TNF Receptor II Antibody - #AF0364

Fig. 6 Atsttrin enhances osteoblastogenesis through TNFR2. a, b MC3T3-E1 cells were treated with Atsttrin (500 ng/ml) for 48 h. The protein level of RUNX2 was assessed by Western blotting (n = 3). Quantification of the band density for RUNX2 based on the Western blotting assay. c, d BMMSCs were treated with Atsttrin (500 ng/ml) for 7 days, and ALP staining was performed. Scale bar, 200 µm. Each experiment was performed three times independently. Quantification of the percentage of the positive area was based on ALP staining. e–g MC3T3-E1 cells were cultured with Atsttrin (500 ng/ml) for 8 h. The mRNA levels of ALP, RUNX2, and Col-1 were detected by real-time PCR (n = 3). h MC3T3-E1 cells were treated with Atsttrin (500 ng/ml) for 7 days. The relative fold ALP activity was detected by an ALP assay kit (n = 3). i Knockout efficiency of TNFR2 using siRNA in MC3T3-E1 cells and BMMSCs, as assayed by immunoblotting analysis. j, k MC3T3-E1 cells transfected with scRNAi or TNFR2 RNAi were treated with Atsttrin (500 ng/ml) for 48 h. The protein was examined by Western blotting with an anti-RUNX2 antibody (n = 3). Quantification of the band density for RUNX2 based on the Western blotting assay. l, m BMMSCs transfected with scRNAi or TNFR2 RNAi were treated with Atsttrin (500 ng/ml) for 7 days, and ALP staining was performed. Scale bar, 200 µm. Each experiment was performed three times independently. Quantification of the percentage of the positive area was based on ALP staining. n–p MC3T3-E1 cells transfected with scRNAi or TNFR2 RNAi were treated with Atsttrin (500 ng/ml) for 8 h. The mRNA levels of ALP, RUNX2, and Col-1 were measured by real-time PCR (n = 3).