Microfibrillar-associated protein 4 as a potential marker of acute relapse in inflammatory demyelinating diseases of the central nervous system: Pathological and clinical aspects.

BACKGROUND: Microfibrillar-associated protein 4 (MFAP4) is an extracellular matrix protein not previously described in the human central nervous system (CNS). OBJECTIVES: We determined MFAP4 CNS expression and measured cerebrospinal fluid (CSF) and serum levels. METHODS: Tissue was sampled at autopsy from patients with acute multiple sclerosis (MS) (n = 3), progressive MS (n = 3), neuromyelitis optica spectrum disorder (NMOSD) (n = 2), and controls (n = 9), including 6 healthy controls (HC). MFAP4 levels were measured in 152 patients: 49 MS, 62 NMOSD, 22 myelin oligodendrocyte glycoprotein-associated disease (MOGAD), and 19 isolated optic neuritis (ION). RESULTS: MFAP4 localized to meninges and vascular/perivascular spaces, intense in the optic nerve. At sites of active inflammation, MFAP4 reactivity was reduced in NMOSD and acute MS and less in progressive MS. CSF MFAP4 levels were reduced during relapse and at the onset of diseases (mean U/mL: MS 14.3, MOGAD 9.7, and ION 14.6 relative to HC 17.9. (p = 0.013, p = 0.000, and p = 0.019, respectively). Patients with acute ON (n = 68) had reduced CSF MFAP4 (mean U/mL: 14.5, p = 0.006). CSF MFAP4 levels correlated negatively with relapse severity (rho = -0.41, p = 0.017). CONCLUSION: MFAP4 immunoreactivity was reduced at sites of active inflammation. CSF levels of MFAP4 were reduced following relapse and may reflect disease activity.

Catabolic activity of osteoblast lineage cells contributes to osteoclastic bone resorption in vitro.

Osteoblast lineage cells in human bone were recently shown to colonize eroded bone surfaces and to closely interact with osteoclasts. They proved to be identical to reversal cells and are believed to differentiate into bone-forming osteoblasts thereby coupling resorption and formation. However, they also exert catabolic activity that contributes to osteoclastic bone resorption, but this has not received much attention. Herein, we used co-cultures of primary human osteoblast lineage cells and human osteoclasts derived from peripheral blood monocytes to investigate whether a catabolic activity of osteoblast lineage cells could impact on osteoclastic bone resorption. Through a combination of immunofluorescence, in situ hybridization and time-lapse experiments, we show that MMP-13-expressing osteoblast lineage cells are attracted to and closely interact with bone-resorbing osteoclasts. This close interaction results in a strong and significant increase in the bone resorptive activity of osteoclasts - especially those making trenches. Importantly, we show that osteoclastic bone resorption becomes sensitive to inhibition of matrix metalloproteinases in the presence, but not in the absence, of osteoblast lineage cells. We propose that this may be due to the direct action of osteoblast-lineage-derived MMP-13 on bone resorption.

Coordination of Fusion and Trafficking of Pre-osteoclasts at the Marrow-Bone Interface.

Fusion is the final osteoclast differentiation step leading to bone resorption. In healthy trabecular bone, osteoclast fusion is restricted to bone surfaces undergoing resorption, and necessarily requires site-specific recruitment of mononucleated pre-osteoclasts originating from the bone marrow. However, the spatiotemporal mechanism coordinating recruitment and fusion is poorly investigated. Herein we identify a collagen/vascular network as a likely structure supporting this mechanism. We therefore used multiplex immunohistochemistry and electron microscopy on human iliac crest bone samples, in combination with functional assays performed in vitro with osteoclasts generated from healthy blood donors. First, we found that putative pre-osteoclasts are in close vicinity of a network of collagen fibers associated with vessels and bone remodeling compartment canopies. Based on 3D-reconstructions of serial sections, we propose that this network may serve as roads leading pre-osteoclasts to resorption sites, as reported for cell migration in other tissues. Importantly, almost all these bone marrow pre-osteoclasts, but only some osteoclasts, express the collagen receptor OSCAR, which is reported to induce fusion competence. Furthermore, differentiating osteoclasts cultured on collagen compared to mineral show higher fusion rates, higher expression of fusogenic cytokines, and a CD47 plasma membrane distribution pattern reported to be typical of a pre-fusion state-thus collectively supporting collagen-induced fusion competence. Finally, these in vitro assays show that collagen induces high cell mobility. The present data lead to a model where collagen fibers/vasculature support the coordination between traffic and fusion of pre-osteoclasts, by serving as a physical road and inducing fusion competence as well as cell mobility.

