Journal: Molecular Medicine

Article Title: Tetrandrine inhibits RANKL-induced osteoclastogenesis by promoting the degradation of TRAIL

doi: 10.1186/s10020-022-00568-4

Figure Lengend Snippet: Tetrandrine inhibit RANKL-induced osteoclastogenesis. A The chemical structural formula of Tetrandrine; B Tetrandrine impact on BMMs viability was detected by CCK-8 assay at 48 h and 96 h. C With different dilutions of Tetrandrine, M-CSF and RANKL for 7 days, BMMs were processed; scale bar = 200 μm. D The number and area of TRAP-positive cells. E TRAP-positive BMMs after the treatment with 1 μM Tetrandrine for the designated days in the process of osteoclastogenesis; scale bar = 200 μm. F To quantify TRAP-positive cells, the number and area of osteoclasts. G Immunofluorescence image for showing the formation of F-actin. All experiments were carried out in triplicate. The results were presented as mean ± SEM. * P < 0.05, ** P < 0.01, *** P < 0.005 and **** P < 0.001 as compared with the control group

Article Snippet: Briefly, 1 × 10 5 cells were added to each well of a 96-well plate and cultivated for 48 and 96 h with Tetrandrine (Selleck, Houston, Texas, United States).

Techniques: CCK-8 Assay, Immunofluorescence, Control

Journal: Molecular Medicine

Article Title: Tetrandrine inhibits RANKL-induced osteoclastogenesis by promoting the degradation of TRAIL

doi: 10.1186/s10020-022-00568-4

Figure Lengend Snippet: Tetrandrine inhibits osteoclastic bone resportion and expression of osteoclastogenesis related genes. A BMMs were planted into slices of bone and treated with different concentrations of Tetrandrine for 7 days and bone resorption pit pictures taken by a scanning electron microscope are presented; scale bar = 500 μm. The pit zone resorption area were shown by the ImageJ. B The detection of expression levels of osteoclastogenesis related genes in BMMs with 0, 0.25, 0.5, 1 μM was carried out with qPCR. C After treating BMMs with Tetrandrine (1 μM) for 0, 1, 3, 5 days, they were detected via RT-qPCR. The mRNA expression levels of NFATc1, c-Fos, DC-STAMP, TRAP, CTSK were evaluated. D For 0, 1, 3, 5 days with or without 1 μM of Tetrandrine, the treatment of BMMs was carried out with RANKL. The protein expression levels of NFATc1, TRAP and CTSK were evaluated. All experiments were carried out in triplicate. The results were presented as mean ± SEM. * P < 0.05, ** P < 0.01, *** P < 0.005 and **** P < 0.001 as compared with the control group

Article Snippet: Briefly, 1 × 10 5 cells were added to each well of a 96-well plate and cultivated for 48 and 96 h with Tetrandrine (Selleck, Houston, Texas, United States).

Techniques: Expressing, Microscopy, Quantitative RT-PCR, Control

Journal: Molecular Medicine

Article Title: Tetrandrine inhibits RANKL-induced osteoclastogenesis by promoting the degradation of TRAIL

doi: 10.1186/s10020-022-00568-4

Figure Lengend Snippet: Tetrandrine affects osteoclastogenesis by promoting the ubiquitination of TRAIL. A The mRNA expression level of TRAIL in BMMs with 0, 0.25, 0.5, 1 μM of Tetrandrine was carried out with RT-qPCR. B The expression of level of TRAIL in BMMs treated with specified concentrations of Tetrandrine for 5 days was detected using western blot. C Immunofluorescence image for showing TRAIL expression in cytoplasm; scale bar = 200 μm. D The co-IP assay was used to detect the effect of Tetrandrine on TRAIL ubiquitination with or without RANKL. E–G The transfection efficiency of TRAIL silencing (si #1, si #2, si #3) and TRAIL overexpression (TRAIL vector) detected via RT-qPCR and western blotting. H–K BMMs were transfected with siTRAIL and TRAIL vector, the expression of the related genes and proteins that were revealed by RT-qPCR and western blotting. All experiments were carried out in triplicate. The results were presented as mean ± SEM. * P < 0.05, ** P < 0.01, *** P < 0.005 and **** P < 0.001 as compared with the control group

Article Snippet: Briefly, 1 × 10 5 cells were added to each well of a 96-well plate and cultivated for 48 and 96 h with Tetrandrine (Selleck, Houston, Texas, United States).

