Nc sirna si nc

Small interfering RNAs (siRNAs) targeting VPS9D1-AS1 (si-VPS9D1-AS1) and negative control (NC) siRNA (si-NC) were obtained from GenePharma Technology (Shanghai, China). miR-525-5p mimic and miR-525-5p inhibitor were synthesized by RiboBio Biotechnology (Guangzhou, China) and used to increase and decrease endogenous expression levels of miR-525-5p, respectively. miRNA mimic NC (miR-NC) and NC inhibitor were used as controls. The HMGA1 overexpression plasmid pcDNA3.1-HMGA1 and empty pcDNA3.1 plasmid were designed and generated by Genearray Biotechnology (Shanghai, China). Lipofectamine™ 2000 was used for all cell transfections according to the manufacturer’s protocol.

Normal gingival epithelial cells (ATCC^® PCS-200-014™) were acquired from the American Type Culture Collection (ATCC; Manassas, VA, USA) and cultured in minimum essential medium (Gibco; Thermo Fisher Scientific, Inc., Waltham, MA, USA) containing 10% fetal bovine serum (FBS; Gibco; Thermo Fisher Scientific). Dulbecco’s modified Eagle’s medium (DMEM; Gibco; Thermo Fisher Scientific) with 10% FBS was employed in the culture of the TSCC cell line CAL-27 (ATCC). The other two TSCC cell lines SCC-9 and SCC-15 were maintained in a 1:1 mixture of DMEM and Ham’s F12 medium (Gibco; Thermo Fisher Scientific) containing 10% FBS and 400 ng/mL hydrocortisone. All cells were incubated at 37°C in a humidified incubator supplied with 5% CO₂.
Small interfering RNAs (siRNAs) for NOP14-AS1 (si-NOP14-AS1) and negative control (NC) siRNA (si-NC) were designed and offered by Shanghai GenePharma Co., Ltd. (Shanghai, China). The overexpressed high mobility group box 3 (HMGB3) plasmid pcDNA3.1-HMGB3 was constructed by RiboBio (Guangzhou, China). miRNA-665 (miR-665) mimic, NC mimic, miR-665 inhibitor, and NC inhibitor were also obtained from Shanghai GenePharma. TSCC cells were seeded into six-well plates, and transfection was performed by employing Lipofectamine 2000 (Invitrogen; Thermo Fisher Scientific).