Early reversal cells in adult human bone remodeling: osteoblastic nature, catabolic functions and interactions with osteoclasts.

The mechanism coupling bone resorption and formation is a burning question that remains incompletely answered through the current investigations on osteoclasts and osteoblasts. An attractive hypothesis is that the reversal cells are likely mediators of this coupling. Their nature is a big matter of debate. The present study performed on human cancellous bone is the first one combining in situ hybridization and immunohistochemistry to demonstrate their osteoblastic nature. It shows that the Runx2 and CD56 immunoreactive reversal cells appear to take up TRAcP released by neighboring osteoclasts. Earlier preclinical studies indicate that reversal cells degrade the organic matrix left behind by the osteoclasts and that this degradation is crucial for the initiation of the subsequent bone formation. To our knowledge, this study is the first addressing these catabolic activities in adult human bone through electron microscopy and analysis of molecular markers. Periosteoclastic reversal cells show direct contacts with the osteoclasts and with the demineralized resorption debris. These early reversal cells show (1) ¾-collagen fragments typically generated by extracellular collagenases of the MMP family, (2) MMP-13 (collagenase-3) and (3) the endocytic collagen receptor uPARAP/Endo180. The prevalence of these markers was lower in the later reversal cells, which are located near the osteoid surfaces and morphologically resemble mature bone-forming osteoblasts. In conclusion, this study demonstrates that reversal cells colonizing bone surfaces right after resorption are osteoblast-lineage cells, and extends to adult human bone remodeling their role in rendering eroded surfaces osteogenic.

High-dose therapy improves the bone remodelling compartment canopy coverage and bone formation in multiple myeloma.

Bone loss in multiple myeloma (MM) is caused by an uncoupling of bone formation to resorption trigged by malignant plasma cells. Increasing evidence indicates that the bone remodelling compartment (BRC) canopy, which normally covers the remodelling sites, is important for coupled bone remodelling. Loss of this canopy has been associated with bone loss. This study addresses whether the bone remodelling in MM is improved by high-dose therapy. Bone marrow biopsies obtained from 20 MM patients, before and after first-line treatment with high-dose melphalan followed by autologous stem cell transplantation, and from 20 control patients with monoclonal gammopathy of undetermined significance were histomorphometrically investigated. This investigation confirmed that MM patients exhibited uncoupled bone formation to resorption and reduced canopy coverage. More importantly, this study revealed that a good response to anti-myeloma treatment increased the extent of formative bone surfaces with canopy, and reduced the extent of eroded surfaces without canopy, reverting the uncoupled bone remodelling, while improving canopy coverage. The association between improved coupling and the canopy coverage supports the notion that canopies are critical for the coupling of bone formation to resorption. Furthermore, this study supports the observation that systemic bone disease in MM can be reversed in MM patients responding to anti-myeloma treatment.

A supra-cellular model for coupling of bone resorption to formation during remodeling: lessons from two bone resorption inhibitors affecting bone formation differently.

