Human mesenchymal stem cells promote human osteoclast differentiation fromCD34(+) bone marrow hematopoietic progenitors

Interactions between osteoclast progenitors and stromal cells derived from mesenchymal stem cells (MSCs) within the bone marrow are important for oste oclast differentiation. In vitro models of osteoclastogenesis are well esta blished in animal species; however, such assays do not necessarily reflect human osteoclastogenesis. We sought to establish a reproducible coculture m odel of human osteoclastogenesis using highly purified human marrow-derived MSCs (hMSCs) and CD34(+) hematopoietic stem cells (HSCs). After 3 weeks, c oculture of hMSCs and HSCs resulted in an increase in hematopoietic cell nu mber with formation of multinucleated osteoclast-like cells Ocls). Cocultur e of hMSCs with HSCs, transduced with a retroviral vector that expresses en hanced green fluorescent protein, produced enhanced green fluorescent prote in(+) Ocls, further demonstrating that Ocls arise from HSCs. These Ocls exp ress calcitonin and vitronectin receptors and tartrate-resistant acid phosp hatase and possess the ability to resorb bone. Ocl formation in this assay is cell contact dependent and is independent of added exogenous factors. Co nditioned medium from the coculture contained high levels of interleukin (I L)-6, IL-11, leukemia inhibitory factor (LIF), and macrophage-colony stimul ating factor. IL-6 and LIF were present at low levels in cultures of hMSCs but undetectable in cultures of HSCs alone. These data suggest that cocultu re with HSCs induce hMSCs to secrete cytokines involved in Ocl formation. A ddition of neutralizing anti-IL6, IL-11, LIF, or macrophage-colony stimulat ing factor antibodies to the coculture inhibited Ocl formation. hMSCs seem to support Ocl formation as undifferentiated progenitor cells, because trea tment of hMSCs with dexamethasone, ascorbic acid, and beta-glycerophosphate (to induce osteogenic differentiation) actually inhibited osteoclastogenes is in this coculture model. In conclusion, we have developed a simple and r eproducible assay using culture-expanded hMSCs and purified HSCs with which to study the mechanisms of human osteoclastogenesis.