Knowledge, attitudes, and preventative practices regarding ischemic heart disease among emergency department patients in northern Tanzania.

OBJECTIVES: The objective of this study is to increase understanding of knowledge, attitudes, and preventative practices regarding ischemic heart disease (IHD) in sub-Saharan Africa in order to develop patient-centered interventions to improve care and outcomes. STUDY DESIGN: This is a prospective observational study. METHODS: Adult patients presenting with chest pain or shortness of breath to an emergency department in northern Tanzania were enrolled. A questionnaire was adapted from existing knowledge attitude and practice surveys regarding cardiovascular disease and the WHO STEPS instrument. Individual five-year risk of cardiovascular event was determined by validated models based on age, sex, systolic blood pressure, body mass index, diabetes, and smoking status. An IHD knowledge score was calculated by giving one point for each correct response to the knowledge-related items, with a maximum score of 10. Associations between IHD knowledge and patient characteristics were assessed by Welch's t-test. RESULTS: A total of 349 patients were enrolled, with median interquartile range (IQR) age 60 (45, 72) years. Of participants, 259 (74.2%) had hypertension, and 228 (65.3%) had greater than 10% five-year risk of cardiovascular event. The mean (SD) knowledge score was 4.8 (3.3). The majority of respondents (224, 64.2%) recognized obesity as a risk factor for heart attack, while a minority (34, 9.7%) knew that a daily aspirin could reduce the risk of cardiovascular event. Greater IHD knowledge was associated with younger age (P = 0.045) and higher levels of education (P < 0.001) but not higher risk of cardiovascular disease (P = 0.123). Most respondents expressed a willingness to diet to improve their health (322, 92.3%) and a preference for treatment from a physician rather than a traditional healer for a heart attack (321, 92.0%). A minority of patients reported exercising regularly (88, 25.2%) or seeing a doctor routinely for checkups (100, 28.7%). CONCLUSIONS: High-risk emergency department patients in northern Tanzania have moderate knowledge regarding IHD but do not consistently engage in healthy preventive practices. Patient-centered interventions are needed to improve IHD knowledge and practices in high-risk populations.

Access to emergency care services: a transversal ecological study about Brazilian emergency health care network.

OBJECTIVES: Studies of health geography are important in the planning and allocation of emergency health services. The geographical distribution of health facilities is an important factor in timely and quality access to emergency services; therefore, the present study analyzed the emergency health care network in Brazil, focusing the analysis at the roles of small hospitals (SHs). STUDY DESIGN: Cross-sectional ecological study. METHODS: Data were collected from 9429 hospitals of which 3524 were SHs and 5905 were high-complexity centers (HCCs). For analytical purposes, we considered four specialties when examining the proxies of emergency care capability: adult, pediatrics, neonatal, and obstetric. We analyzed the spatial distribution of hospitals, identifying municipalities that rely exclusively on SHs and the distance of these cities from HCCs. RESULTS: More than 14 and 30 million people were at least 120 km away from HCCs with an adult intensive care unit (ICU) and pediatric ICU, respectively. For neonatal care distribution, 12% of the population was more than 120 km away from a health facility with a neonatal ICU. The maternities situation is different from other specialties, where 81% of the total Brazilian population was within 1 h or less from such health facilities. CONCLUSION: Our results highlighted a polarization in distribution of Brazilian health care facilities. There is a concentration of hospitals in urban areas more developed and access gaps in rural areas and the Amazon region. Our results demonstrate that the distribution of emergency services in Brazil is not facilitating access to the population due to geographical barriers associated with great distances.

Vascular-targeted agents for the treatment of angiosarcoma.