The bone matrix is maintained functional through the combined action of bone resorbing osteoclasts and bone forming osteoblasts, in so-called bone remodeling units. The coupling of these two activities is critical for securing bone replenishment and involves osteogenic factors released by the osteoclasts. However, the osteoclasts are separated from the mature bone forming osteoblasts in time and space. Therefore the target cell of these osteoclastic factors has remained unknown. Recent explorations of the physical microenvironment of osteoclasts revealed a cell layer lining the bone marrow and forming a canopy over the whole remodeling surface, spanning from the osteoclasts to the bone forming osteoblasts. Several observations show that these canopy cells are a source of osteoblast progenitors, and we hypothesized therefore that they are the likely cells targeted by the osteogenic factors of the osteoclasts. Here we provide evidence supporting this hypothesis, by comparing the osteoclast-canopy interface in response to two types of bone resorption inhibitors in rabbit lumbar vertebrae. The bisphosphonate alendronate, an inhibitor leading to low bone formation levels, reduces the extent of canopy coverage above osteoclasts. This effect is in accordance with its toxic action on periosteoclastic cells. In contrast, odanacatib, an inhibitor preserving bone formation, increases the extent of the osteoclast-canopy interface. Interestingly, these distinct effects correlate with how fast bone formation follows resorption during these respective treatments. Furthermore, canopy cells exhibit uPARAP/Endo180, a receptor able to bind the collagen made available by osteoclasts, and reported to mediate osteoblast recruitment. Overall these observations support a mechanism where the recruitment of bone forming osteoblasts from the canopy is induced by osteoclastic factors, thereby favoring initiation of bone formation. They lead to a model where the osteoclast-canopy interface is the physical site where coupling of bone resorption to bone formation occurs.

The bone resorption inhibitors odanacatib and alendronate affect post-osteoclastic events differently in ovariectomized rabbits.

Odanacatib (ODN) is a bone resorption inhibitor which differs from standard antiresorptives by its ability to reduce bone resorption without decreasing bone formation. What is the reason for this difference? In contrast with other antiresorptives, such as alendronate (ALN), ODN targets only the very last step of the resorption process. We hypothesize that ODN may therefore modify the remodeling events immediately following osteoclastic resorption. These events belong to the reversal phase and include recruitment of osteoblasts, which is critical for connecting bone resorption to formation. We performed a histomorphometric study of trabecular remodeling in vertebrae of estrogen-deficient rabbits treated or not with ODN or ALN, a model where ODN, but not ALN, was previously shown to preserve bone formation. In line with our hypothesis, we found that ODN treatment compared to ALN results in a shorter reversal phase, faster initiation of osteoid deposition on the eroded surfaces, and higher osteoblast recruitment. The latter is reflected by higher densities of mature bone forming osteoblasts and an increased subpopulation of cuboidal osteoblasts. Furthermore, we found an increase in the interface between osteoclasts and surrounding osteoblast-lineage cells. This increase is expected to favor the osteoclast-osteoblast interactions required for bone formation. Regarding bone resorption itself, we show that ODN, but not ALN, treatment results in shallower resorption lacunae, a geometry favoring bone stiffness. We conclude that, compared to standard antiresorptives, ODN shows distinctive effects on resorption geometry and on reversal phase activities which positively affect osteoblast recruitment and may therefore favor bone formation.

Case report: Death caused by multi-organ metastatic calcifications as a result of intramuscular injections with paraffin oil.

In this forensic case report, we present autopsy findings from a young male in his thirties who had been self-injecting paraffin oil into his upper extremities 8 years prior to death. The injections induced an inflammatory response, leading to granuloma formation. This, in turn, resulted in severe hypercalcemia. The external autopsy examination revealed gross macroscopic ulcerations and enlargement of upper extremities, while calcifications of ligaments, heart, kidneys and dura mater was revealed on postmortem CT-scans. Histopathological examination showed extensive multiorgan metastatic calcifications in several tissues including the lungs, heart and kidney. Cause of death was estimated to be the extensive calcific deposits in the heart likely resulting in cardiac arrest. To our knowledge this is the first case reporting findings from an autopsy in which the cause of death was linked to cosmetic oil injections.

A Critical Role of the Bone Marrow Envelope in Human Bone Remodeling.

