Angiogenic Modulators Research Articles

Angiopoietin 2 promotes angiogenesis in tissue-engineered bone and improves repair of bone defects by inducing autophagy.

Angiogenesis plays a key role in the repair of large segmental bone defects with tissue-engineered bones. However, there is no effective method of promoting angiogenesis in tissue-engineered bone. Both angiopoietin 2 (Ang2) and autophagy have been shown to be involved in angiogenesis, but their roles in angiogenesis of tissue-engineered bone remains unknown. In this in vivo study, a radius bone defect was created in New Zealand white rabbits, which were then treated by implantation of a hydroxyapatite/collagen scaffold followed by injection of different concentrations of Ang2. Expression of the autophagic modulators microtubule-associated protein 1 light chain 3 (LC3), Beclin-1, and SQSTM1/P62 were measured via western blotting, while the angiogenic modulators VEGF and CD31 were detected by western blotting and immunohistochemistry, respectively. X-ray imaging combined with general observation was used to evaluate bone defect healing. Expression of LC3 -I/LC3-II, Beclin-1, VEGF, and CD31 in the callus area increased and SQSTM1/p62 decreased in a dose-dependent manner with increasing Ang2 concentration. In the group treated with a high concentration of Ang2, the new callus grew well, accompanied by remarkable angiogenesis, leading to good repair of the bone defects. However, in the low concentration of Ang2 group, in spite of the existence of angiogenesis and new bone formation, the bone defects were not repaired. Furthermore, angiogenesis and osteogenesis were both obstructed in the control group. In conclusion, our study demonstrated that a high concentration of Ang2 promoted angiogenesis in tissue-engineered bone and improved repair of bone defects by inducing autophagy.

Placental growth factor reverses decreased vascular and uteroplacental MMP-2 and MMP-9 and increased MMP-1 and MMP-7 and collagen types I and IV in hypertensive pregnancy.

Preeclampsia is a complication of pregnancy manifested as maternal hypertension (HTN) and fetal intrauterine growth restriction, with unclear mechanisms. Placental ischemia increases antiangiogenic soluble fms-like tyrosine kinase-1 (sFlt-1) relative to angiogenic placental growth factor (PlGF); however, the molecular targets are unclear. To test the hypothesis that placental ischemia-induced changes in sFlt-1 and PlGF target vascular and uteroplacental matrix metalloproteinases (MMPs), we tested whether raising the sFlt-1-to-PlGF ratio by infusing sFlt-1 (10 µg·kg-1·day-1) in pregnant (Preg) rats increases blood pressure (BP) and alters MMPs and whether correcting sFlt-1/PlGF by infusing PlGF (20 µg·kg-1·day-1) in Preg rats with reduced uterine perfusion pressure (RUPP) improves BP and reverses the changes in MMPs. On gestational day 19, BP was higher and the litter size and uterine, placenta, and pup weight were less in Preg + sFlt-1 and RUPP than Preg rats and restored in RUPP + PlGF versus RUPP rats. Gelatin and casein zymography and Western blots revealed decreases in MMP-2 and MMP-9 and increases in MMP-1 and MMP-7 in the aorta, uterine artery, uterus, and placenta of Preg + sFlt-1 and RUPP versus Preg rats, which were reversed in RUPP + PlGF versus RUPP rats. Collagen types I and IV were more abundant in Preg + sFlt-1 and RUPP versus Preg rats and were reversed in RUPP + PlGF versus RUPP rats. Thus, PlGF reverses decreased vascular and uteroplacental MMP-2 and MMP-9 and increased MMP-1, MMP-7, and collagen types I and IV induced by placental ischemia and sFlt-1 in HTN in pregnancy. Angiogenic factors and MMP modulators could rectify changes in MMPs and collagen, restore vascular and uteroplacental remodeling, and improve HTN and intrauterine growth restriction in preeclampsia. NEW & NOTEWORTHY Understanding the mechanisms of preeclampsia could help in its prevention and management. This study shows that correcting soluble fms-like tyrosine kinase-1 (sFlt-1)/placental growth factor (PlGF) imbalance by infusing PlGF reverses the decreases in vascular and uteroplacental matrix metalloproteinase (MMP)-2 and MMP-9 and the increases in MMP-1, MMP-7, and collagen types I and IV induced by placental ischemia and antiangiogenic sFlt-1 in hypertension in pregnancy. Angiogenic factors and MMP modulators could rectify changes in vascular and uteroplacental MMPs and collagen content and ameliorate hypertension and intrauterine growth restriction in preeclampsia.

Platelet Microparticles and miRNA Transfer in Cancer Progression: Many Targets, Modes of Action, and Effects Across Cancer Stages.

