Circulating autotaxin levels in healthy teenagers: Data from the Vitados cohort.

Autotaxin (ATX) is a secreted enzyme with a lysophospholipase D activity, mainly secreted by adipocytes and widely expressed. Its major function is to convert lysophosphatidylcholine (LPC) into lysophosphatidic acid (LPA), an essential bioactive lipid involved in multiple cell processes. The ATX-LPA axis is increasingly studied because of its involvement in numerous pathological conditions, more specifically in inflammatory or neoplastic diseases, and in obesity. Circulating ATX levels gradually increase with the stage of some pathologies, such as liver fibrosis, thus making them a potentially interesting non-invasive marker for fibrosis estimation. Normal circulating levels of ATX have been established in healthy adults, but no data exist at the pediatric age. The aim of our study is to describe the physiological concentrations of circulating ATX levels in healthy teenagers through a secondary analysis of the VITADOS cohort. Our study included 38 teenagers of Caucasian origin (12 males, 26 females). Their median age was 13 years for males and 14 years for females, ranging from Tanner 1 to 5. BMI was at the 25th percentile for males and 54th percentile for females, and median blood pressure was normal. ATX median levels were 1,049 (450-2201) ng/ml. There was no difference in ATX levels between sexes in teenagers, which was in contrast to the male and female differences described in the adult population. ATX levels significantly decreased with age and pubertal status, reaching adult levels at the end of puberty. Our study also suggested positive correlations between ATX levels and blood pressure (BP), lipid metabolism, and bone biomarkers. However, except for LDL cholesterol, these factors were also significantly correlated with age, which might be a confounding factor. Still, a correlation between ATX and diastolic BP was described in obese adult patients. No correlation was found between ATX levels and inflammatory marker C-reactive protein (CRP), Body Mass Index (BMI), and biomarkers of phosphate/calcium metabolism. In conclusion, our study is the first to describe the decline in ATX levels with puberty and the physiological concentrations of ATX levels in healthy teenagers. It will be of utmost importance when performing clinical studies in children with chronic diseases to keep these kinetics in mind, as circulating ATX might become a non-invasive prognostic biomarker in pediatric chronic diseases.

Inhibition of alkaline phosphatase impairs dyslipidemia and protects mice from atherosclerosis.

Calcium accumulation in atherosclerotic plaques predicts cardiovascular mortality, but the mechanisms responsible for plaque calcification and how calcification impacts plaque stability remain debated. Tissue-nonspecific alkaline phosphatase (TNAP) recently emerged as a promising therapeutic target to block cardiovascular calcification. In this study, we sought to investigate the effect of the recently developed TNAP inhibitor SBI-425 on atherosclerosis plaque calcification and progression. TNAP levels were investigated in ApoE-deficient mice fed a high-fat diet from 10 weeks of age and in plaques from the human ECLAGEN biocollection (101 calcified and 14 non-calcified carotid plaques). TNAP was inhibited in mice using SBI-425 administered from 10 to 25 weeks of age, and in human vascular smooth muscle cells (VSMCs) with MLS-0038949. Plaque calcification was imaged in vivo with 18F-NaF-PET/CT, ex vivo with osteosense, and in vitro with alizarin red. Bone architecture was determined with µCT. TNAP activation preceded and predicted calcification in human and mouse plaques, and TNAP inhibition prevented calcification in human VSMCs and in ApoE-deficient mice. More unexpectedly, TNAP inhibition reduced the blood levels of cholesterol and triglycerides, and protected mice from atherosclerosis, without impacting the skeletal architecture. Metabolomics analysis of liver extracts identified phosphocholine as a substrate of liver TNAP, who's decreased dephosphorylation upon TNAP inhibition likely reduced the release of cholesterol and triglycerides into the blood. Systemic inhibition of TNAP protects from atherosclerosis, by ameliorating dyslipidemia, and preventing plaque calcification.