Proper bone remodeling depends not only on a team of bone-resorbing osteoclasts and bone-forming osteoblasts. It also depends on the site-specific delivery of a large amount of osteoblast lineage cells to the bone remodeling site. How this delivery occurs is poorly known. Here, we gained insight into this mechanism by analyzing the distribution of markers of osteoblastogenesis on bone surfaces and in their bone marrow neighborhood in human cancellous bone. We found a CD271-positive/PDGFß-R-positive cell layer surrounding the bone marrow that provides osteoblastogenic potential along all bone surfaces, whether quiescent or remodeling. This bone marrow envelope cell layer takes the appearance of a canopy above remodeling sites, where it then also shows an upregulation of the proliferation marker Ki67, smooth muscle actin (SMA), tenascin C, fibronectin, and MMP13. This indicates that the canopy is a region of the bone marrow envelope where early markers of osteoblastogenesis are activated concurrently with initiation of bone remodeling. Importantly, the high proliferation index in the canopy is not associated with increasing cell densities at the canopy level, but it is at the bone surface level, thereby supporting delivery of cells from the canopy to the bone surface. This delivery route explains why lack of canopies was previously found to coincide with lack of bone formation, and fits current knowledge on the canopies as a target for regulators of bone remodeling. We conclude that the coordination of bone marrow envelope activities and bone surface activities allows integrating osteoblastogenesis and bone remodeling into the same functional unit, and propose that the bone marrow envelope is critical for preserving bone health. © 2023 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research (ASBMR).

Osteoprogenitor recruitment and differentiation during intracortical bone remodeling of adolescent humans.

BACKGROUND: Recruitment and proliferation of osteoprogenitors during the reversal-resorption phase, and their differentiation into mature bone-forming osteoblasts is crucial for initiation of bone formation during bone remodeling. This study investigates the osteoprogenitors' gradual recruitment, proliferation, and differentiation into bone-forming osteoblasts within intracortical remodeling events of healthy adolescent humans. METHODS: The study was conducted on cortical bone specimens from 11 adolescent human controls - patients undergoing surgery due to coxa valga. The osteoprogenitor recruitment route and differentiation into osteoblasts were backtracked using immunostainings and in situ hybridizations with osteoblastic markers (CD271/NGFR, osterix/SP7, COL3A1 and COL1A1). The osteoblastic cell populations were defined based on the pore surfaces, and their proliferation index (Ki67), density and number/circumference were estimated in multiplex-immunofluorescence (Ki67, TRAcP, CD34) stained sections. RESULTS: During the reversal-resorption phase, osteoclasts are intermixed with (COL3A1+NFGR+) osteoblastic reversal cells, which are considered to be osteoprogenitors of (COL1A1+SP7+) bone-forming osteoblasts. Initiation of bone formation requires a critical density of these osteoprogenitors (43 ± 9 cells/mm), which is reached though proliferation (4.4 ± 0.5 % proliferative) and even more so through recruitment of osteoprogenitors, but challenged by the ongoing expansion of the canal circumference. These osteoprogenitors most likely originate from osteoblastic bone lining cells and mainly lumen osteoprogenitors, which expand their population though proliferation (4.6 ± 0.3 %) and vascular recruitment. These lumen osteoprogenitors resemble canopy cells above trabecular remodeling sites, and like canopy cells they extend above bone-forming osteoblasts where they may rejuvenate the osteoblast population during bone formation. CONCLUSION: Initiation of bone formation during intracortical remodeling requires a critical density of osteoprogenitors on eroded surfaces, which is reached though proliferation and recruitment of local osteoprogenitors: bone lining cells and lumen osteoprogenitors.

Trabecular bone deterioration in a postmenopausal female suffering multiple spontaneous vertebral fractures due to a delayed denosumab injection - A post-treatment re-initiation bone biopsy-based case study.

