IntroductionAn increasing number of studies underline the interactions between the bone and immune systems and have led to the emergence of osteoimmunology. 1,2 Excessive bone resorption is frequently associated with chronic infections and autoimmune and inflammatory diseases. [3][4][5][6] The immune system plays a major role in this process, in particular through activated T cells, which secrete proinflammatory cytokines involved in osteoclastogenesis. 7 However, less is known about the involvement of other immune cells in the control of bone resorption. Dendritic cells (DCs) also play an important role in autoimmune and inflammatory diseases. 8 These cells derive from the same myeloid precursor as osteoclasts (OCLs), and both cell types are modulated by common factors, mainly by receptor activator of NF-B ligand (RANK-L). RANK-L is essential for the differentiation of OCLs, 9 the activity and survival of DCs. 10 These data highlight a potential link between DCs and OCLs.Cells from the myelomonocytic lineage, including DCs, display a high developmental and functional plasticity depending on local factors and stimuli experienced during their differentiation and maturation. 11,12 Although they were considered to be terminally differentiated cells, recent studies have suggested that mature splenic DCs can be influenced by their microenvironment to undergo further differentiation. Splenic stromal cells induce mature DCs to differentiate into regulatory DCs, which differ from mature DCs by their phenotype, their cytokine secretion pattern, and their ability to inhibit T-cell proliferation. 13 Moreover, DCs generated in vitro transdifferentiate into endothelial cells when cultured with tumor-conditioned media 14 or into OCLs when cultured with osteoclastogenic factors. 15,16 Although these in vitro studies revealed the capacity of DCs to transdifferentiate into other cell types under specific conditions, it is not clear yet whether this plasticity takes place in vivo.Osteopetrosis is characterized by an impaired bone resorption because of the absence of OCL formation or activity. 17 In the osteopetrotic oc/oc mouse, differentiated OCLs are present but are unable to resorb bone because of a deletion in the Tcirg1 gene encoding the a3 subunit of the vacuolar ATPase. 18 The a3 protein is responsible for the acidification process necessary for the dissolution of the bone matrix leading to the formation of resorption lacunae. In the absence of a3 expression, the bone marrow of oc/oc mice is filled with numerous and disorganized trabeculae, and osteoclastogenesis is highly increased. 19,20 The consequence of this severe osteopetrotic phenotype is a life span less than 3 weeks. Therefore, the oc/oc mouse provides an appropriate model to assess the in vivo capacity of wild-type precursor cells to give rise to functional OCLs.To assess whether DCs have an osteoclastogenic potential, we purified them from normal mice and cultured them with RANK-L and macrophage-colony stimulating factor (M-CSF). We showed that this treatment a...

Bone marrow mesenchymal stromal cells (BM-MSCs) have a critical role in tissue regeneration and in the hematopoietic niche due to their differentiation and self-renewal capacities. These mechanisms are finely tuned partly by small non-coding microRNA implicated in post-transcriptional regulation. The easiest way to quantify them is RT-qPCR followed by normalization on validated reference genes (RGs). This study identified appropriate RG for normalization of miRNA expression in BM-MSCs and HS27a and HS5 cell lines in various conditions including normoxia, hypoxia, co-culture, as model for the hematopoietic niche and after induced differentiation as model for regenerative medicine. Six candidates, namely miR-16-5p, miR-34b-3p, miR-103a-3p, miR-191-5p, let-7a-5p and RNU6A were selected and their expression verified by RT-qPCR. Next, a ranking on stability of the RG candidates were performed with two algorithms geNorm and RefFinder and the optimal number of RGs needed to normalize was determined. Our results indicate miR-191-5p as the most stable miRNA in all conditions but also that RNU6a, usually used as RG is the less stable gene. This study demonstrates the interest of rigorously evaluating candidate miRNAs as reference genes and the importance of the normalization process to study the expression of miRNAs in BM-MSCs or derived cell lines.

Introduction Inflammatory joint diseases such as rheumatoid arthritis are associated with local bone erosions and systemic bone loss, mediated by increased osteoclastic activity. The receptor activator of nuclear factor (NF) κB ligand (RANKL) plays a key role in mediating inflammation-induced bone loss, whereas tumour necrosis factor (TNF) plays a central role in the inflammatory process. Here we tested whether a recently identified class of small molecule inhibitors of RANKL signalling (ABD compounds) also affect TNF signalling and whether these compounds inhibit inflammation in an animal model of rheumatoid arthritis. Methods The inhibitory effects of the ABD compounds on TNF-induced signalling were tested in mouse macrophage cultures by western blotting and in an NFκB luciferase-reporter cell line. The anti-inflammatory effects of the compounds were tested in the mouse collagen-induced arthritis model of rheumatoid arthritis. Results The ABD compounds ABD328 and ABD345 both inhibited TNF-induced activation of the NFκB pathway and the extracellular signal-regulated kinase (ERK) and Jun kinase (JNK) mitogen activated protein kinases (MAPKs). When tested in the mouse collageninduced arthritis model of rheumatoid arthritis, the compounds suppressed inflammatory arthritis, inhibited joint destruction and prevented systemic bone loss. Furthermore, one of the compounds (ABD328) showed oral activity. Conclusions Here we describe a novel class of small molecule compounds that inhibit both RANKL-and TNFinduced NFκB and MAPK signalling in osteoclasts and macrophages, and inflammation and bone destruction in a mouse model of rheumatoid arthritis. These novel compounds therefore represent a promising new class of treatments for inflammatory diseases, such as rheumatoid arthritis.