PURPOSE: Angiosarcomas are rare, aggressive vascular tumours known to express vascular endothelial growth factor (VEGF), a key pro-angiogenic growth factor. The aim of this study was to determine the potential effects of vascular-targeted agents for the treatment of angiosarcoma, using two human cutaneous angiosarcoma cell lines (ASM and ISO-HAS), and human dermal microvascular endothelial cells (HuDMECs) for comparison. METHODS: Protein arrays were used to assess the expression of angiogenesis-related proteins, and potential drug targets were assessed by ELISA and Western blotting. Response to vascular-targeted agents, including bevacizumab an anti-VEGF antibody, axitinib a VEGF-receptor tyrosine kinase inhibitor, everolimus an mTOR inhibitor, selumetinib a MEK inhibitor and vadimezan a vascular-disrupting agent were compared in functional in vitro cellular assays, including viability, differentiation and migration assays. RESULTS: ASM and ISO-HAS cells expressed a broad range of pro-angiogenic growth factors. ASM and ISO-HAS VEGF expression was significantly increased (p = 0.029) compared with HuDMECs. Striking responses were seen to vadimezan with an IC50 of 90 and 150 µg/ml for ASM and ISO-HAS cells, respectively. Selumetinib inhibited ASM with an IC50 of 1,750 ng/ml, but was not effective in ISO-HAS. Everolimus reduced both ASM and ISO-HAS viable cell counts by 20 % (p < 0.001). Minimal responses were observed to bevacizumab and axitinib in assays with ASM and ISO-HAS cells. CONCLUSIONS: Further studies are warranted to investigate mTOR inhibitors, MEK inhibitors and vascular-disrupting agents for the treatment of angiosarcoma.

Anti-tissue factor short hairpin RNA inhibits breast cancer growth in vivo.

In breast cancer, there is a correlation between tissue factor (TF) expression, angiogenesis and disease progression. TF stimulates tumour angiogenesis, in part, through up-regulation of vascular endothelial growth factor (VEGF). Therefore, this study aimed to establish whether TF stimulates angiogenesis and tumour progression directly and independent of VEGF up-regulation. Initially, the effects of TF and VEGF were assessed on endothelial cell migration (Boyden chamber) and differentiation (tubule formation on Matrigel). Subsequently, MDA-MB-436 breast cancer cells, which produce high levels of both TF and VEGF (western blot analysis), were established in vivo, following which tumours were treated three times per week for 3 weeks with intra-tumoural injections of either anti-VEGF siRNA, anti-TF shRNA, the two treatments combined, or relevant controls. Both VEGF and TF significantly stimulated endothelial cell migration and tubule formation (P < 0.02). Breast cancer xenografts (MDA-MB-436) treated with TF or VEGF-specific agents demonstrated significant inhibition in tumour growth (VEGFsiRNA 61%; final volume: 236.2 ± 23.2 mm(3) vs TFshRNA 89%; 161.9 ± 17.4 mm(3) vs combination 93%; 136.3 ± 9.2 mm(3) vs control 400.4 ± 32.7 mm(3); P < 0.005). Microvessel density (MVD), a measure of angiogenesis, was also significantly inhibited in all groups (MVD in control = 29 ± 2.9; TFshRNA = 18 ± 1.1; VEGFsiRNA = 16.7 ± 1.5; both = 12 ± 2.1; P < 0.004), whereas the proliferative index of the tumours was only reduced in the TFshRNA-treated groups (control = 0.51 ± 0.011; TFshRNA = 0.41 ± 0.014; VEGFsiRNA = 0.49 ± 0.013; both = 0.41 ± 0.004; P < 0.008). These data suggest that TF has a direct effect on primary breast cancer growth and angiogenesis, and that specific inhibition of the TF-signalling pathway has potential for the treatment of primary breast cancer.

Angiopoietin-1, angiopoietin-2 and Tie-2 receptor expression in human dermal wound repair and scarring.

BACKGROUND: The angiopoietin (Ang)/Tie-2 ligand/receptor system is known to interact with the vascular endothelial growth factor (VEGF) pathway to determine the fate of blood vessels during angiogenesis. However, the precise contribution of this system to angiogenesis and the mechanisms of vascular maturation and remodelling in human tissue repair have yet to be elucidated. OBJECTIVES: To examine the spatial and temporal expression of Ang-1, Ang-2, Tie-2 and VEGF in relation to angiogenesis in human surgical wounds. METHODS: Punch biopsies were taken either from normal unwounded skin (controls) during surgery or from mastectomy scars between 3 days and 2 years postsurgery. Ang-1, Ang-2, Tie-2 and VEGF fibroblast/myofibroblast and endothelial expression were characterized by immunohistochemistry, analysed semiquantitatively and correlated with microvessel density (MVD) and scar age. RESULTS: The expression of VEGF, Ang-1, Ang-2 and Tie-2 in fibroblasts/myofibroblasts was increased significantly in early scars, decreased in older scars and was related to scar age (P < 0·001) and MVD (P < 0·0004), with strong correlations between all factors. In contrast, vascular expression of Ang-1 was decreased slightly in early scars, vascular Ang-2 remained constant and Tie-2 vascular expression increased, although there were no correlations with scar age or MVD. CONCLUSIONS: These data demonstrate that angiopoietins and their receptor, Tie-2, are expressed in both fibroblasts/myofibroblasts and endothelial cells in healing human wounds. Fibroblast/myofibroblast expression correlates with angiogenesis and VEGF expression, suggesting a role for the angiopoietin/Tie-2 system in normal wound repair and scarring.