Background: Denosumab, is a potent anti-resorptive that, increases bone mineral density, and reduces fracture risk in osteoporotic patients. However, several case studies have reported multiple vertebral fractures in patients discontinuing denosumab. Case presentation: This case report describes a 64-year-old female with postmenopausal osteoporosis treated with denosumab, who had her 11th injection delayed by 4 months. The patient suffered eight spontaneous vertebral fractures. After consent, an iliac crest bone biopsy was obtained following re-initiation of the denosumab treatment and analyzed by micro-computed tomography and histomorphometry. Results: micro-computed tomography analysis revealed a low trabecular bone volume of 10 %, a low trabecular thickness of 97 µm, a low trabecular spacing of 546 µm, a high trabecular number of 1.8/mm, and a high structure model index of 2.2, suggesting trabecular thinning and loss of trabecular plates. Histomorphometric trabecular bone analysis revealed an eroded perimeter per bone perimeter of 33 % and an osteoid perimeter per bone perimeter of 62 %. Importantly, 88 % of the osteoid perimeter was immediately above an eroded-scalloped cement line with no sign of mineralization, and often with no clear bone-forming osteoblasts on the surface. Moreover, only 5 % of the bone perimeter was mineralizing, reflecting that only 8 % of the osteoid perimeter underwent mineralization, resulting in a mineralization lag time of 545 days. Taken together, this indicates limited bone formation and delayed mineralization. Conclusion: We present a case report of multiple vertebral fractures after denosumab discontinuation with histomorphometric evidence that denosumab discontinuation leads to extensive trabecular bone resorption followed by a limited bone formation and delayed mineralization if the denosumab treatment is reinitiated. This highlights the importance of developing optimal discontinuation strategies for patients that are to discontinue treatment.

Local coordination between intracortical bone remodeling and vascular development in human juvenile bone.

Although failure to establish a vascular network has been associated with many skeletal disorders, little is known about what drives development of vasculature in the intracortical bone compartments. Here, we show that intracortical bone resorption events are coordinated with development of the vasculature. We investigated the prevalence of vascular structures at different remodeling stages as well as their 3D organization using proximal femoral cortical bone from 5 girls and 6 boys (aged 6-15 years). A 2D analysis revealed that non-quiescent intracortical pores contained more vascular structures than quiescent pores (p < 0.0001). Type 2 pores, i.e., remodeling of existing pores, had a higher density of vascular structures than type 1 pores, i.e., de novo created pores (p < 0.05). Furthermore, pores at the eroded-formative remodeling stage, had more vascular structures than pores at any other remodeling stage (p < 0.05). A 3D reconstruction of an intracortical remodeling event showed that osteoclasts in the advancing tip of the cutting cone as well as preosteoclasts in the lumen expressed vascular endothelial growth factor-A (VEGFA), while VEGFA-receptors 1 and 2 mainly were expressed in endothelial cells in the adjacent vasculature. Consequently, we propose that the progression of the vascular network in intracortical remodeling events is driven by osteoclasts expressing VEGFA. Moreover, the vasculature is continuously reconfigured according to the demands of the remodeling events at the surrounding bone surfaces.

Novel Developments in the Treatment of Multiple Myeloma-Associated Bone Disease.

Osteolytic bone disease is present in about 80% of patients with multiple myeloma at the time of diagnosis. Managing bone disease in patients with multiple myeloma is a challenge and requires a multi-faceted treatment approach with medication, surgery, and radiation. The established treatments with intravenous or subcutaneous antiresorptives can cause debilitating adverse events for patients, mainly osteonecrosis of the jaw, which, traditionally, has been difficult to manage. Now, oral surgery is recommended and proven successful in 60-85% of patients. Patients with spinal involvement may benefit from surgery in the form of vertebroplasty and kyphoplasty for pain relief, improved mobility, and reestablished sagittal balance, as well as the restoration of vertebral height. These procedures are considered safe, but the full therapeutic impact needs to be investigated further. Ixazomib, the first oral proteasome inhibitor, increases osteoblast differentiation, and recently published preliminary results in patients treated with Ixazomib maintenance have promisingly shown increased trabecular volume caused by prolonged bone formation activity. Other novel potential treatment strategies are discussed as well.

