Osteoclasts are derived from myeloid lineage cells, and their differentiation is supported by various osteotropic factors, including the tumor necrosis factor (TNF) family member TNF-related activation-induced cytokine (TRANCE). Genetic deletion of TRANCE or its receptor, receptor activator of nuclear factor κB (RANK), results in severely osteopetrotic mice with no osteoclasts in their bones. TNF receptor-associated factor (TRAF) 6 is a key signaling adaptor for RANK, and its deficiency leads to similar osteopetrosis. Hence, the current paradigm holds that TRANCE–RANK interaction and subsequent signaling via TRAF6 are essential for the generation of functional osteoclasts. Surprisingly, we show that hematopoietic precursors from TRANCE-, RANK-, or TRAF6-null mice can become osteoclasts in vitro when they are stimulated with TNF-α in the presence of cofactors such as TGF-β. We provide direct evidence against the current paradigm that the TRANCE–RANK–TRAF6 pathway is essential for osteoclast differentiation and suggest the potential existence of alternative routes for osteoclast differentiation.

Osteoclasts are specialized bone resorbing cells that derive from monocyte precursors. We have identified three populations of cells with high osteoclastogenic potential in murine bone marrow, which expressed the phenotype: B220−CD3−CD11b−/low CD115+ and either CD117hi, CD117intermediate or CD117low. We have evaluated these populations for their ability to also generate macrophages and dendritic cells. At a single cell level, the population expressing higher CD117 levels was able to generate bone-resorbing osteoclasts, phagocytic macrophages and antigen-presenting dendritic cells in vitro with efficiencies of over 90 percent, indicating that there exists a common developmental pathway for these cell types. Cells with osteoclastogenic potential also exist in blood and peripheral hematopoietic organs. Their functional meaning and/or their relationship with bone marrow progenitors is not well established. Hence, we characterized murine peripheral cell populations for their ability to form osteoclasts, macrophages and dendritic cells in vitro. The spleen and peripheral blood monocyte progenitors share phenotypic markers with bone marrow progenitors, but differ in their expression of CD11b, which was low in bone marrow but high in periphery. We propose that circulating monocyte progenitors are derived from a common bone marrow osteoclasts/macrophage/dendritic cell progenitor (OcMDC), which we have now characterized at a clonal level. However, the lineage relationship between the bone marrow and peripheral monocyte progenitors has yet to be defined.

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We studied the effects of PTH on the expression of tumor necrosis factor-related activation-induced cytokine (TRANCE), osteoprotegerin (OPG), and receptor activator of NF kappaB (RANK) messenger RNA (mRNA) in cultured murine bone marrow, calvaria, and osteoblasts. TRANCE, OPG, and RANK are recently identified regulators of osteoclast formation. Bone marrow cells were cultured with or without PTH(1-34) for 6 days. TRANCE, OPG, and RANK mRNA were measured by RT-PCR. In 6-day cultures, PTH stimulated the number of OCL/well in a dose-dependent manner. A time course showed significant (P < 0.01) increases in OCL/well after 24 h of PTH (100 ng/ml). TRANCE mRNA expression, like OCL formation, increased dose dependently and was maximal, with 10-100 ng/ml PTH. In contrast, OPG mRNA expression was decreased by 0.1 ng/ml PTH (40%) and completely abolished by 1 ng/ml. TRANCE mRNA expression was rapidly stimulated by PTH (maximal response at 1 h, 8.1-fold over control). Expression declined by 40% at 24 h but was still much greater than control at 6 days (4.6-fold) in a time-course study. PTH caused a transient stimulation of OPG mRNA at 1 h (2-fold), which returned to basal levels by 2 h. After 6 h, PTH completely inhibited OPG mRNA. There were only minor effects of PTH on RANK mRNA expression. PTH had less potent effects on TRANCE and OPG mRNA expression in calvaria organ cultures and osteoblasts. In mouse calvaria cultures, TRANCE expression was detectable in controls and was increased 2.9-fold by PTH at 24 h. PTH treatment of calvaria decreased OPG expression by 30% at 6 h. MC3T3 E-1 osteoblastic cells expressed minimal levels of TRANCE mRNA either before or after PTH treatment. OPG mRNA was present in MC3T3 E-1 cells, but levels were not modulated by PTH. In primary osteoblastic cells, PTH stimulated TRANCE mRNA expression 4-fold at 2 h and inhibited OPG mRNA expression by 46%. These results demonstrate a tight correlation between the ability of PTH to stimulate OCL formation in marrow culture and expression of TRANCE (r = 0.87, P < or = 0.05) and OPG mRNA (r = -0.88, P < or = 0.05). Reciprocal regulation of TRANCE and OPG mRNA by PTH preceded its effects on OCL formation by 18-23 h. Hence, it is likely that PTH regulates bone resorption, at least in part, via its effects on TRANCE and OPG expression.

Background: miR-29 is a positive regulator of osteoblastogenesis, but its role in osteoclastogenesis is undefined. Results: Expression of all miR-29 family members increased during osteoclastic differentiation. miR-29 knockdown impaired migration, osteoclast commitment, and formation. Six novel miR-29 targets were identified. Conclusion: miR-29 promotes osteoclastogenesis. Significance: These data expand our understanding of osteoclastogenesis, providing insight into miR-29 function in hematopoietic cells and other lineages.