Blockade of Platelet CysLT1R Receptor with Zafirlukast Counteracts Platelet Protumoral Action and Prevents Breast Cancer Metastasis to Bone and Lung.

Metastases are the main cause of death in cancer patients, and platelets are largely known for their contribution in cancer progression. However, targeting platelets is highly challenging given their paramount function in hemostasis. Using a high-throughput screening and platelet-induced breast tumor cell survival (PITCS) assay as endpoint, we identified the widely used anti-asthmatic drugs and cysteinyl leukotriene receptor 1 (CysLT1R) antagonists, zafirlukast and montelukast, as new specific blockers of platelet protumoral action. Here, we show that human MDA-B02 breast cancer cells produce CysLT through mechanisms involving microsomal glutathione-S-transferase 1/2/3 (MGST1/2/3) and that can modulate cancer cell-platelet interactions via platelet-CysLT1R. CysLT1R blockade with zafirlukast decreased platelet aggregation and adhesion on cancer cells and inhibited PITCS, migration, and invasion in vitro. Zafirlukast significantly reduced, by 90%, MDA-B02 cell dissemination to bone in nude mice and reduced by 88% 4T1 spontaneous lung metastasis formation without affecting primary tumor growth. Combined treatment of zafirlukast plus paclitaxel totally inhibited metastasis of 4T1 cells to the lungs. Altogether, our results reveal a novel pathway mediating the crosstalk between cancer cells and platelets and indicate that platelet CysLT1R represents a novel therapeutic target to prevent metastasis without affecting hemostasis.

X-Linked Hypophosphatemia, Not Only a Skeletal Disease But Also a Chronic Inflammatory State.

CONTEXT: X-linked hypophosphatemia (XLH) is a rare genetic disease caused by a primary excess of fibroblast growth factor 23 (FGF23). FGF23 has been associated with inflammation and impaired osteoclastogenesis, but these pathways have not been investigated in XLH. OBJECTIVE: This work aimed to evaluate whether XLH patients display peculiar inflammatory profile and increased osteoclastic activity. METHODS: We performed a prospective, multicenter, cross-sectional study analyzing transcript expression of 8 inflammatory markers (Il6, Il8, Il1ß, CXCL1, CCL2, CXCR3, Il1R, Il6R) by real-time quantitative polymerase chain reaction on peripheral blood mononuclear cells (PBMCs) purified from total blood samples extracted from patients and healthy control individuals. The effect of native/active vitamin D on osteoclast formation was also assessed in vitro from XLH patients' PBMCs. RESULTS: In total, 28 XLH patients (17 children, among them 6 undergoing standard of care [SOC] and 11 burosumab therapy) and 19 controls were enrolled. Expression of most inflammatory markers was significantly increased in PBMCs from XLH patients compared to controls. No differences were observed between the burosumab and SOC subgroups. Osteoclast formation was significantly impaired in XLH patients. XLH mature osteoclasts displayed higher levels of inflammatory markers, being however lower in cells derived from the burosumab subgroup (as opposed to SOC). CONCLUSION: We describe for the first time a peculiar inflammatory profile in XLH. Since XLH patients have a propensity to develop arterial hypertension, obesity, and enthesopathies, and because inflammation can worsen these clinical outcomes, we hypothesize that inflammation may play a critical role in these extraskeletal complications of XLH.

Autotaxin/Lysophosphatidic Acid Axis: From Bone Biology to Bone Disorders.

Lysophosphatidic acid (LPA) is a natural bioactive phospholipid with pleiotropic activities affecting multiple tissues, including bone. LPA exerts its biological functions by binding to G-protein coupled LPA receptors (LPA1-6) to stimulate cell migration, proliferation, and survival. It is largely produced by autotaxin (ATX), a secreted enzyme with lysophospholipase D activity that converts lysophosphatidylcholine (LPC) into active LPA. Beyond its enzymatic activity, ATX serves as a docking molecule facilitating the efficient delivery of LPA to its specific cell surface receptors. Thus, LPA effects are the result of local production by ATX in a given tissue or cell type. As a consequence, the ATX/LPA axis should be considered as an entity to better understand their roles in physiology and pathophysiology and to propose novel therapeutic strategies. Herein, we provide not only an extensive overview of the relevance of the ATX/LPA axis in bone cell commitment and differentiation, skeletal development, and bone disorders, but also discuss new working hypotheses emerging from the interplay of ATX/LPA with well-established signaling pathways regulating bone mass.

