The short-chain fatty acid receptors Gpr41/43 regulate bone mass by promoting adipogenic differentiation of mesenchymal stem cells.

Bone is a dynamic tissue that is constantly remodeled throughout adult life. Recently, it has been shown that bone turnover decreases shortly after food consumption. This process has been linked to the fermentation of non-digestible food ingredients such as inulin by gut microbes, which results in the production of the short-chain fatty acids (SCFAs) acetate, propionate and butyrate. SCFAs exert various metabolic functions, which in part can be explained by activation of G protein-coupled receptors (Gpr) 41 and 43. However, the potential relevance of a SCFA-Gpr41/43 signaling axis for bone metabolism has not been established. The aim of our study is to investigate the role of Gpr41/43 in bone metabolism and osteogenic differentiation of mesenchymal stem cells. For this purpose, we analyzed the skeletal phenotype of wild type controls (WT) and Gpr41/43 double knockout (Gpr41/43 dKO) mice fed either a chow or an inulin-enriched diet. In addition, we isolated bone marrow derived mesenchymal stem cells from WT and Gpr41/43 dKO mice and differentiated them into osteoblasts in the absence or presence of acetate. MicroCT scanning of femoral bones of Gpr41/43 dKO mice revealed a significant increase of trabecular bone volume and trabecular compared to WT controls. Treatment of WT bone marrow-derived osteoblasts with acetate resulted in decreased mineralization and substantial downregulation of bone formation markers such as Phex, Ptgs2 and Col1a1. Notably, this effect was strongly attenuated in differentiated osteoblasts lacking Gpr41/43. Inversely, acetate supplementation resulted in higher levels of adipocyte marker genes including Pparg, Lpl and Adipoq in bone marrow-derived cells from WT mice, an effect blunted in differentiated cells isolated from Gpr41/43 dKO mice. Overall, these data indicate that acetate regulates bone architecture via SCFA-Gpr41/43 signaling by modulating the osteogenic versus adipogenic differentiation of mesenchymal stem cells.

Arthroscopic Shaver-based Harvest of Minced Cartilage Results in Reduced Chondrocyte Viability and Reduced Quality of Cartilaginous Repair Tissue Compared With Open Harvest and Conventional Fragmentation.

PURPOSE: To characterize and compare the quality of regenerative cartilage tissue (ReCT) after conventional minced cartilage (CMC) and arthroscopic minced cartilage (AMC), in terms of cell viability, gene expression, and matrix synthesis and to investigate the influence of different shaver types. METHODS: Chondral tissue was harvested from the knees of 8 porcine donors. Porcine specimens were euthanized one day before harvest. AMC was created with 2 shaver blades in 2 operating modes (oscillating vs forward) and compared with a scalpel-fragmented CMC control. Before histologic analysis, 50% of the tissue was digested to prevent dedifferentiation of chondrocytes to fibroblasts. Cells were cultured and analyzed for cell viability, gene expression of cartilage-specific markers (aggrecan [ACAN], collagen type II, alpha1 [COL2A1], collagen type I, alpha1 [COL1A1], fibronectin-1 [FN1]), and matrix synthesis (Alcian-blue). RESULTS: AMC tissue contained fewer viable chondrocytes (41%-54% vs 91%; P = .001-.048) compared with CMC. After culture, CMC showed greater expressions of ACAN (27 virtual copy numbers [VCN]/housekeeping gene) and COL2A1 (30 VCN) compared with AMC (ACAN 2-9 VCN, COL2A1 2-7 VCN, P = .001-.039). AMC presented greater expressions of COL1A1 (9-21 VCN) and FN1 (12-17 VCN) than CMC (1 and 6 VCN, P = .001-.050). The signal intensity of the cartilage matrix formed by CMC (86/mm2) was greater than by AMC (7-10 mm2, P = .001-.032). CONCLUSIONS: CMC contained high numbers of viable chondrocytes, resulting in high-quality, hyaline-like ReCT. In contrast, AMC showed impaired chondrocyte quantity and viability, showing greater expressions of fibroblast markers and a decreased formation of mature cartilage matrix in porcine samples. The high chondrogenic potential of CMC to form hyaline-like ReCT was not confirmed for AMC. CLINICAL RELEVANCE: On the basis of our findings, arthroscopic harvest of minced cartilage leads to reduced chondrocyte viability and ReCT quality. Accordingly, CMC and AMC cannot be regarded as synonymous techniques, as arthroscopic techniques seem to be less efficacious.