KIAA1199 deficiency enhances skeletal stem cell differentiation to osteoblasts and promotes bone regeneration.

Upon transplantation, skeletal stem cells (also known as bone marrow stromal or mesenchymal stem cells) can regulate bone regeneration by producing secreted factors. Here, we identify KIAA1199 as a bone marrow stromal cell-secreted factor in vitro and in vivo. KIAA1199 plasma levels of patients positively correlate with osteoporotic fracture risk and expression levels of KIAA1199 in patient bone marrow stromal cells negatively correlates with their osteogenic differentiation potential. KIAA1199-deficient bone marrow stromal cells exhibit enhanced osteoblast differentiation in vitro and ectopic bone formation in vivo. Consistently, KIAA1199 knockout mice display increased bone mass and biomechanical strength, as well as an increased bone formation rate. They also exhibit accelerated healing of surgically generated bone defects and are protected from ovariectomy-induced bone loss. Mechanistically, KIAA1199 regulates osteogenesis by inhibiting the production of osteopontin by osteoblasts, via integrin-mediated AKT and ERK-MAPK intracellular signaling. Thus, KIAA1199 is a regulator of osteoblast differentiation and bone regeneration and could be targeted for the treatment or management of low bone mass conditions.

Transitory Activation and Improved Transition from Erosion to Formation within Intracortical Bone Remodeling in Hypoparathyroid Patients Treated with rhPTH(1-84).

In hypoparathyroidism, lack of parathyroid hormone (PTH) leads to low calcium levels and decreased bone remodeling. Treatment with recombinant human PTH (rhPTH) may normalize bone turnover. This study aimed to investigate whether rhPTH(1-84) continued to activate intracortical bone remodeling after 30 months and promoted the transition from erosion to formation and whether this effect was transitory when rhPTH(1-84) was discontinued. Cortical histomorphometry was performed on 60 bone biopsies from patients (aged 31 to 78 years) with chronic hypoparathyroidism randomized to either 100 µg rhPTH(1-84) a day (n = 21) (PTH) or similar placebo (n = 21) (PLB) for 6 months as add-on to conventional therapy. This was followed by an open-label extension, where patients extended their rhPTH(1-84) (PTH) (n = 5), continued conventional treatment (CON) (n = 5), or withdrew from rhPTH(1-84) and resumed conventional therapy (PTHw) for an additional 24 months (n = 8). Bone biopsies were collected at months 6 (n = 42) and 30 (n = 18). After 6 and 30 months, the overall cortical microarchitecture (cortical porosity, thickness, pore density, and mean pore diameter) in the PTH group did not differ from that of the PLB/CON and PTHw groups. Still, the PTH group had a significantly and persistently higher percentage of pores undergoing remodeling than the PLB/CON groups. A significantly higher percentage of these pores was undergoing bone formation in the PTH compared with the PLB/CON groups, whereas the percentage of pores with erosion only was not different. This resulted in a shift in the ratio between formative and eroded pores, reflecting a faster transition from erosion to formation in the PTH-treated patients. In the rhPTH(1-84) withdrawal group PTHw, the latter effects of PTH were completely reversed in comparison to those of the PLB/CON groups. In conclusion, rhPTH(1-84) replacement therapy in hypoparathyroidism patients promotes intracortical remodeling and its transition from erosion to formation without affecting the overall cortical microstructure. The effect persists for at least 30 months and is reversible when treatment is withdrawn. © 2023 The Authors. JBMR Plus published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research.

Immunolocalization of RANK and RANKL along the root surface and in the periodontal membrane of human primary and permanent teeth.

