Intermittent parathyroid hormone (iPTH) treatment stimulates Tcell production of the osteogenic Wnt ligand Wnt10b, a factor required for iPTH to activate Wnt signaling in osteoblasts and stimulate bone formation. However, it is unknown whether iPTH induces Wnt10b production and bone anabolism through direct activation of the parathyroid hormone (PTH)/PTH-related protein receptor (PPR) in T cells. Here, we show that conditional silencing of PPR in T cells blunts the capacity of iPTH to induce T-cell production of Wnt10b; activate Wnt signaling in osteoblasts; expand the osteoblastic pool; and increase bone turnover, bone mineral density, and trabecular bone volume. These findings demonstrate that direct PPR signaling in T cells plays an important role in PTH-induced bone anabolism by promoting T-cell production of Wnt10b and suggest that T cells may provide pharmacological targets for bone anabolism.bone mass | T lymphocytes | bone cells P arathyroid hormone (PTH) is a major regulator of calcium metabolism and defends against hypocalcemia, in part, by stimulating bone resorption, and thereby the release of calcium from the skeleton. However, when injected daily, a regimen known as intermittent parathyroid hormone (iPTH) treatment, the hormone markedly stimulates trabecular and cortical bone formation. Although this bone-forming activity is antagonized by a stimulation of bone resorption, the net effect of iPTH treatment is an improvement in bone microarchitecture and increased strength (1, 2). As a result, intermittent treatment with the 1-34 fragment of PTH is a Food and Drug Administration-approved treatment modality for postmenopausal osteoporosis (3).The effects of PTH on bone result from its binding to the PTH/PTH-related protein receptor (PPR or PTHR1) expressed on bone marrow (BM), stromal cells (SCs), osteoblasts (OBs), and osteocytes (1, 4, 5). iPTH stimulates bone formation by increasing the number of OBs (6-8), a phenomenon achieved through activation of quiescent lining cells (9), increased OB proliferation (10, 11) and differentiation (10, 12, 13), attenuation of OB apoptosis (14-17), and signaling in osteocytes (18). However, the specific contribution of each of these effects of iPTH remains controversial. The expansion of the osteoblastic pool induced by iPTH is initiated by the release from the matrix undergoing resorption of TGF-β, insulin-like growth factor 1, and other growth factors that recruit SCs to remodeling areas (19)(20)(21)(22). Subsequent events are driven primarily by the activation of Wnt signaling in osteoblastic cells (23). Activation of Wnt signaling induces OB proliferation (24) and differentiation (23, 25), prevents OB apoptosis (16,17,26), and augments OB production of osteoprotegerin (OPG) (27).iPTH activates Wnt signaling in OBs through multiple mechanisms that include Wnt ligand-independent activation of the Wnt coreceptor LRP6 (28), increased production of Wnt ligands by bone and BM cells (29,30), and suppression of sclerostin production (31-33). Additional effects on the ...

We identified previously in vitro LRP4 (low-density lipoprotein receptor-related protein 4) as a facilitator of the WNT (Winglesstype) antagonist sclerostin and found mutations disrupting this function to be associated with high bone mass in humans similar to patients lacking sclerostin. To further delineate the role of LRP4 in bone in vivo, we generated mice lacking Lrp4 in osteoblasts/osteocytes or osteocytes only. Lrp4 deficiency promoted progressive cancellous and cortical bone gain in both mutants, although more pronouncedly in mice deficient in osteoblast/osteocyte Lrp4, consistent with our observation in human bone that LRP4 is most strongly expressed by osteoblasts and early osteocytes. Bone gain was related primarily to increased bone formation. Interestingly, Lrp4 deficiency in bone dramatically elevated serum sclerostin levels whereas bone expression of Sost encoding for sclerostin was unaltered, indicating that osteoblastic Lrp4 retains sclerostin within bone. Moreover, we generated anti-LRP4 antibodies selectively blocking sclerostin facilitator function while leaving unperturbed LRP4-agrin interaction, which is essential for neuromuscular junction function. These antibodies increased bone formation and thus cancellous and cortical bone mass in skeletally mature rodents. Together, we demonstrate a pivotal role of LRP4 in bone homeostasis by retaining and facilitating sclerostin action locally and provide a novel avenue to bone anabolic therapy by antagonizing LRP4 sclerostin facilitator function.LRP4 | SOST | sclerostin | WNT | bone anabolism