The emerging role of tranexamic acid and its principal target, plasminogen, in skeletal health.

The worldwide burden of skeletal diseases such as osteoporosis, degenerative joint disease and impaired fracture healing is steadily increasing. Tranexamic acid (TXA), a plasminogen inhibitor and anti-fibrinolytic agent, is used to reduce bleeding with high effectiveness and safety in major surgical procedures. With its widespread clinical application, the effects of TXA beyond anti-fibrinolysis have been noticed and prompted renewed interest in its use. Some clinical trials have characterized the effects of TXA on reducing postoperative infection rates and regulating immune responses in patients undergoing surgery. Also, several animal studies suggest potential therapeutic effects of TXA on skeletal diseases such as osteoporosis and fracture healing. Although a direct effect of TXA on the differentiation and function of bone cells in vitro was shown, few mechanisms of action have been reported. Here, we summarize recent findings of the effects of TXA on skeletal diseases and discuss the underlying plasminogen-dependent and -independent mechanisms related to bone metabolism and the immune response. We furthermore discuss potential novel indications for TXA application as a treatment strategy for skeletal diseases.

Increased ß2-adrenergic signaling promotes fracture healing through callus neovascularization in mice.

Traumatic brain injury (TBI) leads to skeletal changes, including bone loss in the unfractured skeleton, and paradoxically accelerates healing of bone fractures; however, the mechanisms remain unclear. TBI is associated with a hyperadrenergic state characterized by increased norepinephrine release. Here, we identified the ß2-adrenergic receptor (ADRB2) as a mediator of skeletal changes in response to increased norepinephrine. In a murine model of femoral osteotomy combined with cortical impact brain injury, TBI was associated with ADRB2-dependent enhanced fracture healing compared with osteotomy alone. In the unfractured 12-week-old mouse skeleton, ADRB2 was required for TBI-induced decrease in bone formation and increased bone resorption. Adult 30-week-old mice had higher bone concentrations of norepinephrine, and ADRB2 expression was associated with decreased bone volume in the unfractured skeleton and better fracture healing in the injured skeleton. Norepinephrine stimulated expression of vascular endothelial growth factor A and calcitonin gene-related peptide-α (αCGRP) in periosteal cells through ADRB2, promoting formation of osteogenic type-H vessels in the fracture callus. Both ADRB2 and αCGRP were required for the beneficial effect of TBI on bone repair. Adult mice deficient in ADRB2 without TBI developed fracture nonunion despite high bone formation in uninjured bone. Blocking ADRB2 with propranolol impaired fracture healing in mice, whereas the ADRB2 agonist formoterol promoted fracture healing by regulating callus neovascularization. A retrospective cohort analysis of 72 patients with long bone fractures indicated improved callus formation in 36 patients treated with intravenous norepinephrine. These findings suggest that ADRB2 is a potential therapeutic target for promoting bone healing.

Tranexamic Acid Attenuates the Progression of Posttraumatic Osteoarthritis in Mice.