OBJECTIVE: Root resorption, impaired tooth eruption and early tooth loss have been described in relation to diseases that involve defects in the RANK-RANKL-OPG-expression. The aim of the present immunhistochemical study was to localize and compare the reactions for RANK and membrane-bound RANKL along root surfaces and in the periodontal membrane in close proximity to the root surface of human primary and permanent teeth. MATERIALS AND METHODS: The material comprised extracted human teeth (11 primary teeth and six permanent teeth) from 10 different patients. Paraffin sections were prepared of each tooth and sections of each tooth were immunohistochemically stained with antibodies specific for membrane-bound RANKL and RANK. RESULTS: The root surface and the periodontal membrane in close proximity to the root surface did not show immunoreactivity for RANKL. RANKL was only located in odontoblasts and in cells along denticles in one primary tooth. RANK was located in mononuclear cells in the pulp and in multinucleated odontoclasts along resorbed root surfaces and along resorbed dentin surfaces in the pulp in primary teeth and one permanent tooth. CONCLUSIONS: This study demonstrated RANK positivity in resorption areas in primary and permanent teeth. RANKL was positive in the pulp of one primary tooth. RANK expression in odontoclasts and RANKL expression in the pulp may indicate that RANK/RANKL play a role during resorption.

2D size of trabecular bone structure units (BSU) correlate more strongly with 3D architectural parameters than age in human vertebrae.

Bone tissue is continuously remodeled. In trabecular bone, each remodeling transaction forms a microscopic bone structural unit (BSU), also known as a hemiosteon or a trabecular packet, which is bonded to existing tissue by osteopontin-rich cement lines. The size and shape of the BSUs are determined by the size and shape of the resorption cavity, and whether the cavity is potentially over- or under-filled by the subsequent bone formation. The present study focuses on the recently formed trabecular BSUs, and how their 2D size and shape changes with age and trabecular microstructure. The study was performed using osteopontin-immunostained frontal sections of L2 vertebrae from 8 young (aged 18.5-37.6 years) and 8 old (aged 69.1-96.4 years) control females, which underwent microcomputed tomography (µCT) imaging prior to sectioning. The contour of 4230 BSU profiles (181-385 per vertebra) within 1024 trabecular profiles were outlined, and their 2D width, length, area, and shape were assessed. Of these BSUs, 22 (0.5%) were generated by modeling-based bone formation (i.e. without prior resorption), while 99.5% were generated by remodeling-based bone formation (i.e. with prior resorption). The distributions of BSU profile width, length, and area were significantly smaller in the old versus young females (p < 0.005), and the median profile width, length, and area were negative correlated with age (p < 0.018). Importantly, these BSU profile size parameters were more strongly correlated with trabecular bone volume (BV/TV, p < 0.002) and structure model index (SMI, p < 0.008) assessed by µCT, than age. Moreover, the 2D BSU size parameters were positively correlated to the area of the individual trabecular profiles (p < 0.0001), which were significantly smaller in the old versus young females (p < 0.024). The BSU shape parameters (aspect ratio, circularity, and solidity) were not correlated with age, BV/TV, or SMI. Collectively, the study supports the notion that not only the BSU profile width, but also its length and area, are more influenced by the age-related bone loss and shift from plates to rods (SMI), than age itself. This implies that BSU profile size is mainly driven by changes in the trabecular microstructure, which affect the size of the resorption cavity that the BSU refills.

FIBCD1 is an endocytic GAG receptor associated with a novel neurodevelopmental disorder.