Pain-like behavior in the collagen antibody-induced arthritis model is regulated by lysophosphatidic acid and activation of satellite glia cells.

Inflammatory and neuropathic-like components underlie rheumatoid arthritis (RA)-associated pain, and lysophosphatidic acid (LPA) is linked to both joint inflammation in RA patients and to neuropathic pain. Thus, we investigated a role for LPA signalling using the collagen antibody-induced arthritis (CAIA) model. Pain-like behavior during the inflammatory phase and the late, neuropathic-like phase of CAIA was reversed by a neutralizing antibody generated against LPA and by an LPA1/3 receptor inhibitor, but joint inflammation was not affected. Autotaxin, an LPA synthesizing enzyme was upregulated in dorsal root ganglia (DRG) neurons during both CAIA phases, but not in joints or spinal cord. Late-phase pronociceptive neurochemical changes in the DRG were blocked in Lpar1 receptor deficient mice and reversed by LPA neutralization. In vitro and in vivo studies indicated that LPA regulates pain-like behavior via the LPA1 receptor on satellite glia cells (SGCs), which is expressed by both human and mouse SGCs in the DRG. Furthermore, CAIA-induced SGC activity is reversed by phospholipid neutralization and blocked in Lpar1 deficient mice. Our findings suggest that the regulation of CAIA-induced pain-like behavior by LPA signalling is a peripheral event, associated with the DRGs and involving increased pronociceptive activity of SGCs, which in turn act on sensory neurons.

Emerging role of cysteinyl LTs in cancer.

Cysteinyl leukotrienes (CysLTs) are inflammatory lipid mediators that play a central role in the pathophysiology of several inflammatory diseases. Recently, there has been an increased interest in determining how these lipid mediators orchestrate tumour development and metastasis through promoting a pro-tumour micro-environment. Up-regulation of CysLTs receptors and CysLTs production is found in a number of cancers and has been associated with increased tumorigenesis. Understanding the molecular mechanisms underlying the role of CysLTs and their receptors in cancer progression will help investigate the potential of targeting CysLTs signalling for anti-cancer therapy. This review gives an overview of the biological effects of CysLTs and their receptors, along with current knowledge of their regulation and expression. It also provides a recent update on the molecular mechanisms that have been postulated to explain their role in tumorigenesis and on the potential of anti-CysLTs in the treatment of cancer.

Peripheral Blood Mononuclear Cells (PBMCs) to Dissect the Underlying Mechanisms of Bone Disease in Chronic Kidney Disease and Rare Renal Diseases.

PURPOSE OF REVIEW: To describe the methods that can be used to obtain functional and mature osteoclasts from peripheral blood mononuclear cells (PBMCs) and report the data obtained with this model in two peculiar diseases, namely pediatric chronic kidney disease-associated mineral and bone disorders (CKD-MBD) and nephropathic cystinosis. To discuss future research possibilities in the field. RECENT FINDINGS: Bone tissue undergoes continuous remodeling throughout life to maintain bone architecture; it involves two processes: bone formation and bone resorption with the coordinated activity of osteoblasts, osteoclasts, and osteocytes. Animal models fail to fully explain human bone pathophysiology during chronic kidney disease, mainly due to interspecies differences. The development of in vitro models has permitted to mimic human bone-related diseases as an alternative to in vivo models. Since 1997, osteoclasts have been generated in cell cultures, notably when culturing PBMCs with specific growth factors and cytokines (i.e., M-CSF and RANK-L), without the need for osteoblasts or stromal cells. These models may improve the global understanding of bone pathophysiology. They can be been used not only to evaluate the direct effects of cytokines, hormones, cells, or drugs on bone remodeling during CKD-MBD, but also in peculiar genetic renal diseases inducing specific bone impairment.