Techniques: Ubiquitin Proteomics, Expressing, Quantitative RT-PCR, Western Blot, Immunofluorescence, Co-Immunoprecipitation Assay, Transfection, Over Expression, Plasmid Preparation, Control

Journal: Molecular Medicine

Article Title: Tetrandrine inhibits RANKL-induced osteoclastogenesis by promoting the degradation of TRAIL

doi: 10.1186/s10020-022-00568-4

Figure Lengend Snippet: TRAIL overexpression partially rescued the inhibitory effect of Tetrandrine on osteoclastogenesis. A The mRNA expression of c-Fos, CTSK, DC-STAMP, NFATc1 and TRAP in BMMs in rescue experiment was detected by RT-qPCR. B Western blotting demonstrated the expression of NFATc1, TRAP and CTSK in BMMs in rescue experiment. C The number and area of TRAP-positive cells, the bone resorption pits and F-actin formation in rescue experiment; Scale bar in TRAP staining and immunofluorescence image = 200 μm; scale bar in bone resorption pit picture = 500 μm. All experiments were carried out in triplicate. The results were presented as mean ± SEM. * P < 0.05, ** P < 0.01, *** P < 0.005 and **** P < 0.001 as compared with the control group

Article Snippet: Briefly, 1 × 10 5 cells were added to each well of a 96-well plate and cultivated for 48 and 96 h with Tetrandrine (Selleck, Houston, Texas, United States).

Techniques: Over Expression, Expressing, Quantitative RT-PCR, Western Blot, Staining, Immunofluorescence, Control

Journal: Molecular Medicine

Article Title: Tetrandrine inhibits RANKL-induced osteoclastogenesis by promoting the degradation of TRAIL

doi: 10.1186/s10020-022-00568-4

Figure Lengend Snippet: Tetrandrine inhibits NF-κB and MAPK pathways by downregulating TRAIL expression. A Treatment of BMMs was carried out with RANKL, with or without 0.25 μM Tetrandrine for 0, 30 and 60 min. The p-P65, p-JNK, p-ERK, p-P38, p-IκBα, p-IKKα/β protein levels were evaluated. B The p-P65, p-JNK, p-P38, p-IκBα, p-IKKα/β protein levels were determined by western blotting in rescue experiment. All experiments were carried out in triplicate. The results were presented as mean ± SEM. * P < 0.05, ** P < 0.01, *** P < 0.005 and **** P < 0.001 as compared with the control group

Article Snippet: Briefly, 1 × 10 5 cells were added to each well of a 96-well plate and cultivated for 48 and 96 h with Tetrandrine (Selleck, Houston, Texas, United States).

Techniques: Expressing, Western Blot, Control

Journal: Molecular Medicine

Article Title: Tetrandrine inhibits RANKL-induced osteoclastogenesis by promoting the degradation of TRAIL

doi: 10.1186/s10020-022-00568-4

Figure Lengend Snippet: Tetrandrine promotes C3H10 osteoblastogenesis and inhibits TRAIL expression in C3H10. A Effect of Tetrandrine on C3H10 viability by CCK-8 assay at 48 h and 96 h. B The TRAIL, OCN, ALP and OPG expression levels in C3H10 treated with Tetrandrine were detected with RT-qPCR. C The protein levels of COL1a, RUNX2, OCN, TRAIL, RANKL and OPG were measured with western blotting. D , E ALP and ARS expression in C3H10 following the treatment with various concentration of Tetrandrine for 7 days. The mineralized extracellular matrix was detected with Alizarin Red S staining for 14 days. The OD values obtained for ALP solutions and mineralized matrix solutions after their treatment with Tetrandrine; scale bar = 200 μm. All experiments were carried out in triplicate. The results were presented as mean ± SEM. * P < 0.05, ** P < 0.01, *** P < 0.005 and **** P < 0.001 as compared with the control group

Article Snippet: Briefly, 1 × 10 5 cells were added to each well of a 96-well plate and cultivated for 48 and 96 h with Tetrandrine (Selleck, Houston, Texas, United States).

