Journal: Frontiers in Pharmacology

Article Title: The Novel Antioxidant Compound JSH-23 Prevents Osteolysis by Scavenging ROS During Both Osteoclastogenesis and Osteoblastogenesis

doi: 10.3389/fphar.2021.734774

Figure Lengend Snippet: JSH-23 reduces RANKL-induced ROS production during osteoclastogenesis by downregulating the TRAF6/Rac1/NOX1 signaling pathway and enhancing the expression of Nrf2/HO-1. (A) Representative images of RANKL-induced ROS generation in BMMs with or without JSH-23 treatment at different concentrations. (B) Quantification of the average DCF intensity per well. (C) BMMs were stimulated with RANKL in the absence or presence of JSH-23 (20 and 40 μM) for 48 h, and then the ROS production pathway was analyzed by western blotting. (D) BMMs were cultured in α-MEM containing M-CSF and RANKL in the presence of JSH-23 (40 μM) at the indicated times (1, 3, 5 days). The expression of Nrf2 was analyzed by western blotting. (E) Western blotting images of the effects of JSH-23 on antioxidant enzyme, HO-1, catalase, NQO1, and GSR expression. (F – H) Quantitative analysis of the western blotting results. (I) Representative immunofluorescence images showing Nrf2 translocation and (J) HO-1 expression. (K – L) , BMMs were transfected with siRNA against Nrf2 or (M, N) HO-1 for 48 h. The silencing efficiency was evaluated by western blotting. (O) Transfected BMMs were treated with RANKL and JSH-23 (40 μM) for 5 days. TRAP staining was measured, and (P) the osteoclasts were counted. The values are shown as the means ± SDs, n = 3; * p < 0.05, ** p < 0.01.

Article Snippet: Alpha minimum essential medium (α-MEM), penicillin/streptomycin, and fetal bovine serum (FBS) were purchased from Excell (Excell Bio, Shanghai, China).

Techniques: Expressing, Western Blot, Cell Culture, Immunofluorescence, Translocation Assay, Transfection, Staining