The bone formation inhibitor sclerostin encoded by SOST binds in vitro to low-density lipoprotein receptor-related protein (LRP) 5/6 Wnt co-receptors, thereby inhibiting Wnt/b-catenin signaling, a central pathway of skeletal homeostasis. Lrp5/LRP5 deficiency results in osteoporosis-pseudoglioma (OPPG), whereas Sost/SOST deficiency induces lifelong bone gain in mice and humans. Here, we analyzed the bone phenotype of mice lacking Sost (Sost À/À ), Lrp5 (Lrp5 À/À ), or both (Sost À/À ;Lrp5 À/À ) to elucidate the mechanism of action of Sost in vivo. Sost deficiency-induced bone gain was significantly blunted in Sost À/À ;Lrp5 À/À mice. Yet the Lrp5 OPPG phenotype was fully rescued in Sost À/À ;Lrp5 À/À mice and most bone parameters were elevated relative to wild-type. To test whether the remaining bone increases in Sost À/À ;Lrp5 À/À animals depend on Lrp6, we treated wild-type, Sost À/À , and Sost À/À ;Lrp5 À/À mice with distinct Lrp6 function blocking antibodies. Selective blockage of Wnt1 class-mediated Lrp6 signaling reduced cancellous bone mass and density in wild-type mice. Surprisingly, it reversed the abnormal bone gain in Sost À/À and Sost À/À ;Lrp5 À/À mice to wild-type levels irrespective of enhancement or blockage of Wnt3a class-mediated Lrp6 activity. Thus, whereas Sost deficiency-induced bone anabolism partially requires Lrp5, it fully depends on Wnt1 class-induced Lrp6 activity. These findings indicate: first, that OPPG syndrome patients suffering from LRP5 loss-of-function should benefit from principles antagonizing SOST/sclerostin action; and second, that therapeutic WNT signaling inhibitors may stop the debilitating bone overgrowth in sclerosing disorders related to SOST deficiency, such as sclerosteosis, van Buchem disease, and autosomal dominant craniodiaphyseal dysplasia, which are rare disorders without viable treatment options.

Better understanding of bone growth and regeneration mechanisms within periosteal tissues will improve understanding of bone physiology and pathology. Macrophage contributions to bone biology and repair have been established but specific investigation of periosteal macrophages has not been undertaken. We used an immunohistochemistry approach to characterize macrophages in growing murine bone and within activated periosteum induced in a mouse model of bone injury. Osteal tissue macrophages (osteomacs) and resident macrophages were distributed throughout resting periosteum. In tissues collected from 4-week-old mice, osteomacs were observed intimately associated with sites of periosteal diaphyseal and metaphyseal bone dynamics associated with normal growth. This included F4/80 + Mac-2 − /low osteomac association with extended tracks of bone formation (modeling) on diphyseal periosteal surfaces. Although this recapitulated endosteal osteomac characteristics, there was subtle variance in the morphology and spatial organization of periosteal modeling-associated osteomacs, which likely reflects the greater structural complexity of periosteum. Osteomacs, resident macrophages and inflammatory macrophages (F4/80 + Mac-2 hi ) were associated with the complex bone dynamics occurring within the periosteum at the metaphyseal corticalization zone. These three macrophage subsets were also present within activated native periosteum after bone injury across a 9-day time course that spanned the inflammatory through remodeling bone healing phases. This included osteomac association with foci of endochondral ossification within the activated native periosteum. These observations confirm that osteomacs are key components of both osteal tissues, in spite of salient differences between endosteal and periosteal structure and that multiple macrophage subsets are involved in periosteal bone dynamics.

No abstract

No abstract