Techniques: Expressing, CCK-8 Assay, Quantitative RT-PCR, Western Blot, Concentration Assay, Staining, Control

Journal: Molecular Medicine

Article Title: Tetrandrine inhibits RANKL-induced osteoclastogenesis by promoting the degradation of TRAIL

doi: 10.1186/s10020-022-00568-4

Figure Lengend Snippet: Tetrandrine prevents OVX-induced bone loss in vivo. A Weight change diagram of mice after Tetrandrine treatment (each; n = 5). B RANKL/OPG, TNFα and IL-6 levels in serum of mice were determined with Elisa assay. C , D Detection of the tetrandrine effects on the expression of TRAIL in the OVX-induced model carried out with RT-qPCR and western blot. E Detection of NFATc1, DC-STAMP, TRAP and CTSK expression levels was carried out with RT-qPCR. F Calcein-alizarin red S labeling to detect the femur mineral apposition rate (MAR) in mice. G TRAP staining was used for sections of femurs and number of osteoclasts per field of each specimen were measured. H The Von kossa staining was used for the sections of femurs and the MS/BS (%) was measured. All experiments were carried out in triplicate. The results were presented as mean ± SEM. * P < 0.05, ** P < 0.01, *** P < 0.005 and **** P < 0.001 as compared with the control group

Article Snippet: Briefly, 1 × 10 5 cells were added to each well of a 96-well plate and cultivated for 48 and 96 h with Tetrandrine (Selleck, Houston, Texas, United States).

Techniques: In Vivo, Enzyme-linked Immunosorbent Assay, Expressing, Quantitative RT-PCR, Western Blot, Labeling, Staining, Control

Journal: Molecular Medicine

Article Title: Tetrandrine inhibits RANKL-induced osteoclastogenesis by promoting the degradation of TRAIL

doi: 10.1186/s10020-022-00568-4

Figure Lengend Snippet: Tetrandrine inhibits OVX-induced osteoporosis in mice model. A H&E staining was used for several major organs of mice in three groups (each; n = 5). B , C The micro-CT was used for the scanning tibias of each mouse, and the BS/BV, Tb.N, Tb.Sp and Tb.Th values were recorded for each specimen. D , E H&E and Masson staining was also used for the femurs. All experiments were carried out in triplicate. The results were presented as mean ± SEM. * P < 0.05 and ** P < 0.01 as compared with the control group

Article Snippet: Briefly, 1 × 10 5 cells were added to each well of a 96-well plate and cultivated for 48 and 96 h with Tetrandrine (Selleck, Houston, Texas, United States).

Techniques: Staining, Micro-CT, Control

Journal: Molecular Medicine

Article Title: Tetrandrine inhibits RANKL-induced osteoclastogenesis by promoting the degradation of TRAIL

doi: 10.1186/s10020-022-00568-4

Figure Lengend Snippet: Schematic illustration for the mechanism by which Tetrandrine inhibits RANKL-induced osteoclastogenesis by promoting the degradation of TRAIL. RANKL recruits TRAF6 by interacting with its receptor RANK, and then activates downstream effectors such as NF-κB and MAPKs to promote mature osteoclast formation. Inflammatory factors including IL-6 and TNFα play an active role in the binding of RANK and RANKL and Tetrandrine has obvious anti-inflammatory effect. More importantly, Tetrandrine promotes TRAIL degradation and TRAIL downregulation inhibits the recruitment of TRAF6 by RANKL and eventually dampen osteoclast different and function. Besides, Tetrandrine can stimulate osteoblast difference

Article Snippet: Briefly, 1 × 10 5 cells were added to each well of a 96-well plate and cultivated for 48 and 96 h with Tetrandrine (Selleck, Houston, Texas, United States).

Techniques: Binding Assay