Response to Cysteamine in Osteoclasts Obtained from Patients with Nephropathic Cystinosis: A Genotype/Phenotype Correlation.

Bone complications of cystinosis have been recently described. The main objectives of this paper were to determine in vitro the impact of CTNS mutations and cysteamine therapy on human osteoclasts and to carry out a genotype-phenotype analysis related to osteoclastic differentiation. Human osteoclasts were differentiated from peripheral blood mononuclear cells (PBMCs) and were treated with increasing doses of cysteamine (0, 50, 200 µM) and then assessed for osteoclastic differentiation. Results are presented as median (min-max). A total of 17 patients (mainly pediatric) were included, at a median age of 14 (2-61) years, and a eGFR of 64 (23-149) mL/min/1.73 m2. Most patients (71%) were under conservative kidney management (CKM). The others were kidney transplant recipients. Three functional groups were distinguished for CTNS mutations: cystinosin variant with residual cystin efflux activity (RA, residual activity), inactive cystinosin variant (IP, inactive protein), and absent protein (AP). PBMCs from patients with residual cystinosin activity generate significantly less osteoclasts than those obtained from patients of the other groups. In all groups, cysteamine exerts an inhibitory effect on osteoclastic differentiation at high doses. This study highlights a link between genotype and osteoclastic differentiation, as well as a significant impact of cysteamine therapy on this process in humans.

Tamoxifen Treatment in the Neonatal Period Affects Glucose Homeostasis in Adult Mice in a Sex-Dependent Manner.

Tamoxifen is a selective estrogen receptor modulator used to activate the CREERT2 recombinase, allowing tissue-specific and temporal control of the somatic mutagenesis to generate transgenic mice. Studies integrating development and metabolism require a genetic modification induced by a neonatal tamoxifen administration. Here, we investigate the effects of a neonatal tamoxifen administration on energy homeostasis in adult male and female C57BL/6J mice. C57BL/6J male and female mouse pups received a single injection of tamoxifen 1 day after birth (NTT) and were fed a high-fat/high-sucrose diet at 6 weeks of age. We measured weight, body composition, glucose and insulin tolerance, basal metabolism, and tibia length and weight in adult mice. The neonatal tamoxifen administration exerted long-term, sex-dependent effects on energy homeostasis. NTT female mice became overweight and developed impaired glucose control in comparison to vehicle-treated littermates. NTT females exhibited 60% increased fat mass, increased food intake, decreased physical activity and energy expenditure, impaired glucose and insulin tolerance, and fasting hyperglycemia and hyperinsulinemia. In contrast, NTT male mice exhibited a modest amelioration of glucose and insulin tolerance and long-term decreased lean mass linked to decreased bone weight. These results suggest that the neonatal tamoxifen administration exerted a marked and sex-dependent influence on adult energy homeostasis and bone weight and must therefore be used with caution for the development of transgenic mouse models regarding studies on energy homeostasis and bone biology.

Inhibition of Osteoclast Differentiation by 1.25-D and the Calcimimetic KP2326 Reveals 1.25-D Resistance in Advanced CKD.

