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Highlights • Bone metastases negatively impact on patients’ quality of life (QoL). • Skeletal related events have a detrimental effect on both QoL and survival. • Both local and systemic treatments are often required to manage bone metastases. • Bone turnover modulators reduce the risk of skeletal complications and improve pain. • Novel agents may deserve further investigation for the management of bone metastases., Bone metastases (BM) are a common complication of cancer, whose management often requires a multidisciplinary approach. Despite the recent therapeutic advances, patients with BM may still experience skeletal-related events and symptomatic skeletal events, with detrimental impact on quality of life and survival. A deeper knowledge of the mechanisms underlying the onset of lytic and sclerotic BM has been acquired in the last decades, leading to the development of bone-targeting agents (BTA), mainly represented by anti-resorptive drugs and bone-seeking radiopharmaceuticals. Recent pre-clinical and clinical studies have showed promising effects of novel agents, whose safety and efficacy need to be confirmed by prospective clinical trials. Among BTA, adjuvant bisphosphonates have also been shown to reduce the risk of BM in selected breast cancer patients, but failed to reduce the incidence of BM from lung and prostate cancer. Moreover, adjuvant denosumab did not improve BM free survival in patients with breast cancer, suggesting the need for further investigation to clarify BTA role in early-stage malignancies. The aim of this review is to describe BM pathogenesis and current treatment options in different clinical settings, as well as to explore the mechanism of action of novel potential therapeutic agents for which further investigation is needed.

Atherosclerosis is a multifocal alteration of the vascular wall of medium and large arteries characterized by a local accumulation of cholesterol and non-resolving inflammation. Atherothrombotic complications are the leading cause of disability and mortality in western countries. Neovascularization in atherosclerotic lesions plays a major role in plaque growth and instability. The angiogenic process is mediated by classical angiogenic factors and by additional factors specific to atherosclerotic angiogenesis. In addition to its role in plaque progression, neovascularization may take part in plaque destabilization and thromboembolic events. Anti-angiogenic agents are effective to reduce atherosclerosis progression in various animal models. However, clinical trials with anti-angiogenic drugs, mainly anti-VEGF/VEGFR, used in anti-cancer therapy show cardiovascular adverse effects, and require additional investigations., Graphical abstract fx1, Highlights • Intimal neovascularization in plaque growth and destabilization. • Role of classical and specific angiogenic factors in atherosclerotic angiogenesis. • Inflammation and oxidative stress promote angiogenesis and plaque destabilization. • Angiogenesis promote intraplaque hemorrhages, afflux of blood cells and lipids. • Anti-angiogenic therapy for atherosclerotic angiogenesis.

sFlt-1 (soluble Flt-1) potently inhibits angiogenesis by binding extracellularly to VEGF (vascular endothelial growth factor). In the present paper, we report that hypoxia down-regulates sFlt-1 expression in HMVECs (human microvascular endothelial cells), a constituent of microvessels where angiogenesis occurs. Hypoxia (5–1% O2) increased VEGF expression in HMVECs. In contrast, the levels of sFlt-1 mRNA and protein in HMVECs decreased significantly as the O2 concentration fell, whereas mFlt-1 (membrane-bound Flt-1) mRNA and protein remained unchanged. This suggested that hypoxia selectively regulates alternative 3′-end processing of sFlt-1 pre-mRNA. We have also demonstrated that sFlt-1 overexpression in lentiviral-construct-infected HMVECs counteracted VEGF-induced endothelial cell growth. We next identified cis-elements involved in sFlt-1 mRNA processing in HMVECs using a human Flt-1 minigene and found that two non-contiguous AUUAAA sequences function as the poly(A) signal. Furthermore, we identified a cis-element in intron 13 that regulates sFlt-1 mRNA processing. Mutagenesis of the U-rich region in intron 13 caused a significant decrease in the soluble-form/membrane-form RNA ratio in the minigene-transfected HMVECs. These results suggest that decreased sFlt-1 expression due to hypoxia contributes to hypoxia-induced angiogenesis and reveals a novel mechanism regulating angiogenesis by alternative mRNA 3′-end processing.

