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TGF-β Inducible Early Gene 1 Regulates Osteoclast Differentiation and Survival by Mediating the NFATc1, AKT, and MEK/ERK Signaling Pathways

Figure 1

Lack of TIEG1 in osteoclast precursors delays osteoclast differentiation and apoptosis in vitro.

A. WT and TIEG1−/− marrow cells were cultured as described in the Methods section and subsequently fixed and stained for TRAP activity and chromatin condensation beginning on day 3 or after feeding the cells with MCSF and RANKL for the indicated time in hours (h). These data are representative of three separate experiments. B. Mean +/− SD of viable osteoclasts over time. These data were obtained from four replicate wells (p<0.05) and are representative of three separate experiments. C. Image of TRAP and Hoechst stained osteoclasts on day 4. Viable osteoclasts are indicated with an arrow and apoptotic osteoclasts are indicated with a star. These data are representative of three separate experiments. D. Mean +/− SD of the number of apoptotic osteoclasts on day 3 after feeding the cells with MCSF and RANKL for the indicated time in hours. Note that apoptosis is not observed until day 4 (24 h after feeding on day 3). E. The ratio of apoptotic osteoclasts to total number of osteoclasts is also presented as mean +/− SD. These data were obtained from four replicate wells. (p<0.05) and are representative of three separate experiments. F. Mean +/− SD of proliferation of WT and TIEG1−/− osteoclast progenitors. WT and TIEG1−/− non-adherent bone marrow cells were seeded at 1×105 cells/well in 96-well plates and grown at 37°C for 3 h at day 0, 1, 2 and 3 of differentiation. Proliferation was determined using an absorbance of 490 nm and the CellTiter 96® AQueous One Solution Assay kit. These data were obtained from eight replicate wells (p<0.05) and are representative of three separate experiments. G. The mean +/− SD of number of colony forming units (CFU)-GM. These data are from three replicate wells (p<0.05) and are representative of three separate experiments.

Figure 1

doi: https://doi.org/10.1371/journal.pone.0017522.g001