Cardiovascular Safety of Anti-Sclerostin Therapy in Chronic Kidney Disease

The significance of sclerostin for bone and cardiovascular health in patients with chronic kidney disease (CKD) is complex and incompletely understood. Experimental evidence suggests that anti-sclerostin therapy shows diminished efficacy on bone in the setting of CKD. Limited clinical evidence suggests that the osteoanabolic and anti-resorptive activity is attenuated, but hypocalcemia is more prevalent in patients with advanced CKD (eGFR < 30 mL/min) treated with anti-sclerostin (romosozumab) therapy as compared to patients without kidney disease. Furthermore, sclerostin is prominently expressed in uremic arteries. Whether the inhibition of sclerostin has adverse effects on cardiovascular health in CKD is currently unknown. This review summarizes the current understanding of the physiology and pathophysiology of sclerostin in CKD, with a focus on the cardiovascular safety of anti-sclerostin therapy in patients with or without CKD.

Lumican Inhibits Osteoclastogenesis and Bone Resorption by Suppressing Akt Activity

Lumican, a ubiquitously expressed small leucine-rich proteoglycan, has been utilized in diverse biological functions. Recent experiments demonstrated that lumican stimulates preosteoblast viability and differentiation, leading to bone formation. To further understand the role of lumican in bone metabolism, we investigated its effects on osteoclast biology. Lumican inhibited both osteoclast differentiation and in vitro bone resorption in a dose-dependent manner. Consistent with this, lumican markedly decreased the expression of osteoclastogenesis markers. Moreover, the migration and fusion of preosteoclasts and the resorptive activity per osteoclast were significantly reduced in the presence of lumican, indicating that this protein affects most stages of osteoclastogenesis. Among RANKL-dependent pathways, lumican inhibited Akt but not MAP kinases such as JNK, p38, and ERK. Importantly, co-treatment with an Akt activator almost completely reversed the effect of lumican on osteoclast differentiation. Taken together, our findings revealed that lumican inhibits osteoclastogenesis by suppressing Akt activity. Thus, lumican plays an osteoprotective role by simultaneously increasing bone formation and decreasing bone resorption, suggesting that it represents a dual-action therapeutic target for osteoporosis.

Influence of Cu2+ on Osteoclast Formation and Activity In Vitro

Background: Copper-containing biomaterials are increasingly applied for bone regeneration due to their pro-angiogenetic, pro-osteogenetic and antimicrobial properties. Therefore, the effect of Cu2+ on osteoclasts, which play a major role in bone remodeling was studied in detail. Methods: Human primary osteoclasts, differentiated from human monocytes were differentiated or cultivated in the presence of Cu2+. Osteoclast formation and activity were analyzed by measurement of osteoclast-specific enzyme activities, gene expression analysis and resorption assays. Furthermore, the glutathione levels of the cells were checked to evaluate oxidative stress induced by Cu2+. Results: Up to 8 µM Cu2+ did not induce cytotoxic effects. Activity of tartrate-resistant acid phosphatase (TRAP) was significantly increased, while other osteoclast specific enzyme activities were not affected. However, gene expression of TRAP was not upregulated. Resorptive activity of osteoclasts towards dentin was not changed in the presence of 8 µM Cu2+ but decreased in the presence of extracellular bone matrix. When Cu2+ was added to mature osteoclasts TRAP activity was not increased and resorption decreased only moderately. The glutathione level of both differentiating and mature osteoclasts was significantly decreased in the presence of Cu2+. Conclusions: Differentiating and mature osteoclasts react differently to Cu2+. High TRAP activities are not necessarily related to high resorption.

