Total joint replacement, although considered an excellent surgical procedure, can be complicated by osteolysis induced by implant particles and subsequent aseptic loosening of the implant. The pathogenesis of implant-associated osteolysis includes inflammatory and osteolytic processes. The sustained chronic inflammatory response initiated by particulate debris at the implant-bone interface is manifested by recruitment of a wide array of cell types. These cells include macrophages, fibroblasts, giant cells, neutrophils, lymphocytes, and -most importantly -osteoclasts, which are the principal bone resorbing cells. The 'cellular response' entails secretion of osteoclastogenic and inflammatory cytokines that favor exacerbated osteoclast activity and enhanced osteolysis. An appreciation of the complex network that leads to these cellular and inflammatory responses will form a foundation on which to develop therapeutic interventions to combat inflammatory periprosthetic bone loss. IntroductionAseptic loosening of joint implants is a disabling condition that can affect patients 10 to 20 years after joint replacement surgery (Figure 1). Total joint replacement for end-stage joint diseases such as osteoarthritis and inflammatory rheumatoid arthritis is an effective surgical intervention [1]. Unfortunately, wear debris, primarily generated from the prosthetic joint articular surface, remains the major factor limiting the survival of joint implants. Subtle progression of tissue destruction around the implant imposes a major challenge, because signs and symptoms may not be clinically apparent until late stages of failure.The basis of periprosthetic tissue destruction, the so-called 'biologic response' to implant debris, is complex. However, this response is the outcome of multiple factors, including physical and biologic components [2,3]. At the core of the biologic response that leads to osteolysis is activation of the receptor activator of nuclear factor-κB (RANK)/RANK ligand (RANKL) axis, which is indicated by expression of RANK, RANKL, and osteoprotegerin (OPG) in periprosthetic membranes [4,5]. This activation culminates in enhanced osteoclast recruitment and activity adjacent to bone-implant interfaces, leading to osteolysis [6,7]. Aseptic loosening and osteolysis: etiology, pathogenesis, and cellular responsesWear debris is formed at prosthetic joint articulations, modular interfaces, and nonarticulating interfaces [1,8]. Although a wide range of particles has been found, the majority of particles formed are less than 5 µm in diameter and are randomly shaped. Studies have suggested that the cellular response to particles may vary with size, shape, composition, charge, and number of particles [9,10]. Furthermore, it was proposed that particle phagocytosis represents an important component of the cellular response to implants; hence, the size of these particles is significant. In this regard, several reports have estimated that particles ranging from 0.2 to 10 µm in diameter undergo phagocytosis by macrophages ...

Summary Intense effort has been devoted to understanding predisposition to chronic systemic inflammation as this contributes to cardiometabolic disease. We demonstrate that deletion of the macrophage vitamin D receptor (VDR) in mice (KODMAC) is sufficient to induce insulin resistance by promoting M2 macrophage accumulation in the liver, as well as increase cytokine secretion and hepatic glucose production. Moreover, VDR deletion increases atherosclerosis by enabling lipid-laden M2 monocytes to adhere, migrate, and carry cholesterol into the atherosclerotic plaque, and by increasing macrophage cholesterol uptake and esterification. Increased foam cell formation results from lack of VDR-SERCA2b interaction, causing SERCA dysfunction, activation of ER stress-CaMKII-JNKp-PPARγ signaling, and induction of the scavenger receptors CD36 and SR-A1. BM transplant of VDR-expressing cells into KODMAC mice improved insulin sensitivity, suppressed atherosclerosis, and decreased foam cell formation. The immunomodulatory effects of vitamin D in macrophages are thus critical in diet-induced insulin resistance and atherosclerosis in mice. Graphical Abstract