BACKGROUND: Posttraumatic osteoarthritis (OA) is a common disorder associated with a high socioeconomic burden, particularly in young, physically active, and working patients. Tranexamic acid (TXA) is commonly used in orthopaedic trauma surgery as an antifibrinolytic agent to control excessive bleeding. Previous studies have reported that TXA modulates inflammation and bone cell function, both of which are dysregulated during posttraumatic OA disease progression. PURPOSE: To evaluate the therapeutic effects of systemic and topical TXA treatment on the progression of posttraumatic OA in the knee of mice. STUDY DESIGN: Controlled laboratory study. METHODS: OA was induced via anterior cruciate ligament (ACL) transection on the right knee of female mice. Mice were treated with TXA or vehicle intraperitoneally daily or intra-articularly weekly for 4 weeks, starting on the day of surgery. Articular cartilage degeneration, synovitis, bone erosion, and osteophyte formation were scored histologically. Micro-computed tomography evaluation was conducted to measure the subchondral bone microstructure and osteophyte volume. Cartilage thickness and bone remodeling were assessed histomorphometrically. RESULTS: Both systemic and topical TXA treatment significantly reduced cartilage degeneration, synovitis, and bone erosion scores and increased the ratio of hyaline to calcified cartilage thickness in posttraumatic OA. Systemic TXA reversed ACL transection-induced subchondral bone loss and osteophyte formation, whereas topical treatment had no effect. Systemic TXA decreased the number and surface area of osteoclasts, whereas those of osteoblasts were not affected. No effect of topical TXA on osteoblast or osteoclast parameters was observed. CONCLUSION: Both systemic and topical TXA exerted protective effects on the progression of posttraumatic OA. Drug repurposing of TXA may, therefore, be useful for the prevention or treatment of posttraumatic OA, particularly after ACL surgery. CLINICAL RELEVANCE: TXA might be beneficial in patients with posttraumatic OA of the knee.

Transcript-dependent effects of the CALCA gene on the progression of post-traumatic osteoarthritis in mice.

Osteoarthritis represents a chronic degenerative joint disease with exceptional clinical relevance. Polymorphisms of the CALCA gene, giving rise to either a procalcitonin/calcitonin (PCT/CT) or a calcitonin gene-related peptide alpha (αCGRP) transcript by alternative splicing, were reported to be associated with the development of osteoarthritis. The objective of this study was to investigate the role of both PCT/CT and αCGRP transcripts in a mouse model of post-traumatic osteoarthritis (ptOA). WT, αCGRP-/- and CALCA-/- mice were subjected to anterior cruciate ligament transection (ACLT) to induce ptOA of the knee. Mice were sacrificed 4 and 8 weeks post-surgery, followed by micro-CT and histological evaluation. Here we show that the expression of both PCT/CT and αCGRP transcripts is induced in ptOA knees. CALCA-/- mice show increased cartilage degeneration and subchondral bone loss with elevated osteoclast numbers compared to αCGRP-/- and WT mice. Osteophyte formation is reduced to the same extent in CALCA-/- and αCGRP-/- mice compared to WT controls, while a reduced synovitis score is noticed exclusively in mice lacking CALCA. Our data show that expression of the PCT/CT transcript protects from the progression of ptOA, while αCGRP promotes osteophyte formation, suggesting that CALCA-encoded peptides may represent novel targets for the treatment of ptOA.

Piezo1 expression in chondrocytes controls endochondral ossification and osteoarthritis development.

Piezo proteins are mechanically activated ion channels, which are required for mechanosensing functions in a variety of cell types. While we and others have previously demonstrated that the expression of Piezo1 in osteoblast lineage cells is essential for bone-anabolic processes, there was only suggestive evidence indicating a role of Piezo1 and/or Piezo2 in cartilage. Here we addressed the question if and how chondrocyte expression of the mechanosensitive proteins Piezo1 or Piezo2 controls physiological endochondral ossification and pathological osteoarthritis (OA) development. Mice with chondrocyte-specific inactivation of Piezo1 (Piezo1Col2a1Cre), but not of Piezo2, developed a near absence of trabecular bone below the chondrogenic growth plate postnatally. Moreover, all Piezo1Col2a1Cre animals displayed multiple fractures of rib bones at 7 days of age, which were located close to the growth plates. While skeletal growth was only mildly affected in these mice, OA pathologies were markedly less pronounced compared to littermate controls at 60 weeks of age. Likewise, when OA was induced by anterior cruciate ligament transection, only the chondrocyte inactivation of Piezo1, not of Piezo2, resulted in attenuated articular cartilage degeneration. Importantly, osteophyte formation and maturation were also reduced in Piezo1Col2a1Cre mice. We further observed increased Piezo1 protein abundance in cartilaginous zones of human osteophytes. Finally, we identified Ptgs2 and Ccn2 as potentially relevant Piezo1 downstream genes in chondrocytes. Collectively, our data do not only demonstrate that Piezo1 is a critical regulator of physiological and pathological endochondral ossification processes, but also suggest that Piezo1 antagonists may be established as a novel approach to limit osteophyte formation in OA.