Active vitamin D analogs and calcimimetics are the main therapies used for treating secondary hyperparathyroidism (SHPT) in patients with chronic kidney disease (CKD). Peripheral blood mononuclear cells of 19 pediatric patients with CKD1-5D and 6 healthy donors (HD) were differentiated into mature osteoclasts with receptor activator of NF-κB ligand (RANKL) and macrophage colony-stimulating factor (M-CSF). The effects of single or combined treatment with active vitamin D (1.25-D) and/or calcimimetic KP2326 were evaluated on osteoclastic differentiation and osteoclastic-mediated bone resorption. Although 1.25-D inhibited osteoclastic differentiation, a significant resistance to 1.25-D was observed when glomerular filtration rate decreased. A significant albeit less important inhibitory effect of KP2326 on osteoclastic differentiation was also found both in cells derived from HD and CKD patients, through a putative activation of the Erk pathway. This inhibitory effect was not modified by CKD stage. Combinatorial treatment with 1.25-D and KP2326 did not result in synergistic effects. Last, KP2326 significantly inhibited osteoclast-mediated bone resorption. Both 1.25-D and KP2326 inhibit osteoclastic differentiation, however, to a different extent. There is a progressive resistance to 1.25-D in advanced CKD that is not found with KP2326. KP2326 also inhibits bone resorption. Given that 1.25-D has no effect on osteoclastic resorption activity and that calcimimetics also have direct anabolic effects on osteoblasts, there is an experimental rationale that could favor the use of decreased doses of 1.25-D with low doses of calcimimetics in SHPT in dialysis to improve the underlying osteodystrophy. However, this last point deserves confirmatory clinical studies. © 2020 American Society for Bone and Mineral Research.

Expression of the type 1 lysophosphatidic acid receptor in osteoblastic cell lineage controls both bone mineralization and osteocyte specification.

Lysphosphatidic acid (LPA) is a major natural bioactive lipid mediator whose biological functions affect multiple organs. These include bone as demonstrated by global Lpar1-knockout mice (Lpar1-/-) which present a bone growth defect. LPA acts on all bone cells including osteoblasts, that are responsible for bone formation, and osteoclasts, which are specialized cells that resorb bone. LPA appears as a potential new coupling molecule during bone remodeling. LPA1 is the most ubiquitous LPA receptor among the six LPA receptor family members (LPA1-6). To better understand the specific role of LPA via its receptor LPA1 in osteoblastic cell lineage we generated osteoblast-specific Lpar1 knockout mice (Lpar1-∆Ob) by crossing Lpar1flox/flox and Osx:Cre+ mouse lines. Lpar1-∆Ob mice do not recapitulate the bone defects of Lpar1-/- mice but revealed reduced bone mineralization and decreased cortical thickness, as well as increased bone porosity associated with an augmentation in the lacunae areas of osteocyte and their apoptotic yield. In vitro, primary Lpar1-∆Ob and immortalized cl1-Ob-Lpar1-/- osteoblasts revealed a remarkable premature expression of alkaline phosphatase, reduced cell proliferation associated with decreased YAP-P nuclear accumulation, and reduced mineralization activity. Osteocyte specification is markedly impaired as demonstrated by reduced expression of early (E11) and late (DMP1, DKK1, SOST) osteocyte markers ex vivo in enriched osteocytic fractions of Lpar1-∆Ob mouse bone explants. In addition, E11 expression and dendrite formation induced by FGF2 are markedly impaired in both primary Lpar1-∆Ob and immortalized cl1-Ob-Lpar1-/- osteoblasts. Taken together these results suggest a new role for LPA in bone mass control via bone mineralization and osteocyte function.

The Bone Morphogenetic Protein Signaling Inhibitor LDN-193189 Enhances Metastasis Development in Mice.

Breast cancer with bone metastasis is essentially incurable with current anticancer therapies. The bone morphogenetic protein (BMP) pathway is an attractive therapeutic candidate, as it is involved in the bone turnover and in cancer cell formation and their colonization of distant organs such as the bone. We previously reported that in breast cancer cells, the ZNF217 oncogene drives BMP pathway activation, increases the metastatic growth rate in the bone, and accelerates the development of severe osteolytic lesions in mice. In the present study, we aimed at investigating the impact of the LDN-193189 compound, a potent inhibitor of the BMP type I receptor, on metastasis development in vivo. ZNF217-revLuc cells were injected into the left ventricle of nude mice (n = 16) while control mice (n = 13) were inoculated with control pcDNA6-revLuc cells. Mice from each group were treated or not with LDN-193189 for 35 days. We found that systemic LDN-193189 treatment of mice significantly enhanced metastasis development, by increasing both the number and the size of metastases. In pcDNA6-revLuc-injected mice, LDN-193189 also affected the kinetics of metastasis emergence. Altogether, these data suggest that in vivo, LDN-193189 might affect the interaction between breast cancer cells and the bone environment, favoring the emergence and development of multiple metastases. Hence, our report highlights the importance of the choice of drugs and therapeutic strategies used in the management of bone metastases.