Platelet-derived microparticles (PMPs) have long been known to increase in circulation in the presence of cancer, and have been considered to be cancer promoting by multiple mechanisms including shrouding of circulating tumor cells allowing immune evasion, inducing a procoagulant state associated with increased risk for venous thromboembolic events in cancer patients, and supporting metastatic dissemination by establishment of niches for anchorage of circulating tumor cells. These modes of PMP-enhanced progression of late stage cancer are generally based on the adhesive and procoagulant surfaces of PMPs. However, it is now clear that PMPs can also act as intercellular signaling vesicles, by fusion with target cells and transfer of a broad array of platelet-derived molecular contents including growth factors, angiogenic modulators, second messengers, lipids, and nucleic acids. It is also now well established that PMPs are major repositories of microRNAs (miRNAs). In recent years, new roles of PMPs in cancer have begun emerging, primarily reflecting their ability to transfer miRNA contents and modulate gene expression in target cells, allowing PMPs to affect cancer development at many stages. PMPs have been shown to interact with and transfer miRNAs to various blood vascular cells including endothelium, macrophages and neutrophils. As each of these contributes to cancer progression, PMP-mediated miRNA transfer can affect immune response, NETosis, tumor angiogenesis, and likely other cancer-associated processes. Recently, PMP miRNA transfer was found to suppress primary tumor growth, via PMP infiltration in solid tumors, anchorage to tumor cells and direct miRNA transfer, resulting in tumor cell gene suppression and inhibition of tumor growth. This mini-review will summarize current knowledge of PMP-miRNA interactions with cancer-associated cells and effects in cancer progression, and will indicate new research directions for understanding platelet-cancer interactions.

The Role of Biomaterials as Angiogenic Modulators of Spinal Cord Injury: Mimetics of the Spinal Cord, Cell and Angiogenic Factor Delivery Agents.

Spinal cord injury (SCI) represents an extremely debilitating condition for which no efficacious treatment is available. One of the main contributors to the inhospitable environment found in SCI is the vascular disruption that happens at the moment of injury that compromises the blood-spinal cord barrier (BSCB) and triggers a cascade of events that includes infiltration of inflammatory cells, ischemia and intraparenchymal hemorrhage. Due to the unsatisfactory nature of revascularization following SCI, restoring vascular perfusion and the BSCB seems an interesting way of modulating the lesion environment into a regenerative phenotype, with a potential increase in functional recovery. Certain biomaterials possess interesting features to enhance SCI therapies, and in fact have been applied as angiogenic promoters in other pathologies. The present mini-review intends to highlight the contribution that biomaterials could make in the development of novel therapeutic solutions able to restore proper vascularization and the BSCB.

Advanced tube formation assay using human endothelial colony forming cells for in vitro evaluation of angiogenesis.

The tube formation assay is a widely used in vitro experiment model to evaluate angiogenic properties by measuring the formation of tubular structures from vascular endothelial cells (ECs). in vitro experimental results are crucial when considered the advisability of moving forward to in vivo studies. Thus, the additional attentions to the in vitro assay is necessary to improve the quality of the pre-clinical data, leading to better decision-making for successful drug discovery. In this study, we improved the tube formation assay system in three aspects. First, we used human endothelial colony forming cells (ECFCs), which are endothelial precursors that have a robust proliferative capacity and more defined angiogenic characteristics compared to mature ECs. Second, we utilized a real-time cell recorder to track the progression of tube formation for 48 hours. Third, to minimize analysis error due to the limited observation area, we used image-stitching software to increase the microscope field of view to a 2×2 stitched area from the 4× object lens. Our advanced tube formation assay system successfully demonstrated the time-dependent dynamic progression of tube formation in the presence and absence of VEGF and FGF-2. Vatalanib, VEGF inhibitor, was tested by our assay system. Of note, IC50 values of vatalanib was different at each observation time point. Collectively, these results indicate that our advanced tube formation assay system replicates the dynamic progression of tube formation in response to angiogenic modulators. Therefore, this new system provides a sensitive and versatile assay model for evaluating pro- or anti-angiogenic drugs.

Cgnl1, an endothelial junction complex protein, regulates GTPase mediated angiogenesis.

AimsThe formation of cell–cell and cell–extra cellular matrix (ECM) contacts by endothelial cells (ECs) is crucial for the stability and integrity of a vascular network. We previously identified cingulin-like 1 (Cgnl1) in a transcriptomic screen for new angiogenic modulators. Here we aim to study the function of the cell–cell junction associated protein Cgnl1 during vessel formation.Methods and resultsUnlike family member cingulin, Cgnl1 expression is enriched in ECs during vascular growth. Cgnl1 is important for the formation of multicellular tubule structures, as shown in vitro using loss-of function assays in a 3D matrix co-culture system that uses primary human ECs and supporting mural cells. Further studies revealed that Cgnl1 regulates vascular growth by promoting Ve-cadherin association with the actin cytoskeleton, thereby stabilizing adherens junctions. Cgnl1 also regulates focal adhesion assembly in response to ECM contact, promoting vinculin and paxillin recruitment and focal adhesion kinase signalling. In vivo, we demonstrate in a postnatal retinal vascular development model in mice that Cgnl1 function is crucial for sustaining neovascular growth and stability.ConclusionsOur data demonstrate a functional relevance for Cgnl1 as a defining factor in new vessel formation both in vitro and in vivo.