Monocytes differentiate into osteoclasts through stimulation of receptor activator of NF-κB (RANK). Many downstream effectors of RANK play a positive role in osteoclastogenesis, but their relative importance in osteoclast differentiation is unclear. We report the discovery that activation of a single pathway downstream of RANK is sufficient for osteoclast differentiation. In this regard, introduction of constitutively activated IKKβ (IKKβSSEE) but not wild-type IKKβ into monocytes stimulates differentiation of bona fide osteoclasts in the absence of RANK ligand (RANKL). This phenomenon is independent of upstream signals because IKKβSSEE induced the development of bone-resorbing osteoclasts from RANK and IKKα knockout monocytes and in conditions in which NEMO-IKKβ association was inhibited. NF-κB p100 and p105, but not RelB, were critical mediators of this effect. Inflammatory autocrine signaling by tumor necrosis factor α (TNF-α) and interleukin 1 (IL-1) were dispensable for the spontaneous osteoclastogenesis driven by IKKβSSEE. More important, adenoviral gene transfer of IKKβSSEE induced osteoclasts and osteolysis in calvariae and knees of mice. Our data establish the sufficiency of IKKβ activation for osteolysis and suggest that IKKβ hyperactivation may play a role in conditions of pathologic bone destruction refractory to RANK/RANKL proximal therapeutic interventions. © 2010 American Society for Bone and Mineral Research.

NF-kappaB is a vital component of the molecular programs for immune cell development and activation, inflammatory responses, and osteoclast differentiation. This transcriptional regulatory family is activated by diverse immunological and inflammatory stimuli and contributes to both positive feedback of the immune and osteolytic responses as well as their resolution. The ubiquilous expression of NF-kappaB components in osteoclasts and other immune cells creates an opportunity to gain a better understanding of the complex interplay between the immune and skeletal systems in physiological and pathological conditions and also makes NF-kappaB an important target in the treatment of autoimmune, inflammatory, and osteolytic diseases. Indeed, many genetic murine models have recently been developed which highlight the importance of NF-kappaB in basic processes including lymphocyte development, macrophage activation, and osteoclast differentiation. Furthermore, inhibition of NF-kappaB signaling has been demonstrated to ameliorate tissue inflammation and osteolysis in mouse models of inflammatory disease. A more complete understanding of the immunological factors that regulate NF-kappaB and the role that NF-kappaB plays in the immune and skeletal systems will elucidate potential avenues for intervening therapeutically in the pathological conditions of inflammation and osteolysis.

The signal regulatory protein a (SIRPa)/CD47 axis has emerged as an important innate immune checkpoint that enables cancer cell escape from macrophage phagocytosis. SIRPa expression is limited to macrophages, dendritic cells, and neutrophils-cells enriched in the tumor microenvironment. In this study, we present novel anti-SIRP Abs, SIRP-1 and SIRP-2, as an approach to targeting the SIRPa/CD47 axis. Both SIRP-1 and SIRP-2 bind human macrophage SIRPa variants 1 and 2, the most common variants in the human population. SIRP-1 and SIRP-2 are differentiated among reported anti-SIRP Abs in that they induce phagocytosis of solid and hematologic tumor cell lines by human monocyte-derived macrophages as single agents. We demonstrate that SIRP-1 and SIRP-2 disrupt SIRPa/CD47 interaction by two distinct mechanisms: SIRP-1 directly blocks SIRPa/CD47 and induces internalization of SIRPa/Ab complexes that reduce macrophage SIRPa surface levels and SIRP-2 acts via disruption of higher-order SIRPa structures on macrophages. Both SIRP-1 and SIRP-2 engage FcgRII, which is required for single-agent phagocytic activity. Although SIRP-1 and SIRP-2 bind SIRPg with varying affinity, they show no adverse effects on T cell proliferation. Finally, both Abs also enhance phagocytosis when combined with tumor-opsonizing Abs, including a highly differentiated anti-CD47 Ab, AO-176, currently being evaluated in phase 1 clinical trials, NCT03834948 and NCT04445701. SIRP-1 and SIRP-2 are novel, differentiated SIRP Abs that induce in vitro single-agent and combination phagocytosis and show no adverse effects on T cell functionality. These data support their future development, both as single agents and in combination with other anticancer drugs.

