Minimal essential medium alpha αmem

PDLCs were obtained from impacted human wisdom teeth removed at the Department of Dentoalevolar Surgery of the Semmelweis University (Budapest, Hungary) under ethical guidelines approved by the Ethical Committee of the Hungarian Medical Research Council. This study was approved by the Semmelweis University Regional and Institutional Committee of Science and Research Ethics, the number of the ethical permissions are 17458/2012/EKU and 25459/2019/EKU. Isolation of PDLCs was performed according to our previously published protocol [15 (link),81 (link)].
The PDLCs were cultured in Minimal Essential Medium Alpha (αMEM, Gibco, Thermo Fisher, Waltham, USA) supplemented with 10% fetal bovine serum (FBS, Gibco), 2 mM L-glutamine (Gibco), 100 units/mL penicillin and 100 mg/mL streptomycin (Gibco).
The SH-SY5Y human neuroblastoma cell line was purchased from Sigma-Aldrich (Budapest, Hungary) and maintained according to the manufacturer’s protocol. The growth medium of these cells consisted of a 1:1 mixture of Eagle’s MEM and Ham’s F12 medium supplemented with 15% FBS (Gibco), 2 mM L-glutamine (Gibco), 100 units/mL penicillin, and 100 mg/mL streptomycin (Gibco), as well as 1% non-essential amino acids (NEAA, Gibco).

Escherichia coli-derived lipopolysaccharide (LPS) was obtained from InvivoGen (San Diego, CA, USA). Recombinant mouse RANKL and macrophage colony-stimulating factor (M-CSF) were purchased from R&D (Minneapolis, MN, USA). Penicillin, streptomycin, and minimal essential medium alpha (α-MEM) were purchased from Gibco BRL (Gaithersburg, MD, USA). Fetal bovine serum (FBS) was purchased from Avantor (Radnor, PA, USA). The Cell Counting Kit-8 (CCK-8) was obtained from Beyotime Biotechnology (Shanghai, China). Methylisothiourea sulfate (SMT) was purchased from Selleck (Houston, TX, USA). Dihydroethidium (DHE) was purchased from Med Chem Express (Shanghai, China). The Prime Script RT reagent Kit and TB Green^TM Premix Ex Taq^TM II were obtained from Takara Biomedical Technology (Beijing, China). The primary antibody against iNOS (Thermofisher #PA1-036) was purchased from Thermofisher Scientific (Waltham, MA, USA). Primary antibodies against phospho-p65 (CST #3033), p65 (CST #8242), Keap1 (CST #4678), Nrf2 (CST #12721), HO-1 (CST #26416), and NQO1 (CST #62262) were purchased from Cell Signaling Technology (CST, Danvers, MA, USA). Primary antibodies against β-Actin (Affinity #AF7018), GAPDH (Affinity #AF7021), NFATc1 (Affinity #DF6446), CTSK (Affinity #DF6614), and TNF-α (Affinity #AF7014) were purchased from Affinity Biosciences (Cincinnati, OH, USA).

The murine-derived macrophage cell line RAW 264.7 cells (RAW cells, Cell Bank, purchased from the Chinese Academy of Sciences) and human bone marrow mesenchymal stem cells (hBMMSCs) were purchased from Cyagen Biosciences Inc. (Guangzhou, China). The RAW 264.7 cells were cultured using the Dulbecco’s modified Eagle’s medium (DMEM, Gibco) supplemented with 10% (v/v) fetal bovine serum (FBS, Gibco) and antibiotics (100 U/ml of penicillin and 100 mg/ml of streptomycin) (Thermo Fisher Scientific, United States). The hBMMSCs were cultured using Minimal Essential Medium Alpha (α-MEM, Gibco) supplemented with 10% (v/v) FBS and antibiotics (100 U/ml of penicillin and 100 mg/ml of streptomycin).