The selective norepinephrine reuptake inhibitor reboxetine promotes late-stage fracture healing in mice.

Impaired fracture healing is of high clinical relevance, as up to 15% of patients with long-bone fractures display non-unions. Fracture patients also include individuals treated with selective norepinephrine reuptake inhibitors (SNRI). As SNRI were previously shown to negatively affect bone homeostasis, it remained unclear whether patients with SNRI are at risk of impaired bone healing. Here, we show that daily treatment with the SNRI reboxetine reduces trabecular bone mass in the spine but increases cortical thickness and osteoblast numbers in the femoral midshaft. Most importantly, reboxetine does not impair bone regeneration in a standardized murine fracture model, and even improves callus bridging and biomechanical stability at late healing stages. In sum, reboxetine affects bone remodeling in a site-specific manner. Treatment does not interfere with the early and intermediate stages of bone regeneration and improves healing outcomes of the late-stage fracture callus in mice.

Fracture healing in a mouse model of Hajdu-Cheney-Syndrome with high turnover osteopenia results in decreased biomechanical stability.

Notch signaling regulates cell fate in multiple tissues including the skeleton. Hajdu-Cheney-Syndrome (HCS), caused by gain-of-function mutations in the Notch2 gene, is a rare inherited disease featuring early-onset osteoporosis and increased risk for fractures and non-union. As the impact of Notch2 overactivation on fracture healing is unknown, we studied bone regeneration in mice harboring a human HCS mutation. HCS mice, displaying high turnover osteopenia in the non-fractured skeleton, exhibited only minor morphologic alterations in the progression of bone regeneration, evidenced by static radiological and histological outcome measurements. Histomorphometry showed increased osteoclast parameters in the callus of HCS mice, which was accompanied by an increased expression of osteoclast and osteoblast markers. These observations were accompanied by inferior biomechanical stability of healed femora in HCS mice. Together, our data demonstrate that structural indices of bone regeneration are normal in HCS mice, which, however, exhibit signs of increased callus turnover and display impaired biomechanical stability of healed fractures.

Increased beta2-adrenergic signaling is a targetable stimulus essential for bone healing by promoting callus neovascularization.

Traumatic brain injury (TBI) is associated with a hyperadrenergic state and paradoxically causes systemic bone loss while accelerating fracture healing. Here, we identify the beta2-adrenergic receptor (Adrb2) as a central mediator of these skeletal manifestations. While the negative effects of TBI on the unfractured skeleton can be explained by the established impact of Adrb2 signaling on bone formation, Adrb2 promotes neovascularization of the fracture callus under conditions of high sympathetic tone, including TBI and advanced age. Mechanistically, norepinephrine stimulates the expression of Vegfa and Cgrp primarily in periosteal cells via Adrb2, both of which synergistically promote the formation of osteogenic type-H vessels in the fracture callus. Accordingly, the beneficial effect of TBI on bone repair is abolished in mice lacking Adrb2 or Cgrp, and aged Adrb2-deficient mice without TBI develop fracture nonunions despite high bone formation in uninjured bone. Pharmacologically, the Adrb2 antagonist propranolol impairs, and the agonist formoterol promotes fracture healing in aged mice by regulating callus neovascularization. Clinically, intravenous beta-adrenergic sympathomimetics are associated with improved callus formation in trauma patients with long bone fractures. Thus, Adrb2 is a novel target for promoting bone healing, and widely used beta-blockers may cause fracture nonunion under conditions of increased sympathetic tone.

Inactivation of the gene encoding procalcitonin prevents antibody-mediated arthritis.