Expression and enhancement of FABP4 in septoclasts of the growth plate in FABP5-deficient mouse tibiae

In our previous study, fatty acid-binding protein 5 (FABP5) was expressed in septoclasts with long processes which are considered to resorb uncalcified matrix of the growth plate (GP) cartilage, and no apparent abnormalities were detected in the histo-architecture of the GP of FABP5-deficient (FABP5−/−) mice. Those finding lead us to hypothesize that another FABP can compensate the deletion of FABP5 in septoclasts of its gene-mutant mice. Based on the hypothesis, the present study examined the expression levels of several other FABPs in septoclasts and their morphology in FABP5−/− mouse tibiae. Processes of FABP5−/− septoclasts tend to be shorter than wild septoclasts. FABP4-positive septoclasts in FABP5−/− mice were more numerous than those cells in wild mice.Peroxisome proliferator-activated receptor (PPAR) γ was expressed in FABP4-positive septoclasts of FABP5−/− mice as well as mice administered with GW1929, a PPARγ agonist, suggesting that the occurrence of PPARγ induces an increase of FABP4-positive septoclasts. The present finding suggests that the functional exertion of FABP5 in septoclasts is supplemented by FABP4 in normal and FABP5−/− mice, and that the expression of FABP4 is up-regulated in accompany with PPARγ in FABP5−/− for maintenance of resorptive activity in the GP.

Multitasking by the OC Lineage during Bone Infection: Bone Resorption, Immune Modulation, and Microbial Niche

Bone infections, also known as infectious osteomyelitis, are accompanied by significant inflammation, osteolysis, and necrosis. Osteoclasts (OCs) are the bone-resorbing cells that work in concert with osteoblasts and osteocytes to properly maintain skeletal health and are well known to respond to inflammation by increasing their resorptive activity. OCs have typically been viewed merely as effectors of pathologic bone resorption, but recent evidence suggests they may play an active role in the progression of infections through direct effects on pathogens and via the immune system. This review discusses the host- and pathogen-derived factors involved in the in generation of OCs during infection, the crosstalk between OCs and immune cells, and the role of OC lineage cells in the growth and survival of pathogens, and highlights unanswered questions in the field.

Zoledronic Acid in a Mouse Model of Human Fibrous Dysplasia: Ineffectiveness on Tissue Pathology, Formation of “Giant Osteoclasts” and Pathogenetic Implications

We compared the effects of a nitrogen-containing bisphosphonate (N-BP), zoledronic acid (ZA), and an anti-mouse RANKL antibody (anti-mRANKL Ab) on the bone tissue pathology of a transgenic mouse model of human fibrous dysplasia (FD). For comparison, we also reviewed the histological samples of a child with McCune–Albright syndrome (MAS) treated with Pamidronate for 3 years. EF1α-GsαR201C mice with FD-like lesions in the tail vertebrae were treated with either 0.2 mg/kg of ZA at day 0, 7, and 14 or with 300 μg/mouse of anti-mRANKL Ab at day 0 and 21. All mice were monitored by Faxitron and histological analysis was performed at day 42. ZA did not affect the progression of the radiographic phenotype in EF1α-GsαR201C mice. FD-like lesions in the ZA group showed the persistence of osteoclasts, easily detectable osteoclast apoptotic activity and numerous “giant osteoclasts”. In contrast, in the anti-mRANKL Ab-treated mice, osteoclasts were markedly reduced/absent, the radiographic phenotype reverted and the FD-like lesions were extensively replaced by newly formed bone. Numerous “giant osteoclasts” were also detected in the samples of the child with MAS. This study supports the hypothesis that osteoclasts per se, independently of their resorptive activity, are essential for development and expansion of FD lesions.

SUN-347 Glucagon-like Peptide 1 (GLP-1) Acts Directly On Human Osteoclasts To Increase Differentiation And Bone Resorptive Activity

