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CHAPTER 6 The involvement of M-CSF and RANKL in mitochondrial biogenesis during osteoclast formation and macrophage differentiation

Bone resorption is characterized by a high release of protons, a process associated with a serious energy demand and thus many mitochondria. Although the high number of these organelles is a well-known characteristic of osteoclasts, surprisingly little is known about their biogenesis during osteoclast differentiation. In this study we investigated the role of macrophage colony-stimulating factor (M-CSF) and receptor activator of nuclear factor kappa B ligand (RANKL), important cytokines during osteoclastogenesis, in mitochondrial biogenesis and activity. Murine bone marrow cells were cultured for up to 7 days with or without M-CSF and/or RANKL. We analyzed the mRNA expression of mitochondria-related genes, quantified the mitochondrial activity per cell and analyzed morphological differences between mitochondria in the different cultures. We could distinguish macrophage differentiation and osteoclast differentiation in the M-CSF and M-CSF/RANKL cultures and found a differential up-regulation of mitochondria-related genes during these processes. In particular, D-3 hydroxybutyrate dehydrogenase (BDH) and peroxisome proliferator-activator receptor-γ co-activator-1β (PGC-1β) were more than 10-fold induced during osteoclastogenesis compared to the macrophage culture. Differences between mitochondria in those two cell types also existed at the morphological level. The mitochondria of the osteoclasts were larger and more mitochondria were formed per cytoplasmic surface area compared to the other cultures. On the other hand, it was M-CSF that strongly increased the percentage of cells with a high number of active mitochondria. Taken together, our data suggest a differential biogenesis of mitochondria during macrophage and osteoclast differentiation wherein M-CSF and RANKL play different roles.