BACKGROUND: Procalcitonin (PCT) is applied as a sensitive biomarker to exclude bacterial infections in patients with rheumatoid arthritis (RA) flare-ups. Beyond its diagnostic value, little is known about the pathophysiological role of PCT in RA. METHODS: Collagen antibody-induced arthritis (CAIA) was induced in Calca-deficient mice (Calca-/-), lacking PCT (n = 15), and wild-type (WT) mice (n = 13), while control (CTRL) animals (n = 8 for each genotype) received phosphate-buffered saline. Arthritis severity and grip strength were assessed daily for 10 or 48 days. Articular inflammation, cartilage degradation, and bone lesions were assessed by histology, gene expression analysis, and µ-computed tomography. RESULTS: Serum PCT levels and intra-articular PCT expression increased following CAIA induction. While WT animals developed a full arthritic phenotype, Calca-deficient mice were protected from clinical and histological signs of arthritis and grip strength was preserved. Cartilage turnover markers and Tnfa were exclusively elevated in WT mice. Calca-deficient animals expressed increased levels of Il1b. Decreased bone surface and increased subchondral bone porosity were observed in WT mice, while Calca-deficiency preserved bone integrity. CONCLUSION: The inactivation of Calca and thereby PCT provided full protection from joint inflammation and arthritic bone loss in mice exposed to CAIA. Together with our previous findings on the pathophysiological function of Calca-derived peptides, these data indicate an independent pro-inflammatory role of PCT in RA.

Efficacy of zoledronic acid for the elimination of disseminated tumor cells in a clinically relevant, spontaneously metastatic prostate cancer xenograft model.

Bone metastases develop in >90 % of patients with castration-resistant prostate cancer (PCa) through complex interactions between the bone microenvironment and tumor cells. Previous androgen-deprivation therapy (ADT), which is known to cause bone loss, as well as anti-resorptive agents such as zoledronic acid (ZA), used to prevent skeletal complications, may influence these interactions and thereby the growth of disseminated tumor cells (DTC) in the bone marrow (BM). Here, a spontaneously metastatic xenograft tumor model of human PCa was further optimized to mimic the common clinical situation of ADT (castration) combined with primary tumor resection in vivo. The effects of these interventions, alone or in combination with ZA treatment, on tumor cell dissemination to the BM and other distant sites were analyzed. Metastatic burden was quantified by human-specific Alu-qPCR, bioluminescence imaging (BLI), and immunohistochemistry. Further, bone remodeling was assessed by static histomorphometry and serum parameters. Initial comparative analysis between NSG and SCID mice showed that spontaneous systemic dissemination of subcutaneous PC-3 xenograft tumors was considerably enhanced in NSG mice. Primary tumor resection and thereby prolonged observational periods resulted in a higher overall metastatic cell load at necropsy and tumor growth alone caused significant bone loss, which was further augmented by surgical castration. In addition, castrated mice showed a strong trend towards higher bone metastasis loads. Weekly treatment of mice with ZA completely prevented castration- and tumor-induced bone loss but had no effect on bone metastasis burden. Conversely, the total lung metastasis load as determined by BLI was significantly decreased upon ZA treatment. These findings provide a basis for future research on the role of ZA not only in preventing skeletal complications but also in reducing metastasis to other organs.

Increased concentrations of conjugated bile acids are associated with osteoporosis in PSC patients.

Primary sclerosing cholangitis (PSC) is an idiopathic cholestatic liver disease characterized by chronic inflammation and progressive fibrosis of intra- and extrahepatic bile ducts. Osteoporosis is a frequent comorbidity in PSC, and we could previously demonstrate that IL17-dependent activation of bone resorption is the predominant driver of bone loss in PSC. Since we additionally observed an unexpected heterogeneity of bone mineral density in our cohort of 238 PSC patients, the present study focused on a comparative analysis of affected individuals with diagnosed osteoporosis (PSCOPO, n = 10) or high bone mass (PSCHBM, n = 7). The two groups were not distinguishable by various baseline characteristics, including liver fibrosis or serum parameters for hepatic function. In contrast, quantification of serum bile acid concentrations identified significant increases in the PSCOPO group, including glycoursodeoxycholic acid (GUDCA), an exogenous bile acid administered to both patient groups. Although cell culture experiments did not support the hypothesis that an increase in circulating bile levels is a primary cause of PSC-associated osteoporosis, the remarkable differences of endogenous bile acids and GUDCA in the serum of PSCOPO patients strongly suggest a yet unknown impairment of biliary metabolism and/or hepatic bile acid clearance in this patient subgroup, which is independent of liver fibrosis.

Systemic calcitonin gene-related peptide receptor antagonism decreases survival in a porcine model of polymicrobial sepsis: blinded randomised controlled trial.