Using human MSCs (mesenchymal stem cells) lacking VEGF (vascular endothelial growth factor) receptors, we show that the pro-angiogenic receptor neuropilin-1 associates with phosphorylated PDGFRs [PDGF (platelet-derived growth factor) receptors], thereby regulating cell signalling, migration, proliferation and network assembly. Neuropilin-1 co-immunoprecipitated and co-localized with phosphorylated PDGFRs in the presence of growth factors. Neuropilin-1 knockdown blocked PDGF-AA-induced PDGFRα phosphorylation and migration, reduced PDGF-BB-induced PDGFRβ activation and migration, blocked VEGF-A activation of both PDGFRs, and attenuated proliferation. Neuropilin-1 prominently co-localized with both PDGFRs within MSC networks assembled in Matrigel™ and in the chorioallantoic membrane vasculature microenvironment, and its knockdown grossly disrupted network assembly and decreased PDGFR signalling. Thus neuropilin-1 regulates MSCs by forming ligand-specific receptor complexes that direct PDGFR signalling, especially the PDGFRα homodimer. This receptor cross-talk may control the mobilization of MSCs in neovascularization and tissue remodelling.

The PRH (proline-rich homeodomain) [also known as Hex (haematopoietically expressed homeobox)] protein is a critical regulator of vertebrate development. PRH is able to regulate cell proliferation and differentiation and is required for the formation of the vertebrate body axis, the haematopoietic and vascular systems and the formation of many vital organs. PRH is a DNA-binding protein that can repress and activate the transcription of its target genes using multiple mechanisms. In addition, PRH can regulate the nuclear transport of specific mRNAs making PRH a member of a select group of proteins that control gene expression at the transcriptional and translational levels. Recent biophysical analysis of the PRH protein has shown that it forms homo-oligomeric complexes in vivo and in vitro and that the proline-rich region of PRH forms a novel dimerization interface. Here we will review the current literature on PRH and discuss the complex web of interactions centred on this multifunctional protein.

STS (soft tissue sarcomas) are rare malignant tumours deriving from cells of mesenchymal origin and represent only 1% of all malignant neoplasms. It has been extensively demonstrated that angiogenesis has an important role in cancer malignancy. Particularly, a lot of studies demonstrate the importance of angiogenesis in the development of carcinomas, whereas little is known about the role of angiogenesis in sarcomas and especially in STS. This review aims at summarizing the new discoveries about the nature and the importance of angiogenesis in STS and the new possible therapeutic strategies involved. Only a few studies concerning STS focus on tumour neovascularization and proangiogenic factors and look for a correlation with the patients prognosis/survival. These studies demonstrate that intratumoural MVD (microvessels density) may not accurately represent the angiogenic capacity of STS. Nevertheless, this does not exclude the possibility that angiogenesis could be important in STS. The importance of neoangiogenesis in soft tissue tumours is confirmed by the arising number of publications comparing angiogenesis mediators with clinical features of patients with STS. The efficacy of anti-angiogenic therapies in other types of cancer is well documented. The understanding of the involvement of the angiogenic process in STS, together with the necessity to improve the therapy for this often mortal condition, prompted the exploration of anti-tumour compounds targeting this pathway. In conclusion, this review emphasizes the importance to better understand the mechanisms of angiogenesis in STS in order to subsequently design-specific target therapies for this group of poorly responding tumours., Angiogenesis is important for tumour growth and metastatization. The better understanding of the mechanisms of angiogenesis in soft tissue sarcomas can lead the design of specific target therapies for this group of poorly responding tumours.

Diabetic mice are characterized by a disrupted expression pattern of VEGF (vascular endothelial growth factor), and impaired vasculogenesis during healing. Experimental evidence suggests that RLX (relaxin) can improve several parameters associated with wound healing. Therefore we investigated the effects of porcine-derived RLX in diabetes-related wound-healing defects in genetically diabetic mice. An incisional wound model was produced on the back of female diabetic C57BL/KsJ-m+/+Leptdb (db+/db+) mice and their normal littermates (db+/+m). Animals were treated daily with porcine RLX (25 μg/mouse per day, subcutaneously) or its vehicle. Mice were killed on 3, 6 and 12 days after skin injury for measurements of VEGF mRNA and protein synthesis, SDF-1α (stromal cell-derived factor-1α) mRNA and eNOS (endothelial NO synthase) expression. Furthermore, we evaluated wound-breaking strength, histological changes, angiogenesis and vasculogenesis at day 12. Diabetic animals showed a reduced expression of VEGF, eNOS and SDF-1α compared with non-diabetic animals. At day 6, RLX administration resulted in an increase in VEGF mRNA expression and protein wound content, in eNOS expression and in SDF-1α mRNA. Furthermore, the histological evaluation indicated that RLX improved the impaired wound healing, enhanced the staining of MMP-11 (matrix metalloproteinase-11) and increased wound-breaking strength at day 12 in diabetic mice. Immunohistochemistry showed that RLX in diabetic animals augmented new vessel formation by stimulating both angiogenesis and vasculogenesis. RLX significantly reduced the time to complete skin normalization and this effect was abrogated by a concomitant treatment with antibodies against VEGF and CXCR4 (CXC chemokine receptor 4), the SDF-1α receptor. These data strongly suggest that RLX may have a potential application in diabetes-related wound disorders.