Osteoclast-Derived Autotaxin, a Distinguishing Factor for Inflammatory Bone Loss.

OBJECTIVE: The severity of rheumatoid arthritis (RA) correlates directly with bone erosions arising from osteoclast (OC) hyperactivity. Despite the fact that inflammation may be controlled in patients with RA, those in a state of sustained clinical remission or low disease activity may continue to accrue erosions, which supports the need for treatments that would be suitable for long-lasting inhibition of OC activity without altering the physiologic function of OCs in bone remodeling. Autotaxin (ATX) contributes to inflammation, but its role in bone erosion is unknown. METHODS: ATX was targeted by inhibitory treatment with pharmacologic drugs and also by conditional inactivation of the ATX gene Ennp2 in murine OCs (ΔATXC tsk ). Arthritic and erosive diseases were studied in human tumor necrosis factor-transgenic (hTNF+/- ) mice and mice with K/BxN serum transfer-induced arthritis. Systemic bone loss was also analyzed in mice with lipopolysaccharide (LPS)-induced inflammation and estrogen deprivation. Joint inflammation and bone erosion were assessed by histology and micro-computed tomography. The role of ATX in RA was also examined in OC differentiation and activity assays. RESULTS: OCs present at sites of inflammation overexpressed ATX. Pharmacologic inhibition of ATX in hTNF+/- mice, as compared to vehicle-treated controls, significantly mitigated focal bone erosion (36% decrease; P < 0.05) and systemic bone loss (43% decrease; P < 0.05), without affecting synovial inflammation. OC-derived ATX was revealed to be instrumental in OC bone resorptive activity and was up-regulated by the inflammation elicited in the presence of TNF or LPS. Specific loss of ATX in OCs from mice subjected to ovariectomy significantly protected against the systemic bone loss and erosion that had been induced with LPS and K/BxN serum treatments (30% reversal of systemic bone loss [P < 0.01]; 55% reversal of erosion [P < 0.001]), without conferring bone-protective properties. CONCLUSION: Our results identify ATX as a novel OC factor that specifically controls inflammation-induced bone erosions and systemic bone loss. Therefore, ATX inhibition offers a novel therapeutic approach for potentially preventing bone erosion in patients with RA.

Osteoimmunology of Bone Loss in Inflammatory Rheumatic Diseases.

Over the past two decades, the field of osteoimmunology has emerged in response to a range of evidence demonstrating the reciprocal relationship between the immune system and bone. In particular, localized bone loss, in the form of joint erosions and periarticular osteopenia, as well as systemic osteoporosis, caused by inflammatory rheumatic diseases including rheumatoid arthritis, the prototype of inflammatory arthritis has highlighted the importance of this interplay. Osteoclast-mediated resorption at the interface between synovium and bone is responsible for the joint erosion seen in patients suffering from inflammatory arthritis. Clinical studies have helped to validate the impact of several pathways on osteoclast formation and activity. Essentially, the expression of pro-inflammatory cytokines as well as Receptor Activator of Nuclear factor κB Ligand (RANKL) is, both directly and indirectly, increased by T cells, stimulating osteoclastogenesis and resorption through a crucial regulator of immunity, the Nuclear factor of activated T-cells, cytoplasmic 1 (NFATc1). Furthermore, in rheumatoid arthritis, autoantibodies, which are accurate predictors both of the disease and associated structural damage, have been shown to stimulate the differentiation of osteoclasts, resulting in localized bone resorption. It is now also evident that osteoblast-mediated bone formation is impaired by inflammation both in joints and the skeleton in rheumatoid arthritis. This review summarizes the substantial progress that has been made in understanding the pathophysiology of bone loss in inflammatory rheumatic disease and highlights therapeutic targets potentially important for the cure or at least an alleviation of this destructive process.