We examined OVX-induced bone loss in three TLD mouse models. In TLD mice, OVX caused trabecular bone loss equivalent to that of WT. In contrast, cortical bone loss with OVX was variable. We conclude that T lymphocytes do not influence OVX-induced trabecular bone loss. Introduction:We examined ovariectomy (OVX)-induced bone loss in three T lymphocyte-deficient (TLD) mouse models: nude mice, recombination activating gene 2-deficient (RAG2 KO) mice, and ⌻ cell receptor ␣ chain-deficient (TCR␣ KO) mice. Materials and Methods: Bone mass was examined by DXA, CT, and histomorphometry. We also examined the effect of OVX on T lymphocytes in the bone marrow and spleens of wildtype (WT) mice and on in vitro osteoclastogenesis and colony forming unit-granulocyte macrophage (CFU-GM) activity in the bone marrow of WT and nude mice. Results: In WT mice, OVX did not alter T lymphocyte number in the bone marrow but did increase T lymphocytes in the spleen. Comparison of bone mass in nude, RAG2 KO, and TCR␣ KO mice with WT as measured by DXA showed decreased femoral bone mass in nude mice and increased vertebral bone mass in RAG2 KO mice. In TCR␣ KO mice, femoral, tibial, and vertebral bone mass were decreased. In vertebrae and long bones, bone loss with OVX was consistently present in WT mice but variably present in TLD mice as measured by DXA. In contrast, CT and histomorphometry showed similar trabecular bone loss after OVX in all mice. However, femoral cortical bone loss occurred only in WT and RAG2 KO mice. OVX produced similar trabecular bone loss in WT and TCR␣ ⌲⌷ mice and also induced cortical bone loss in both. Histomorphometry showed that TRACP + area in bones was increased by OVX in femurs from both WT and nude mice as was in vitro osteoclast-like cell formation and CFU-GM activity. Conclusions: These results show that OVX caused similar trabecular bone loss in both WT and TLD mice. The ability of DXA and measurement of cortical bone loss to show OVX-induced effects on bone mass was variable. It seems that T lymphocytes are not critical for OVX-induced trabecular bone loss in these mouse models.

Osteoporosis is an age-related disorder that affects both women and men, although estrogen deficiency induced by menopause accelerates bone loss in older women. As the demographic shifts to a more aged population, a growing number of men and women will be afflicted with osteoporosis. Since the current drug therapies available have multiple side effects, including increased risk of developing certain types of cancer or complications, a search for potential nonpharmacologic alternative therapies for osteoporosis is of prime interest. Soy isoflavones (SI) have demonstrated potential bone-specific effects in a number of studies. This article provides a systematic review of studies on osteoporotic bone loss in relation to SI intake from diet or supplements to comprehensively explain how SI affect the modulation of bone remodeling. Evidence from epidemiologic studies supports that dietary SI attenuate menopause-induced osteoporotic bone loss by decreasing bone resorption and stimulating bone formation. Other studies have also illustrated that bone site-specific trophic and synergistic effects combined with exercise intervention might contribute to improve the bioavailability of SI or strengthen the bonespecific effects. To date, however, the effects of dietary SI on osteoporotic bone loss remain inconclusive, and study results vary from study to study. The current review will discuss the potential factors that result in the conflicting outcomes of these studies, including dosages, intervention materials, study duration, race, and genetic differences. Further well-designed studies are needed to fully understand the underlying mechanism and evaluate the effects of SI on osteoporosis in humans.

To investigate the role of T lymphocytes in osteoclastogenesis, we performed in vivo depletion of CD4 and/or CD8 T lymphocyte subsets and evaluated in vitro osteoclast-like cell (OCL) formation. T lymphocyte depletion (TLD) with mAbs was confirmed 24 h later by flow cytometry. OCL formation was stimulated with 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3) in bone marrow and with recombinant mouse (rm) receptor activator of NF-κB ligand (RANK-L) and rmM-CSF in bone marrow and spleen cell cultures. OCL formation was up to 2-fold greater in 1,25-(OH)2D3-stimulated bone marrow cultures from TLD mice than in those from intact mice. In contrast, TLD did not alter OCL formation in bone marrow or spleen cell cultures that were stimulated with rmRANK-L and rmM-CSF. The effects of TLD seemed to be mediated by enhanced PG synthesis, because the PGE2 concentration in the medium of 1,25-(OH)2D3-stimulated bone marrow cultures from TLD mice was 5-fold higher than that in cultures from intact mice, and indomethacin treatment abolished the stimulatory effect of TLD on OCL formation. There was a 2-fold increase in RANK-L expression and an almost complete suppression of osteoprotegerin expression in 1,25-(OH)2D3-stimulated bone marrow cultures from TLD mice compared with those from intact mice. Although there was a small (20%) increase in IL-1α expression in 1,25-(OH)2D3-stimulated bone marrow cultures from TLD mice, TLD in mice lacking type I IL-1R and wild-type mice produced similar effects on OCL formation. Our data demonstrate that TLD up-regulates OCL formation in vitro by increasing PG production, which, in turn, produces reciprocal changes in RANK-L and osteoprotegerin expression. These results suggest that T lymphocytes influence osteoclastogenesis by altering bone marrow stromal cell function.