Cardiovascular disease (CVD) is the leading cause of mortality in patients with type 2 diabetes mellitus (T2DM). Vitamin D deficiency is not only more prevalent in diabetics but also doubles the risk of developing CVD. However, it is unknown whether 25-hydroxy vitamin D [25(OH)D3] replacement slows monocyte adhesion and migration, critical mechanisms involved in atherosclerosis progression. In this study, monocytes from vitamin D-deficient diabetic patients were cultured either in the patient’s serum or in vitamin D-deficient media with or without 25(OH)D3 treatment. Adding 25(OH)D3 to monocytes cultured in vitamin D-deficient serum or media decreased monocyte adhesion to fibronectin and migration stimulated by monocyte chemotactic protein 1 (MCP-1). Accordingly, 25(OH)D3 decreased adhesion marker β1- and β2-integrin expression and migration receptor chemokine (C-C motif) receptor 2 (CCR2) expression. 25(OH)D3 treatment downregulated monocyte endoplasmic reticulum (ER) stress and scavenger receptor class A, type 1 (SR-A1) expression. The absence of SR-A1 prevented the increased macrophage adhesion and migration induced by vitamin D deficiency. Moreover, the absence of SR-A1 prevented the induction of adhesion and migration and expression of their associated membrane receptors by Thapsigargin, an ER stress inducer. These results identify cellular activation of monocyte/macrophage vitamin D signaling through 25(OH)D3 as a potential mechanism that could modulate adhesion and migration in diabetic subjects.

The transcription factor NF-B is crucial for numerous cellular functions such as survival, differentiation, immunity, and inflammation. A key function of this family of transcription factors is regulation of osteoclast differentiation and function, which in turn controls skeletal homeostasis. The IB kinase (IKK) complex, which contains IKK␣, IKK␤, and IKK␥, is required for activation of NF-B, and deletion of either IKK␣ or IKK␤ resulted with defective osteoclast differentiation and survival. We have recently investigated the details of the mechanisms governing the role of IKK␤ in osteoclastogenesis and found that constitutively active IKK␤ in which serine residues 177/181 were mutated into negatively charged glutamic acids instigates spontaneous bona fide receptor activator of NF-B ligand (RANKL)-independent osteoclastogenesis. To better understand and define the functional role of IKK␤ domains capable of regulating the osteoclastogenic activity of IKK, we investigated key motifs in the activation T loop of IKK␤, which are potentially capable of modulating its osteoclastogenic activity. We discovered that dual serine (traditional serine residues 177/181) and tyrosine (188/199) phosphorylation events are crucial for IKK␤ activation. Mutation of the latter tyrosine residues blunted the NF-B activity of wild type and constitutively active IKK␤, and tyrosine 188/199-deficient IKK␤ inhibited osteoclastogenesis. Thus, tyrosines 188/199 are a novel target for regulating IKK␤ activity, at least in osteoclasts.

Cross-sectional studies indicate consistent associations between low 25(OH)D concentration and increased risk of cardiovascular disease (CVD), but results of randomized control trials (RCTs) are mixed. However, the majority of the RCTs do not focus on type 2 diabetics, potentially obscuring the effects of vitamin D in this population. In vitro 1,25(OH)D downregulates macrophage cholesterol deposition, but the in vivo effects are unknown. To explore potential mechanisms of the effects of vitamin D on CVD risk in patients with type 2 diabetes, we isolated monocytes in a subset of 26 patients from our RCT of diabetics with baseline serum 25(OH)D <25ng/mL randomized to vitamin D 4000 IU/day or placebo for 4 months. Upon enrollment, the mean 25(OH)D level was 17ng/mL, which increased to 36ng/mL after vitamin D and remained unchanged in the placebo group. Before randomization, groups demonstrated similar mean hemoglobin A1c and plasma lipids levels, none of which was significantly altered by vitamin D supplementation. Moreover, assessment of oxidized LDL uptake in monocytes cultured in the patient's own serum before vs. after treatment resulted in >50% reduction in the vitamin D group with no change in the placebo group. This was mediated through suppression of endoplasmic reticulum stress and scavenger receptor CD36 protein expression. The reduction in monocyte cholesterol uptake was reflected in a 19% decrease in total monocyte cholesterol content. Interestingly, cross-sectional analysis of circulating monocytes from vitamin D-deficient vs. sufficient diabetic patients revealed 8-fold higher cholesteryl ester content, confirming the capacity of these monocytes to uptake and carry cholesterol in the circulation. This study identifies a unique circulating cholesterol pool within monocytes that is modulated by vitamin D and has the potential to contribute to CVD in type 2 diabetes.