased from Proteintech (United States). Antibodies against cyclin A2, cyclin B1, p-Akt(S473), p-Akt(T308), pan-AKT, p-ERK (T202/Y204), and pan-ERK were purchased from Cell Signaling Technology (United States). Anti-CDK2 antibody was purchased from Abcam (United Kingdom). HSG was provided by Beijing Kangrentang Pharmaceutical Co., Ltd. The concentration of the prepared HSG mother solution was 0.1 g/mL. For the biological experiments, the 0.1 g/mL stock decoction could be subjected to 0.22-µm filter-sterilization and dilution as needed.