Penicillin streptomycin

All animal experiments were performed based on the Animal Ethics Committee standards of Guangxi Medical University (Protocol Number: SCXK-Gui-2019-0211). BMSCs were extracted from approximately week-old Sprague-Dawley (SD) rats. The BMSCs were cultured in α-minimal essential medium (α-MEM, Biosharp, China) with 10% foetal bovine serum (Tianhang, China) and 1% streptomycin/penicillin (Biosharp, China) in an incubator with 5% CO₂ at 37 ℃. The medium was replaced every 3 days.

Fourteen HNSCC cell lines, namely HSC-4, CAL-27, UM1, HN4, HN12, HN30, CAL-33, SCC-47, SCC-9, H314, H103, H357, H376, and SCC-25 were utilized for this study. Eight of these lines (HSC-4, CAL-27, UM1, HN4, HN12, HN30, CAL-33, and SCC-47) were maintained in DMEM/high-glucose (Cytiva, Logan, USA) supplemented with 10% fetal bovine serum (FBS, VivaCell, Shanghai, China) and 1% streptomycin–penicillin (Biosharp, Hefei, China). The remaining six lines, SCC-9, H314, H103, H357, H376, and SCC25, were grown in F12 medium (VivaCell, Shanghai, China) enriched with 10% FBS, 1% streptomycin–penicillin, and 0.5 µg/mL hydrocortisone. The cells were incubated in a humidified environment at 37 °C with 5% CO2. All these cells were carefully maintained in the central cell bank of State Key Laboratory of Oral Diseases, West China Hospital of Stomatology in Sichuan University. Cell lines were routinely tested to be negative for mycoplasma, and low passage cultures were applied in this experiment.

The murine hippocampal cell line, HT22 was purchased from BNCC (Bena Culture Collection, China). Cells were grown in high glucose Dulbecco's modified Eagle's medium (DMEM; Gibco) supplemented with 10% fetal bovine serum (CLARK) and streptomycin/penicillin (Biosharp, China) in a humidified incubator with 5% CO₂ at 37°C.
Homer1b/c overexpression plasmid and siRNA were designed and manufactured by Shanghai Genechem Co., Ltd. (Shanghai, China) and HANBI Co., Ltd. (Shanghai, China), respectively. HT22 cells were transfected using Lipofectamine 2000 (Invitrogen) according to the manufacturer's instructions. Experiments using transfected cells were performed 24 h after transfection.

U87MG and U251 cells were purchased from Procell (Wuhan, China) and cultivated in high glucose DMEM (Gibco, Carlsbad, CA, USA), appended by 10% fetal bovine serum (CLARK Bioscience, Richmond, VA, USA) and 1% antibiotics (streptomycin/penicillin, Biosharp, Hefei, Anhui, China) in a humidified incubator at 37 °C and 5% CO₂.

Caki-1, 786-O, RCC-JF, RCC-23 (ccRCC cell lines), HK-2 (adult renal tubular epithelial cells) and 293 T (normal human embryonic kidney cell lines) were purchased from Meisen CTCC (Zhejiang, China). All cell lines were authenticated via STR profiling. 293 T and HK-2 cells were cultured in DMEM (HyClone, Logan, Utah, USA), Caki-1 cells were cultured in McCoy’s 5A (Basal Media, Shanghai, China), and 786-O, RCC-23 and RCC-JF cells were cultured in RPMI-1640 (HyClone). All media were supplemented with 10% FBS (PAN, Aidenbach, Germany), 1% penicillin/streptomycin (Biosharp, China) and 1‰ Mycoplasma Elimination Reagent (Yeasen, Shanghai, China) to remove mycoplasma. The cells were cultured in 5% CO₂ at 37 °C.

Antibodies used in the experiment included p-STAT3 (Tyr705) (Cat. 9145, CTS; Clone D3A7; 1:3,000 dilution for IHF staining), anti-TNF-α (Cat. GB11188, Service; 1:100 dilution for IHF staining), anti-IL-1β (Cat. GB11113, Service; 1:100 dilution for IHF staining) and p-HSL (Cat. 4126S, CST;1:100 dilution for IHF staining). Celastrol (98.0%) was purchased from Sigma. Glycyrrhetinic acid was from Huaian Brothers Biological Science. Foetal bovine serum (FBS) was from NTC (NCT-HK014, Natocor, Cordoba, Argentina). DMEM (SH30022.01) was purchased from Hyclone. Penicillin/streptomycin was obtained from Biosharp (Shanghai, China). Hydrocortisone and recombinant hEGF were purchased from Sangon Biotech (Shanghai, China). FITC, MTT and DMSO were all purchased from Sigma-Aldrich (China).