Angelica Dahurica ethanolic extract improves impaired wound healing by activating angiogenesis in diabetes.

Abnormal angiogenesis plays an important role in impaired wound healing and development of chronic wounds in diabetes mellitus. Angelica dahurica radix is a common traditional Chinese medicine with wide spectrum medicinal effects. In this study, we analyzed the potential roles of Angelica dahurica ethanolic extract (ADEE) in correcting impaired angiogenesis and delayed wound healing in diabetes by using streptozotocin-induced diabetic rats. ADEE treatment accelerated diabetic wound healing through inducing angiogenesis and granulation tissue formation. The angiogenic property of ADEE was subsequently verified ex vivo using aortic ring assays. Furthermore, we investigated the in vitro angiogenic activity of ADEE and its underlying mechanisms using human umbilical vein endothelial cells. ADEE treatment induced HUVECs proliferation, migration, and tube formation, which are typical phenomena of angiogenesis, in dose-dependent manners. These effects were associated with activation of angiogenic signal modulators, including extracellular signal-regulated kinase 1/2 (ERK1/2), Akt, endothelial nitric oxide synthase (eNOS) as well as increased NO production, and independent of affecting VEGF expression. ADEE-induced angiogenic events were inhibited by the MEK inhibitor PD98059, the PI3K inhibitor Wortmannin, and the eNOS inhibitor L-NAME. Our findings highlight an angiogenic role of ADEE and its ability to protect against impaired wound healing, which may be developed as a promising therapy for impaired angiogenesis and delayed wound healing in diabetes.

Local and Systemic Inflammatory Biomarkers of Diabetic Retinopathy: An Integrative Approach.

To review the usefulness of local and systemic inflammatory biomarkers of diabetic retinopathy (DR) to implement a more personalized treatment. An integrated research (from ophthalmologist and diabetologist point of view) of most significant literature on serum, vitreous, and aqueous humor (AH) biochemical biomarkers related to inflammation at early and advanced stages of DR (including diabetic macular edema [DME] and proliferative DR) was performed. Moreover, novel imaging retinal biomarkers of local "inflammatory condition" were described. Multiple inflammatory cytokines and chemokines are increased in DR in both serum as well as in the eye (vitreous and AH). Nevertheless, local rather than systemic production of proinflammatory cytokines seems more relevant in the pathogenesis of both DR and DME. In the eye, retinal glia cells (macroglia and microglia) together with RPE are major sources of proinflammatory and angiogenic modulators. Retinal imaging allows for noninvasive clinical evaluation of retinal inflammatory response induced by diabetes mellitus. Proinflammatory cytokines/chemokines play an essential role in the pathogenesis of DR. Therefore, circulating biomarkers and retinal imaging aimed at assessing inflammation have emerged as useful tools for monitoring the onset and progression of DR. In addition, "liquid biopsy" of AH seems a good option in patients with advanced stages of DR requiring intravitreous injections. This strategy may permit us to implement a more personalized treatment with better visual function outcome. Further evaluation and validation of circulating and local biomarkers, as well as multimodal imaging is needed to gain new insights into this issue.

Disruption of Angiogenesis by Anthocyanin-Rich Extracts of Hibiscus sabdariffa.

Abnormal vessel formations contribute to the progression of specific angiogenic diseases including age-related macular degeneration. Adequate vessel growth and maintenance represent the coordinated process of endothelial cell proliferation, matrix remodeling, and differentiation. However, the molecular mechanism of the proper balance between angiogenic activators and inhibitors remains elusive. In addition, quantitative analysis of vessel formation has been challenging due to complex angiogenic morphology. We hypothesized that conjugated double bond containing-natural products, including anthocyanin extracts from Hibiscus sabdariffa, may control the proper angiogenesis. The current study was designed to determine whether natural molecules from African plant library modulate angiogenesis. Further, we questioned how the proper balance of anti- or pro-angiogenic signaling can be obtained in the vascular microenvironment by treating anthocyanin or fatty acids using chick chorioallantoic membrane angiogenesis model in ovo. The angiogenic morphology was analyzed systematically by measuring twenty one angiogenic indexes using Angiogenic Analyzer software. Chick chorioallantoic model demonstrated that anthocyanin-rich extracts inhibited angiogenesis in time- and concentration-dependent manner. Molecular modeling analysis proposed that hibiscetin as a component in Hibiscus may bind to the active site of vascular endothelial growth factor receptor 2 (VEGFR2) with ΔG= -8.42 kcal/mol of binding energy. Our results provided the evidence that anthocyanin is an angiogenic modulator that can be used to treat uncontrolled neovascular-related diseases, including age-related macular degeneration.