A novel fragment derived from the beta chain of human fibrinogen, beta43-63, is a potent inhibitor of activated endothelial cells in vitro and in vivo.

BACKGROUND: Angiogenesis and haemostasis are closely linked within tumours with many haemostatic proteins regulating tumour angiogenesis. Indeed we previously identified a fragment of human fibrinogen, fibrinogen E-fragment (FgnE) with potent anti-angiogenic properties in vitro and cytotoxic effects on tumour vessels in vivo. We therefore investigated which region of FgnE was mediating vessel cytotoxicity. METHODS: Human dermal microvascular endothelial cells (ECs) were used to test the efficacy of peptides derived from FgnE on proliferation, migration, differentiation, apoptosis and adhesion before testing the efficacy of an active peptide on tumour vasculature in vivo. RESULTS: We identified a 20-amino-acid peptide derived from the beta chain of FgnE, beta43-63, which had no effect on EC proliferation or migration but markedly inhibited the ability of activated ECs to form tubules or to adhere to various constituents of the extracellular matrix - collagen IV, fibronectin and vitronectin. Furthermore, our data show that beta43-63 interacts with ECs, in part, by binding to alpha(v)beta(3), so soluble alpha(v)beta(3) abrogated beta43-63 inhibition of tubule formation by activated ECs. Finally, when injected into mice bearing tumour xenografts, beta43-63 inhibited tumour vascularisation and induced formation of significant tumour necrosis. CONCLUSIONS: Taken together, these data suggest that beta43-63 is a novel anti-tumour peptide whose anti-angiogenic effects are mediated by alpha(v)beta(3).

Angiogenesis, vascular endothelial growth factor and its receptors in human surgical wounds.

BACKGROUND: Angiogenesis plays an essential role in tissue repair. Vascular endothelial growth factor (VEGF) mediates angiogenesis through receptor kinases VEGF-R1 and VEGF-R2, and co-receptors, neuropilins Np1 and Np2. This study examined the spatial and temporal expression of these factors in relation to angiogenesis in surgical wounds. METHODS: Scar biopsies were obtained from patients between 3 days and 2 years after surgery. Normal skin control biopsies were taken during surgery. Microvessel density (MVD) was quantified using a Chalkley grid. VEGF, VEGF-R1, VEGF-R2, Np1 and Np2 endothelial expression was determined by immunohistochemistry, and correlated with MVD and scar age. RESULTS: Cumulative MVD was significantly greater in scars than controls (P = 0.011), and was related to scar age (P = 0.007). Expression of VEGF, VEGF-R2, Np1 and Np2 was increased significantly in all scars and correlated with MVD. In contrast, scar VEGF-R1 expression was decreased, and correlated with increased VEGF and VEGF-R2. CONCLUSION: Levels of VEGF, VEGF-R2, Np1 and Np2 are increased, whereas VEGF-R1 expression is decreased in angiogenesis, suggesting a role for VEGF-receptor complexes in early wound healing. This altered protein expression and increased presence of vessels is prolonged, suggesting that structural remodelling continues for at least 2 years after surgery.

Angiogenesis is associated with the onset of hyperplasia in human ductal breast disease.

BACKGROUND: The precise timing of the angiogenic switch and the role of angiogenesis in the development of breast malignancy is currently unknown. METHODS: Therefore, the expression of CD31 (pan endothelial cells (ECs)), endoglin (actively proliferating ECs), hypoxia-inducible factor-1 (HIF-1alpha), vascular endothelial growth factor-A (VEGF) and tissue factor (TF) were quantified in 140 surgical specimens comprising normal human breast, benign and pre-malignant hyperplastic tissue, in situ and invasive breast cancer specimens. RESULTS: Significant increases in angiogenesis (microvessel density) were observed between normal and benign hyperplastic breast tissue (P<0.005), and between in situ and invasive carcinomas (P<0.0005). In addition, significant increases in proliferating ECs were observed in benign hyperplastic breast compared with normal breast (P<0.05) cancers and in invasive compared with in situ cancers (P<0.005). Hypoxia-inducible factor-1alpha, VEGF and TF expression were significantly associated with increases in both angiogenesis and proliferating ECs (P<0.05). Moreover, HIF-1alpha was expressed by 60-75% of the hyperplastic lesions, and a significant association was observed between VEGF and TF in ECs (P<0.005) and invasive tumour cells (P<0.01). CONCLUSIONS: These findings are the first to suggest that the angiogenic switch, associated with increases in HIF-1alpha, VEGF and TF expression, occurs at the onset of hyperplasia in the mammary duct, although the greatest increase in angiogenesis occurs with the development of invasion.

