Fig. 2

The influence of TNFα expressed by BV2 cells for VSMC calcification. a-f Alizarin red staining analysis of VSMC calcification in differently stimulated condition (40×), a group1 added 10% BV2 culture supernatant and 10 mM β-glycerophosphate, b group3 added 10 mM β-glycerophosphate and 10 ng/ml mouse TNFα, c group5 10 mM β-glycerophosphate, d group2 added 10% BV2 culture supernatant, which was incubated with rabbit anti-mouse TNFα antibody, and 10 mM β-glycerophosphate, e group4 added 10 ng/ml mouse TNFα, f group6 added equal volume PBS. g-i qRT-PCR analysis for the expression of Runx2 and α-SMA in VSMC, g the mRNA expression of Runx2 and α-SMA in groups 1 and 2, h the mRNA expression of Runx2 and α-SMA in groups 3 and 4, i the mRNA expression of Runx2 and α-SMA in groups 5 and 6. Bars represent mean ± S.D. (n ≥ 3). Significant difference marked with one (p < 0.05) or two (p < 0.01) asterisks