Glucagon-like peptide 1 (GLP-1) is an intestinal hormone released in response to nutrient intake that promotes glucose-dependent insulin secretion by acting upon the pancreatic GLP-1 receptor (GLP-1R). GLP-1R agonists (GLP-1RAs) are widely used in treatment of type 2 diabetes. Preclinical data indicate that GLP-1RAs could be repurposed to treat low bone mass as GLP-1R-depleted mice have higher bone resorption and thinner cortical bones, while insulinopenic and insulin resistant rats have improved bone formation and reduced bone mass deterioration when treated with GLP-1 or GLP-1RAs. However, the effect of GLP-1 and GLP-1RAs on human bone cells remains undetermined. We aimed to elucidate the effect of GLP-1 on primary human osteoclast (OC) and osteoblast (OB) cultures. OCs were differentiated over 10 days from human blood-derived CD14+ monocytes and OBs over 4–6 weeks from human bone. Cells were seeded on bovine bone slices and studies performed using fetal bovine serum, MCSF and RANKL (OC monocultures) or MCSF only (OB monocultures and OB+OC co-cultures). We first investigated the effect of GLP-1 on bone resorptive activity of mature OCs on bovine bone slices. GLP-1 increased the eroded bone surface percentage compared to vehicle in both OC monocultures (1nM P=0.002; 10nM P=0.023; n=8 donors) and OC+OB co-cultures (1nM P=0.013; 10nM P=0.012; n=8 donors). We then tested the effects of GLP-1 on osteoblast activity in OC+OB co-cultures by measuring alkaline phosphatase (ALP). We found that GLP-1 increased ALP in OC+OB cultures (1nM, P=0.049; 10nM, P=0.019) and these effects were reversed by the GLP-1R antagonist exendin 9–39 (1nM, P=0.93, 10nM, P=0.64). However, in OB monocultures GLP-1 had no effects on ALP (1nM P=0.93, 10nM P=0.64) indicating a GLP-1-driven increase in osteoblast activity through osteoclast-osteoblast coupling. We then assessed the effect of GLP-1 on OC differentiation by assessing TRAcP activity. Although there was a trend towards increased TRAcP activity upon stimulation with GLP-1 on day 10 of osteoclastogenesis, this was not statistically significant (1nM P=0.12; n=8 donors; 10nM P=0.29, n=4 donors). Our studies indicated GLP-1 may have a direct effect on osteoclasts, and we therefore sought to characterise GLP-1-mediated signalling in these cells. We assessed the effect of GLP-1 on cAMP signalling using LANCE assays and assessed phosphorylation of ERK proteins by Western blot analysis in human OC cultures. OCs treated with 10nM GLP-1 for 30 minutes had increased cAMP signaling (P=0.004, n=12 bone slices from 2 donors) when compared to vehicle. Furthermore, 10nM GLP-1 induced rapid increases in phosphorylated ERK (P=0.03 following 2 minutes exposure, n=4 blots). In conclusion, our studies reveal that GLP-1 increases activity in primary mature human OCs, and OBs, via OCs. Our signaling studies in OCs indicate this is mediated by direct action of GLP-1 on human bone cells.

Interaction of Brucella abortus with Osteoclasts: a Step toward Understanding Osteoarticular Brucellosis and Vaccine Safety

ABSTRACT Osteoarticular disease is a frequent complication of human brucellosis. Vaccination remains a critical component of brucellosis control, but there are currently no vaccines for use in humans, and no in vitro models for assessing the safety of candidate vaccines in reference to the development of bone lesions currently exist. While the effect of Brucella infection on osteoblasts has been extensively evaluated, little is known about the consequences of osteoclast infection. Murine bone marrow-derived macrophages were derived into mature osteoclasts and infected with B. abortus 2308, the vaccine strain S19, and attenuated mutants S19vjbR and B. abortus ΔvirB2. While B. abortus 2308 and S19 replicated inside mature osteoclasts, the attenuated mutants were progressively killed, behavior that mimics infection kinetics in macrophages. Interestingly, B. abortus 2308 impaired the growth of osteoclasts without reducing resorptive activity, while osteoclasts infected with B. abortus S19 and S19vjbR were significantly larger and exhibited enhanced resorption. None of the Brucella strains induced apoptosis or stimulated nitric oxide or lactose dehydrogenase production in mature osteoclasts. Finally, infection of macrophages or osteoclast precursors with B. abortus 2308 resulted in generation of smaller osteoclasts with decreased resorptive activity. Overall, Brucella exhibits similar growth characteristics in mature osteoclasts compared to the primary target cell, the macrophage, but is able to impair the maturation and alter the resorptive capacity of these cells. These results suggest that osteoclasts play an important role in osteoarticular brucellosis and could serve as a useful in vitro model for both analyzing host-pathogen interactions and assessing vaccine safety.