Angiogenesis in pre-malignant conditions.

Evidence from human studies suggests that angiogenesis commences during the pre-malignant stages of cancer. Inhibiting angiogenesis may, therefore, be of potential value in preventing progression to invasive cancer. Understanding the mechanisms inducing angiogenesis in these lesions and identification of those important in human tumourigenesis are necessary to develop translational strategies that will help realise the goal of angioprevention.

Neuropilins in physiological and pathological angiogenesis.

Neuropilin-1 (Np1) and neuropilin-2 (Np2) are transmembrane glycoproteins with large extracellular domains that interact with both class 3 semaphorins and vascular endothelial growth factor (VEGF), and are involved in the regulation of many physiological pathways, including angiogenesis. The neuropilins also interact directly with the classical receptors for VEGF, VEGF-R1 and -R2, mediating signal transduction. The heart, glomeruli and osteoblasts express both Np1 and Np2, but there is differential expression in the adult vasculature, with Np1 expressed mainly by arterial endothelium, whereas Np2 is only expressed by venous and lymphatic endothelium. Both neuropilins are commonly over-expressed in regions of physiological (wound-healing) and pathological (tumour) angiogenesis, but the signal transduction pathways, neuropilin-mediated gene expression and the definitive role of neuropilins in angiogenic processes are not fully characterized. This review details the current evidence for the role of neuropilins in angiogenesis, and suggests future research directions that may enhance our understanding of the mechanisms of action of this unique family of proteins.

Identification of key residues involved in mediating the in vivo anti-tumor/anti-endothelial activity of Alphastatin.

BACKGROUND: We have recently shown that Alphastatin, a 24-amino-acid peptide (ADSGEGDFLAEGGGVRGPRVVERH) derived from human fibrinogen has anti-endothelial properties in vitro and in vivo. OBJECTIVES: The aim of this study was to determine the activity of a terminally modified (stabilized) form of Alphastatin in vitro and in vivo and to identify the key residues required for this activity. METHODS: The in vitro activity of modified Alphastatin, truncates and mutants was determined by endothelial cell (HuDMEC) tubule formation and migration. Active peptides were then assessed in vivo using syngeneic murine subcutaneous 4T1 mammary carcinomas. RESULTS: Modified Alphastatin-inhibited HuDMEC migration and tubule formation in response to multiple growth factors and caused a 45% inhibition in tumor growth when administered intravenously at 0.25 mg kg(-1) (three times per week). Intravenous (i.v.) administration proved non-toxic at all doses investigated, whereas oral and intraperitoneal (i.p.) administration demonstrated neither anti-tumor activity nor toxicity. Truncations of Alphastatin revealed an 11-amino-acid peptide (DFLAEGGGVRG), termed AHN419, which inhibited endothelial cell activity in vitro; however, intravenous AHN419 caused a non-significant growth inhibition in vivo. Single amino acid substitutions to alanine along the entire length of Alphastatin indicated that additional residues outside the AHN419 sequence were required for full activity. CONCLUSIONS: Terminal modification of Alphastatin altered the in vivo efficacy and these studies suggest that a hydrophobic cluster (Phe8, Leu9, Ala10 and Val15) is essential for the biological activity, but additional residues, including Ser3-Gly14, Pro18-Val20 and Arg23 are required for full inhibitory activity of Alphastatin.

Fibrinogen E fragment selectively disrupts the vasculature and inhibits the growth of tumours in a syngeneic murine model.

We recently demonstrated that a fragment of human fibrinogen, fibrinogen E fragment, inhibits the migration and differentiation of human endothelial cells in vitro. Here we show that it exerts similar effects on murine endothelial cells in vitro, and selectively disrupts tumour endothelium in vivo, causing widespread intravascular thrombosis and retarding the growth of CT26 tumours in a syngeneic murine model.