Fetal bovine serum (fbs)

The HL-1 cell line that is derived from mouse atrial myocytes was purchased from ATCC (USA). Cells were seeded into cell culture dishes and cultured in DMEM/F12 (Sigma-Aldrich, USA) supplemented with 10% fetal bovine serum (Clark Bioscience, USA) and 1% penicillin/streptomycin (Solarbio Biotechnology, China). The cells were maintained in an incubator containing 5% CO₂ at 37°C.
The cells were divided into 4 groups: CON, Ang II, SXSM, and SXSM + PMA. The CON group was the control group without intervention. The cells of the other groups were cultured for 12 hours and then treated with Ang II (Sigma-Aldrich, USA) for 12 hours. At the same time, SXSM was added to the cells of the SXSM group while SXSM and the PKC agonist PMA (Sigma-Aldrich, USA) were added to the SXSM + PMA group. The concentration of Ang II and PMA was 1 μM and the concentration of SXSM oral liquid that did not result in a cytotoxic effect was as determined in preliminary experiments.

HeLa and 293T cells were cultured in DMEM (Corning Cellgro, 15-017-CVR, Manassas, VA, USA) supplemented with 10% Fetal Bovine Serum (CLARK Bioscience, Richmond, VA, USA), 2 mM GlutaMAX (Gibco, Carlsbad, CA, USA), 100 units/mL of Penicillin-100 µg/mL of Streptomycin (HyClone, Logan, UT, USA). HeLa-DRD1-GFP-3FLAG, HeLa-DRD2-GFP-3FLAG, HeLa-GFP-DRD3-3FLAG, HeLa-DRD5-GFP-3FLAG, HeLa-GIPC1-GFP-3FLAG cells were maintained in DMEM complete medium with 1 μg/mL puromycin (Selleck, Washington, DC, USA). HeLa-GFP-DRD3-3FLAG cells were established as previously described [32 (link),58 (link)]. In addition, HeLa-DRD1/DRD2/DRD5/GIPC1-GFP-3FLAG cells were established similarly. The cDNA for DRD1/DRD2/DRD5/GIPC1 were amplified from HeLa cells and cloned into MSCV vector with GFP-3FLAG in their C-terminus.

Mouse mammary epithelial cells (mMECs) were purchased from ATCC (ATCC^® CRL-3063™) and cultured in DMEM F12 medium (Hyclone) supplemented with 10% (v/v) fetal bovine serum (CLARK) and antibiotics (100 U/ml penicillin and 100 µg/ml streptomycin sulfate) (Hyclone) at 37°C humidified incubator 5% CO₂.

The FLT3-ITD AML cell lines MOLM-13 and MV4-11 were purchased from AddexBio (San Diego, CA, USA; 2012) and American Type Culture Collection (ATCC, Manassas, VA, USA; 2006), respectively. FLT3 wild-type (FLT3-wt) AML cell line THP-1 was purchased from ATCC (2014 and 2002, respectively), while OCI-AML3 was purchased from the German Collection of Microorganisms and Cell Cultures (Braunschweig, Germany; 2011). All cell lines were cultured in RPMI 1640 media (except OCI-AML3, which was cultured in alpha-MEM), with 10–20% fetal bovine serum (CLARK Bioscience, Claymont, DE, USA), plus 2 mM L-glutamine, 100 U/mL penicillin, and 100 μg/mL streptomycin, in a humidified, 5%CO₂/95% environment at 37 °C. All lines were tested monthly for mycoplasma utilizing the PCR method described by Uphoff and Drexler^{15 (link)} and were authenticated in 2017 at the Karmanos Cancer Institute’s Genomics Core via the PowerPlex® 16 System (Promega, Madison, WI, USA).

HEK293T and YT cells were cultured in Dulbecco’s Modified Eagle Medium (DMEM) and Roswell Park Memorial Institute (RPMI) 1640, respectively (ThermoFisher, MA, USA) containing 10% fetal bovine serum (Clark Bioscience, VA, USA). SNK-6 cell line was cultured in RPMI 1640 containing 5% advanced cell culture supplement (AventaCell BioMedical Co., GA, USA) and 1000 U/ml recombinant human IL-2. In addition, 100 U/ml penicillin and streptomycin (ThemoFisher Scientific, MA, USA) were added in cell culture medium at 37 °C with 5% CO₂.

The murine hippocampal cell line HT22 was purchased from BNCC (Bena Culture Collection, Beijing, China) and grown in high glucose Dulbecco’s modified eagle medium (Gibco, California, United States), supplemented with 10% fetal bovine serum (CLARK, Virginia, United States) and streptomycin/penicillin (Biosharp, Anhui, China) in a humidified incubator with 5% CO₂ at 37°C.

In total, 87 patients with CRC were enrolled from the affiliated hospital of Jiangnan University, China between July 2008 and December 2009. Fresh tissues were harvested from the patients, snap-frozen, and preserved at −80°C. The clinical characteristics of the colorectal cancer patients were included in Table 1. The SW480 and CaCo-2 human colorectal cancer cell lines were maintained in our lab. The SW480 cells were cultured in RPMI 1640 medium. The CaCo-2 cells were maintained in MEM. We constructed SW480 cells that expressed high levels of B7-H3 (SW480-B7-H3) and CaCo-2 cells that were stably transfected with a B7-H3 shRNA (CaCo-2-shB7-H3). Cells that had been transfected with a mock vector were used as negative controls (SW480-NC and CaCo-2-NC). All of the media (HyClone, GE Healthcare Life Sciences, South Logan, UT, United States) were supplemented with 10% fetal bovine serum (Clark Bioscience, Houston, TX, United States). The cells were incubated at 37°C in a humidified atmosphere with 5% CO₂.