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. 2017 Jun 14;12(6):e0179512. doi: 10.1371/journal.pone.0179512

Fig 1. In vitro analysis of WT and Col1a1-C5aR1 osteoblasts.

Fig 1

(A) Western blot showing C5aR1 expression in primary osteoblasts of wildtype (WT) and Col1a1-C5aR1 mice after cultivating for 14 d in normal proliferation medium (NM) and osteogenic differentiation medium (ODM). (B) Cell proliferation of WT and Col1a1-C5aR1 osteoblasts determined by BrdU incorporation. (C) Osteogenic differentiation capacity of osteoblasts assessed by alkaline-phosphatase staining after cultivating for 14 d in ODM with or without C5a and (D) by the expression of the osteogenic marker genes Alpl (alkaline phosphatase), Ibsp (bone sialoprotein) and Bglap (bone gamma carboxyglutamate protein/osteocalcin). (E) Western blot showing increased expression of phospho (p)-ERK 1/2 and phospho (p)-p38 after osteoblast stimulation with C5a for 6 h. F) Relative gene expression of interleukin (IL)-6, receptor activator of NF-κB ligand (RANKL) and osteoprotegerin (OPG). *p < 0.05; ***p < 0.001, n = 5–6 per group and treatment.