Leptin protects rat articular chondrocytes from cytotoxicity induced by TNF-α in the presence of cyclohexamide

Abstract

Based on our findings, we suggest that the leptin present in the articular joint fluid protects articular chondrocytes against cumulative mechanical load and detrimental stresses throughout a lifetime, delaying the onset of degenerative changes in chondrocytes. We can further hypothesize that leptin protects articular chondrocytes against destructive stimuli even in the joints of osteoarthritis (OA) patients.

Alpha B-Crystallin Protects Rat Articular Chondrocytes against Casein Kinase II Inhibition-Induced Apoptosis

“…As described previously [5], five-week-old male, specific pathogen-free Sprague Dawley rats were obtained from Samtoko (Osan, Korea). Rat articular chondrocytes for primary culture were isolated from knee joint cartilage slices via enzymatic digestion for 1 h with 0.2% type II collagenase (381 units/mg) in DMEM.…”

“…As described previously [5], cell viability was determined with a Vi-Cell (Beckman Coulter, CA, USA) cell counter, which performs an automated trypan blue exclusion assay.…”

“…As described previously [5], cell suspensions were cytospun onto clean fat-free glass slides using a cytocentrifuge. Centrifuged samples were fixed in 4% paraformaldehyde for 10 min and stained with 10 μg/ml PI or in 4 μg/ml Hoechst 33342 for 30 min at 4°C.…”

“…As described previously [5], cells cultured on coverslips were incubated with a primary antibody for 1 h at 37°C, washed 3 times for 5 min each with PBS, incubated with a FITC-conjugated secondary antibody for 1 h at room temperature and counterstained with propidium iodide. For double immunofluorescent staining, cells were incubated with two different primary antibodies for 1 h at 37°C and washed three times for 5 min each with PBS.…”

“…As described previously [5], MMP disruption was measured using a specific fluorescent probe, JC-1, added directly to the cell culture medium (5 μg/ml final concentration) and incubated for 15 min at 37°C. Flow cytometry was used to measure JC-1 signals (Epics XL; Beckman Coulter).…”