Whole-exome sequencing of two patients with idiopathic complex neurodevelopmental disorder (NDD) identified biallelic variants of unknown significance within FIBCD1, encoding an endocytic acetyl group-binding transmembrane receptor with no known function in the central nervous system. We found that FIBCD1 preferentially binds and endocytoses glycosaminoglycan (GAG) chondroitin sulphate-4S (CS-4S) and regulates GAG content of the brain extracellular matrix (ECM). In silico molecular simulation studies and GAG binding analyses of patient variants determined that such variants are loss-of-function by disrupting FIBCD1-CS-4S association. Gene knockdown in flies resulted in morphological disruption of the neuromuscular junction and motor-related behavioural deficits. In humans and mice, FIBCD1 is expressed in discrete brain regions, including the hippocampus. Fibcd1 KO mice exhibited normal hippocampal neuronal morphology but impaired hippocampal-dependent learning. Further, hippocampal synaptic remodelling in acute slices from Fibcd1 KO mice was deficient but restored upon enzymatically modulating the ECM. Together, we identified FIBCD1 as an endocytic receptor for GAGs in the brain ECM and a novel gene associated with an NDD, revealing a critical role in nervous system structure, function and plasticity.

The Mechanism Switching the Osteoclast From Short to Long Duration Bone Resorption.

The current models of osteoclastic bone resorption focus on immobile osteoclasts sitting on the bone surface and drilling a pit into the bone matrix. It recently appeared that many osteoclasts also enlarge their pit by moving across the bone surface while resorbing. Drilling a pit thus represents only the start of a resorption event of much larger amplitude. This prolonged resorption activity significantly contributes to pathological bone destruction, but the mechanism whereby the osteoclast engages in this process does not have an answer within the standard bone resorption models. Herein, we review observations that lead to envision how prolonged resorption is possible through simultaneous resorption and migration. According to the standard pit model, the "sealing zone" which surrounds the ruffled border (i.e., the actual resorption apparatus), "anchors" the ruffled border against the bone surface to be resorbed. Herein, we highlight that continuation of resorption demands that the sealing zone "glides" inside the cavity. Thereby, the sealing zone emerges as the structure responsible for orienting and displacing the ruffled border, e.g., directing resorption against the cavity wall. Importantly, sealing zone displacement stringently requires thorough collagen removal from the cavity wall - which renders strong cathepsin K collagenolysis indispensable for engagement of osteoclasts in cavity-enlargement. Furthermore, the sealing zone is associated with generation of new ruffled border at the leading edge, thereby allowing the ruffled border to move ahead. The sealing zone and ruffled border displacements are coordinated with the migration of the cell body, shown to be under control of lamellipodia at the leading edge and of the release of resorption products at the rear. We propose that bone resorption demands more attention to osteoclastic models integrating resorption and migration activities into just one cell phenotype.

Bisphosphonates impair the onset of bone formation at remodeling sites.

Bisphosphonates are widely used anti-osteoporotic drugs targeting osteoclasts. They strongly inhibit bone resorption, but also strongly reduce bone formation. This reduced formation is commonly ascribed to the mechanism maintaining the resorption/formation balance during remodeling. The present study provides evidence for an additional mechanism where bisphosphonates actually impair the onset of bone formation after resorption. The evidence is based on morphometric parameters recently developed to assess the activities reversing resorption to formation. Herein, we compare these parameters in cancellous bone of alendronate- and placebo-treated postmenopausal osteoporotic patients. Alendronate increases the prevalence of eroded surfaces characterized by reversal cells/osteoprogenitors at low cell density and remote from active bone surfaces. This indicates deficient cell expansion on eroded surfaces - an event that is indispensable to start formation. Furthermore, alendronate decreases the coverage of these eroded surfaces by remodeling compartment canopies, a putative source of reversal cells/osteoprogenitors. Finally, alendronate strongly decreases the activation frequency of bone formation, and decreases more the formative compared to the eroded surfaces. All these parameters correlate with each other. These observations lead to a model where bisphosphonates hamper the osteoprogenitor recruitment required to initiate bone formation. This effect results in a larger eroded surface, thereby explaining the well-known paradox that bisphosphonates strongly inhibit bone resorption without strongly decreasing eroded surfaces. The possible mechanism for hampered osteoprogenitor recruitment is discussed: bisphosphonates may decrease the release of osteogenic factors by the osteoclasts, and/or bisphosphonates released by osteoclasts may act directly on neighboring osteoprogenitor cells as reported in preclinical studies.