BACKGROUND: Calcitonin gene-related peptide (CGRP) and procalcitonin, which are overexpressed in sepsis, exert distinct immunomodulatory effects mediated through the CGRP receptor. The CGRP receptor antagonist olcegepant improves survival in murine sepsis. This study evaluated whether CGRP receptor antagonism is similarly beneficial in a porcine model of polymicrobial sepsis. METHODS: We conducted a prospective randomised, controlled, investigator-blinded trial in adult pigs of either sex, that were anaesthetised and ventilated before sepsis was induced by polymicrobial (autologous) faecal peritonitis. After the onset of early septic shock (systolic blood pressure <90 mm Hg or >10% decline from baseline MAP), pigs were resuscitated (i.v. fluid/antibiotics/vasopressors) and randomised to receive either i.v. olcegepant (n=8) or vehicle control (n=8). The primary outcome was time to death, euthanasia required up to 72 h after surgery (according to predefined severe cardiorespiratory failure), or both. Secondary outcomes included haemodynamic changes, and systemic as well as organ inflammation (mRNA expression). RESULTS: Septic shock developed 8.7 h (inter-quartile range, 5.8-11.1 h) after the onset of faecal peritonitis. Olcegepant worsened survival, with 6/8 pigs randomised to the control group surviving 72.0 h (50.9-72.0 h), compared with 3/8 pigs receiving olcegepant surviving 51.3 h (12.5-72.0 h; P=0.01). At 48 h, lower MAP and higher cardiac output occurred in pigs receiving olcegepant. Cardiac, hepatic, and renal injury was not different between pigs randomised to receive olcegepant or vehicle. Olcegepant reduced mRNA expression of several inflammation-related cytokines and CD68+ macrophages in liver but not in lung tissue. CONCLUSIONS: CGRP receptor antagonism with olcegepant was not beneficial in this porcine model of polymicrobial sepsis, which closely mimics human sepsis.

Spine Metastases in Immunocompromised Mice after Intracardiac Injection of MDA-MB-231-SCP2 Breast Cancer Cells.

Breast cancer cells frequently metastasize to bone, where their interaction with bone remodeling cell types enhances osteolytic bone destruction. Importantly, however, whereas skeletal analyses of xenograft models are usually restricted to hindlimb bones, human skeletal metastases are far more frequent in the spine, where trabecular bone mass is higher compared to femur or tibia. Here, we addressed whether breast cancer cells injected into immunocompromised mice metastasize to the spine and if this process is influenced by the amount of trabecular bone. We also took advantage of mice carrying the Col1a1-Krm2 transgene, which display severe osteoporosis. After crossing this transgene into the immunocompromised NSG background we injected MDA-MB-231-SCP2 breast cancer cells and analyzed their distribution three weeks thereafter. We identified more tumor cells and clusters of different size in spine sections than in femora, which allowed influences on bone remodeling cell types to be analyzed by comparing tumor-free to tumor-burdened areas. Unexpectedly, the Col1a1-Krm2 transgene did not affect spreading and metastatic outgrowth of MDA-MB-231-SCP2 cells, suggesting that bone tumor interactions are more relevant at later stages of metastatic progression.

G6b-B regulates an essential step in megakaryocyte maturation.

G6b-B is a megakaryocyte lineage-specific immunoreceptor tyrosine-based inhibition motif-containing receptor, essential for platelet homeostasis. Mice with a genomic deletion of the entire Mpig6b locus develop severe macrothrombocytopenia and myelofibrosis, which is reflected in humans with null mutations in MPIG6B. The current model proposes that megakaryocytes lacking G6b-B develop normally, whereas proplatelet release is hampered, but the underlying molecular mechanism remains unclear. We report on a spontaneous recessive single nucleotide mutation in C57BL/6 mice, localized within the intronic region of the Mpig6b locus that abolishes G6b-B expression and reproduces macrothrombocytopenia, myelofibrosis, and osteosclerosis. As the mutation is based on a single-nucleotide exchange, Mpig6bmut mice represent an ideal model to study the role of G6b-B. Megakaryocytes from these mice were smaller, displayed a less-developed demarcation membrane system, and had a reduced expression of receptors. RNA sequencing revealed a striking global reduction in the level of megakaryocyte-specific transcripts, in conjunction with decreased protein levels of the transcription factor GATA-1 and impaired thrombopoietin signaling. The reduced number of mature MKs in the bone marrow was corroborated on a newly developed Mpig6b-null mouse strain. Our findings highlight an unexpected essential role of G6b-B in the early differentiation within the megakaryocytic lineage.

