The human chondrocytes cultures were obtained from patients who underwent joint
replacement therapy. The osteoarthritic cartilage harvested from patients was
turned into pieces and digested with an enzymatic solution [8 mg/ml
hyaluronidase (Sigma-Aldrich, St. Louis, MO, USA), 8 mg/ml collagenase
(Sigma-Aldrich), and 2.5 mg/ml trypsin (Sigma-Aldrich)] for 6 h at 37°C. The
cellular suspension was centrifuged at 1,500 rpm for 5 min and resuspended into
Dulbecco’s modified Eagle’s medium (DMEM)/F12 (Gibco BRL, New York, NY, USA)
with 10% fetal bovine serum (FBS; Gibco BRL), and 1%
Penicillin-Streptomycin-Amphotericin B (PSA) (Biological Industries, Beit
Haemek, Israel) in a humidified atmosphere containing 5% CO2.
Thereafter, the primary OA chondrocytes were cultured, passaged, and maintained
in DMEM/F12 medium.
Based on previous reports showing various inhibitory activities of nicotine
against bone density19 (link), human renal proximal tubular epithelial cells20 (link), and myoblast differentiation21 (link), we firstly established in vitro model of nicotine
(Sigma 36733) OA, by treatment of various doses of nicotine (100, 500, and 1,000
µM) to chondrocytes (passage 2) in the culture medium. After validating OA
properties in nicotine-treated chondrocytes, we further treated them with PDB
for 7 days.
replacement therapy. The osteoarthritic cartilage harvested from patients was
turned into pieces and digested with an enzymatic solution [8 mg/ml
hyaluronidase (Sigma-Aldrich, St. Louis, MO, USA), 8 mg/ml collagenase
(Sigma-Aldrich), and 2.5 mg/ml trypsin (Sigma-Aldrich)] for 6 h at 37°C. The
cellular suspension was centrifuged at 1,500 rpm for 5 min and resuspended into
Dulbecco’s modified Eagle’s medium (DMEM)/F12 (Gibco BRL, New York, NY, USA)
with 10% fetal bovine serum (FBS; Gibco BRL), and 1%
Penicillin-Streptomycin-Amphotericin B (PSA) (Biological Industries, Beit
Haemek, Israel) in a humidified atmosphere containing 5% CO2.
Thereafter, the primary OA chondrocytes were cultured, passaged, and maintained
in DMEM/F12 medium.
Based on previous reports showing various inhibitory activities of nicotine
against bone density19 (link), human renal proximal tubular epithelial cells20 (link), and myoblast differentiation21 (link), we firstly established in vitro model of nicotine
(Sigma 36733) OA, by treatment of various doses of nicotine (100, 500, and 1,000
µM) to chondrocytes (passage 2) in the culture medium. After validating OA
properties in nicotine-treated chondrocytes, we further treated them with PDB
for 7 days.