Autotaxin Implication in Cancer Metastasis and Autoimunne Disorders: Functional Implication of Binding Autotaxin to the Cell Surface.

Autotaxin (ATX) is an exoenzyme which, due to its unique lysophospholipase D activity, is responsible for the synthesis of lysophosphatidic acid (LPA). ATX activity is responsible for the concentration of LPA in the blood. ATX expression is increased in various types of cancers, including breast cancer, where it promotes metastasis. The expression of ATX is also remarkably increased under inflammatory conditions, particularly in the osteoarticular compartment, where it controls bone erosion. Biological actions of ATX are mediated by LPA. However, the phosphate head group of LPA is highly sensitive to degradation by the action of lipid phosphate phosphatases, resulting in LPA inactivation. This suggests that for efficient action, LPA requires protection, which is potentially achieved through docking to a carrier protein. Interestingly, recent reports suggest that ATX might act as a docking molecule for LPA and also support the concept that binding of ATX to the cell surface through its interaction with adhesive molecules (integrins, heparan sulfate proteoglycans) could facilitate a rapid route of delivering active LPA to its cell surface receptors. This new mechanism offers a new vision of how ATX/LPA works in cancer metastasis and inflammatory bone diseases, paving the way for new therapeutic developments.

Autotaxin-ß interaction with the cell surface via syndecan-4 impacts on cancer cell proliferation and metastasis.

Autotaxin (ATX) promotes cancer cell metastasis through the production of lysophosphatidic acid (LPA). ATX binds to αvß3 integrins controlling metastasis of breast cancer cells. We screened a series of cancer cell lines derived from diverse human and mouse solid tumors for the capacity of binding to ATX and found only a modest correlation with their level of αvß3 integrin expression. These results strongly suggested the existence of another cell surface ATX-interacting factor. Indeed, ATXα has been shown to bind heparan-sulfate chains because of its unique polybasic insertion sequence, although the biological significance is unknown. We demonstrated here, that among all cell surface heparan-sulfate proteoglycans, syndecan-4 (SDC4) was essential for cancer cell interaction with ATXß but was restrained by heparan-sulfate chains. In addition, exogenous ATXß-induced MG63 osteosarcoma cell proliferation required physical interaction of ATXß with the cell surface via an SDC4-dependent mechanism. In a preclininal mouse model, targeting SDC4 on 4T1 mouse breast cancer cells inhibited early bone metastasis formation. Furthermore, SDC4-prometastatic activity was totally abolished in absence of ATX expression. In conclusion our results determined that ATX and SDC4 are engaged in a reciprocal collaboration for cancer cell metastasis providing the rational for the development of novel anti-metastasis therapies.

BMP9, but not BMP10, acts as a quiescence factor on tumor growth, vessel normalization and metastasis in a mouse model of breast cancer.