Procalcitonin is expressed in osteoblasts and limits bone resorption through inhibition of macrophage migration during intermittent PTH treatment.

Intermittent injections of parathyroid hormone (iPTH) are applied clinically to stimulate bone formation by osteoblasts, although continuous elevation of parathyroid hormone (PTH) primarily results in increased bone resorption. Here, we identified Calca, encoding the sepsis biomarker procalcitonin (ProCT), as a novel target gene of PTH in murine osteoblasts that inhibits osteoclast formation. During iPTH treatment, mice lacking ProCT develop increased bone resorption with excessive osteoclast formation in both the long bones and axial skeleton. Mechanistically, ProCT inhibits the expression of key mediators involved in the recruitment of macrophages, representing osteoclast precursors. Accordingly, ProCT arrests macrophage migration and causes inhibition of early but not late osteoclastogenesis. In conclusion, our results reveal a potential role of osteoblast-derived ProCT in the bone microenvironment that is required to limit bone resorption during iPTH.

The calcitonin receptor protects against bone loss and excessive inflammation in collagen antibody-induced arthritis.

Pharmacological application of teleost calcitonin (CT) has been shown to exert chondroprotective and anti-resorptive effects in patients with rheumatoid arthritis (RA). However, the role of endogenous CT that signals through the calcitonin receptor (CTR) remains elusive. Collagen II antibody-induced arthritis (CAIA) was stimulated in wild type (WT) and CTR-deficient (Calcr-/-) mice. Animals were monitored over 10 or 48 days. Joint inflammation, cartilage degradation, and bone erosions were assessed by clinical arthritis score, histology, histomorphometry, gene expression analysis, and µ-computed tomography. CAIA was accompanied by elevated systemic CT levels and CTR expression in the articular cartilage. Inflammation, cartilage degradation, and systemic bone loss were more pronounced in Calcr-/- CAIA mice. Expression of various pro-inflammatory, bone resorption, and catabolic cartilage markers were exclusively increased in Calcr-/- CAIA mice. Endogenous CT signaling through the mammalian CTR has the potential to protect against joint inflammation, cartilage degradation, and excessive bone remodeling in experimental RA.

Tumor cell E-selectin ligands determine partialefficacy of bortezomib on spontaneous lung metastasis formation of solid human tumors in vivo.

Extravasation of circulating tumor cells (CTCs) is critical for metastasis and is initiated by adhesive interactions between glycoligands on CTCs and E-selectin on endothelia. Here, we show that the clinically approved proteasome inhibitor bortezomib (BZM; Velcade) counteracts the cytokine-dependent induction of E-selectin in the lung mediated by the primary tumor, thereby impairing endothelial adhesion and thus spontaneous lung metastasis in vivo. However, the efficacy of BZM crucially depends on the tumor cells' E-selectin ligands, which determine distinct adhesion patterns. The canonical ligands sialyl-Lewis A (sLeA) and sLeX mediate particularly high-affinity E-selectin binding so that the incomplete E-selectin-reducing effect of BZM is not sufficient to disrupt adhesion or metastasis. In contrast, tumor cells lacking sLeA/X nevertheless bind E-selectin, but with low affinity, so that adhesion and lung metastasis are significantly diminished. Such low-affinity E-selectin ligands apparently consist of sialylated MGAT5 products on CD44. BZM no longer has anti-metastatic activity after CD44 knockdown in sLeA/X-negative tumor cells or E-selectin knockout in mice. sLeA/X can be determined by immunohistochemistry in cancer samples, which might aid patient stratification. These data suggest that BZM might act as a drug for inhibiting extravasation and thus distant metastasis formation in malignancies expressing low-affinity E-selectin ligands.