It has been proposed that either excessive inflammation or an imbalance in angiogenic factors cause pre-eclampsia. In the present review, the arguments for and against the role of inflammation and/or angiogenic imbalance as the cause of pre-eclampsia are discussed on the basis of the Bradford–Hill criteria for disease causation. Although both angiogenic imbalance and systemic inflammation are implicated in pre-eclampsia, the absence of temporality of inflammatory markers with pre-eclampsia challenges the concept that excessive inflammation is the cause of pre-eclampsia. In contrast, the elevation of anti-angiogenic factors that precede the clinical signs of pre-eclampsia fulfils the criterion of temporality. The second most important criterion is the dose–response relationship. Although such a relationship has not been proven between pro-inflammatory cytokines and pre-eclampsia, high levels of anti-angiogenic factors have been shown to correlate with increased incidence and disease severity, hence satisfying this condition. Finally, as the removal of circulating sFlt-1 (soluble Fms-like tyrosine kinase receptor-1) from pre-eclamptic patients significantly improves the clinical outcome, it fulfils the Hill's experiment principle, which states that removal of the cause by an appropriate experimental regimen should ameliorate the condition. In contrast, treatment with high doses of corticosteroid fails to improve maternal outcome in pre-eclampsia, despite suppressing inflammation. Inflammation may enhance the pathology induced by the imbalance in the angiogenic factors, but does not by itself cause pre-eclampsia. Development of therapies based on the angiogenic and cytoprotective mechanisms seems more promising.

Pre-eclampsia is a pregnancy-related condition characterized by hypertension, proteinuria and endothelial dysfunction. VEGF(165)b, formed by alternative splicing of VEGF (vascular endothelial growth factor) pre-mRNA, inhibits VEGF(165)-mediated vasodilation and angiogenesis, but has not been quantified in pregnancy. ELISAs were used to measure means+/-S.E.M. plasma VEGF(165)b, sEng (soluble endoglin) and sFlt-1 (soluble fms-like tyrosine kinase-1). At 12 weeks of gestation, the plasma VEGF(165)b concentration was significantly up-regulated in plasma from women who maintained normal blood pressure throughout their pregnancy (normotensive group, 4.90+/-1.6 ng/ml; P0.01, as determined using a Mann-Whitney U test) compared with non-pregnant women (0.40+/-0.22 ng/ml). In contrast, in patients who later developed pre-eclampsia, VEGF(165)b levels were lower than in the normotensive group (0.467+/-0.209 ng/ml), but were no greater than non-pregnant women. At term, plasma VEGF(165)b concentrations were greater than normal in both pre-eclamptic (3.75+/-2.24 ng/ml) and normotensive (10.58 ng/ml+/-3.74 ng/ml; P0.1 compared with pre-eclampsia) pregnancies. Patients with a lower than median plasma VEGF(165)b at 12 weeks had elevated sFlt-1 and sEng pre-delivery. Concentrations of sFlt-1 (1.20+/-0.07 and 1.27+/-0.18 ng/ml) and sEng (4.4+/-0.18 and 4.1+/-0.5 ng/ml) were similar at 12 weeks of gestation in the normotensive and pre-eclamptic groups respectively. Plasma VEGF(165)b levels were elevated in pregnancy, but this increase is delayed in women that subsequently develop pre-eclampsia. In conclusion, low VEGF(165)b may therefore be a clinically useful first trimester plasma marker for increased risk of pre-eclampsia.