BACKGROUND: Angiogenesis has become an attractive target for cancer therapy. However, despite the initial success of anti-VEGF (Vascular endothelial growth factor) therapies, the overall survival appears only modestly improved and resistance to therapy often develops. Other anti-angiogenic targets are thus urgently needed. The predominant expression of the type I BMP (bone morphogenetic protein) receptor ALK1 (activin receptor-like kinase 1) in endothelial cells makes it an attractive target, and phase I/II trials are currently being conducted. ALK1 binds with strong affinity to two ligands that belong to the TGF-ß family, BMP9 and BMP10. In the present work, we addressed their specific roles in tumor angiogenesis, cancer development and metastasis in a mammary cancer model. METHODS: For this, we used knockout (KO) mice for BMP9 (constitutive Gdf2-deficient), for BMP10 (inducible Bmp10-deficient) and double KO mice (Gdf2 and Bmp10) in a syngeneic immunocompetent orthotopic mouse model of spontaneous metastatic breast cancer (E0771). RESULTS: Our studies demonstrate a specific role for BMP9 in the E0771 mammary carcinoma model. Gdf2 deletion increased tumor growth while inhibiting vessel maturation and tumor perfusion. Gdf2 deletion also increased the number and the mean size of lung metastases. On the other hand, Bmp10 deletion did not significantly affect the E0771 mammary model and the double deletion (Gdf2 and Bmp10) did not lead to a stronger phenotype than the single Gdf2 deletion. CONCLUSIONS: Altogether, our data show that in a tumor environment BMP9 and BMP10 play different roles and thus blocking their shared receptor ALK1 is maybe not appropriate. Indeed, BMP9, but not BMP10, acts as a quiescence factor on tumor growth, lung metastasis and vessel normalization. Our results also support that activating rather than blocking the BMP9 pathway could be a new strategy for tumor vessel normalization in order to treat breast cancer.

Platelets, autotaxin and lysophosphatidic acid signalling: win-win factors for cancer metastasis.

Platelets play a crucial role in the survival of metastatic cells in the blood circulation. The interaction of tumour cells with platelets leads to the production of plethoric factors among which our review will focus on lysophosphatidic acid (LPA), because platelets are the highest producers of this bioactive lysophospholipid in the organism. LPA promotes platelet aggregation, and blocking platelet function decreases LPA signalling and leads to inhibition of breast cancer cell metastasis. Autotaxin (ATX), a lysophospholipase D responsible for the basal concentration of LPA in blood, was detected in platelet α-granules. Functionally, active ATX is eventually released following tumour cell-induced platelet aggregation, thereby promoting metastasis. Megakaryocytes do not express ATX but respond to LPA stimulation. Whether LPA-primed megakaryocytes contribute to the recently reported negative action of megakaryocytes on cancer metastasis is not yet known. However, an understanding of the ATX/LPA signalling pathways in platelets, cancer cells and megakaryocytes opens up new approaches for fighting cancer metastasis.

Bone disease in nephropathic cystinosis is related to cystinosin-induced osteoclastic dysfunction.

Background: Bone impairment is a poorly described complication of nephropathic cystinosis (NC). The objectives of this study were to evaluate in vitro effects of cystinosin (CTNS) mutations on bone resorption and of cysteamine treatment on bone cells [namely human osteoclasts (OCs) and murine osteoblasts]. Methods: Human OCs were differentiated from peripheral blood mononuclear cells (PBMCs) of patients and healthy donors (HDs). Cells were treated with increasing doses of cysteamine in PBMCs or on mature OCs to evaluate its impact on differentiation and resorption, respectively. Similarly, cysteamine-treated osteoblasts derived from murine mesenchymal stem cells were assessed for differentiation and activity with toxicity and proliferation assays. Results: CTNS was expressed in human OCs derived from HDs; its expression was regulated during monocyte colony-stimulating factor- and receptor activator of nuclear factor-κB-dependent osteoclastogenesis and required for efficient bone resorption. Cysteamine had no impact on osteoclastogenesis but inhibited in vitro HD osteoclastic resorption; however, NC OC-mediated bone resorption was impaired only at high doses. Only low concentrations of cysteamine (50 µM) stimulated osteoblastic differentiation and maturation, while this effect was no longer observed at higher concentrations (200 µM). Conclusion: CTNS is required for proper osteoclastic activity. In vitro low doses of cysteamine have beneficial antiresorptive effects on healthy human-derived OCs and may partly correct the CTNS-induced osteoclastic dysfunction in patients with NC. Moreover, in vitro low doses of cysteamine also stimulate osteoblastic differentiation and mineralization, with an inhibitory effect at higher doses, likely explaining, at least partly, the bone toxicity observed in patients receiving high doses of cysteamine.