Antiangiogenic, wound healing and antioxidant activity of Cladosporium cladosporioides (Endophytic Fungus) isolated from seaweed (Sargassum wightii)

ABSTRACTEndophytic fungi from marine seaweeds are the less studied group of organisms with vast medical applications. The aim of the present study was to evaluate antioxidant, antiangiogenic as well as wound healing potential of the endophytic fungus isolated from the seaweed Sargassum wightii. The morphological characters and the rDNA internal transcribed spacer sequence analysis (BLAST search in Gen Bank database) was used for the identification of endophytic fungus. The antioxidant potential of the ethyl acetate extract of endophytic fungus was assessed by, 1,1-diphenyl-2-picryl-hydrazyl radical scavenging method. The fungal extract was also analysed for reducing power, total phenolic and flavonoid content. Antiangiogenic activity of the fungal extract was studied in vitro by inhibition of wound healing scratch assay and in vivo by Chick chorioallantoic membrane assay. The endophytic fungus was identified as Cladosporium cladosporioides (Gen Bank ID – KT384175). The ethyl acetate extract of C. cladosporioides showed a significant antioxidant and angiosuppressive activity. The ESI-LC-MS analysis of the extract revealed the presence of wide range of secondary metabolites. Results suggest that C. cladosporioides extract could be exploited as a potential source for angiogenic modulators.

Angiogenic Rg1/Sr-Doped TiO2Nanowire/Poly(Propylene Fumarate) Bone Cement Composites

A new approach is provided for preparing radiopaque and angiogenic poly(propylene fumarate) (PPF) bone cements by integrating Sr-doped n-TiO2 nanowires and ginsenoside Rg1 suitable for treating osteonecrosis. High aspect ratio radiopaque TiO2 -nanowires are synthesized by strontium doping in supercritical CO2 for the first time, showing a new phase, SrTiO3 . PPF is synthesized using a transesterification method by reacting diethyl fumarate and propylene glycol, then functionalized using maleic anhydride to produce terminal carboxyl groups, which are subsequently linked to the nanowires. The strong interfacial adhesion between functionalized PPF and nanowires is examined by scanning electron microscopy, Fourier transform infrared, X-ray photoelectron spectroscopy, thermal analysis, and mechanical testing. An angiogenic modulator, ginsenoside Rg1 , is integrated into the bone cement formulation with the mechanical properties, radiopacity, drug release, and angiogenesis behavior of the formed composites explored. The results show superior radiopacity and excellent release of ginsenoside Rg1 in vitro, as well as a dose-dependent increase in the branching point numbers. The present study suggests this new methodology provides sufficient mechanical properties, radiopacity, and angiogenic activity to be suitable for cementation of necrotic bone.

Identification of RSK and TTK as Modulators of Blood Vessel Morphogenesis Using an Embryonic Stem Cell-Based Vascular Differentiation Assay

SummaryBlood vessels are formed through vasculogenesis, followed by remodeling of the endothelial network through angiogenesis. Many events that occur during embryonic vascular development are recapitulated during adult neoangiogenesis, which is critical to tumor growth and metastasis. Current antiangiogenic tumor therapies, based largely on targeting the vascular endothelial growth factor pathway, show limited clinical benefits, thus necessitating the discovery of alternative targets. Here we report the development of a robust embryonic stem cell-based vascular differentiation assay amenable to small-molecule screens to identify novel modulators of angiogenesis. In this context, RSK and TTK were identified as angiogenic modulators. Inhibition of these pathways inhibited angiogenesis in embryoid bodies and human umbilical vein endothelial cells. Furthermore, inhibition of RSK and TTK reduced tumor growth, vascular density, and improved survival in an in vivo Lewis lung carcinoma mouse model. Our study suggests that RSK and TTK are potential targets for antiangiogenic therapy, and provides an assay system for further pathway screens.

Role of Isoprenoid Compounds on Angiogenic Regulation: Opportunities and Challenges.

Isoprenoids represent one of the largest classes of phytochemicals. The structural diversity of these compounds, as well as their remarkable biological activities, makes them suitable candidates for the development of novel therapeutic agents. Several isoprenoids have demonstrated promising potential in the modulation of angiogenesis processes, and therefore provide an appealing alternative and/or addition to the available pharmacotherapies. These compounds could be used per se or combined with standard therapies, which can potentially reduce the undesired secondary effects. Compounds like the sesquiterpenoid artemisinin, and its derivatives, or the diterpenoid triptolide have been successfully tested in a broad range of models (in vitro and in vivo). Moreover, sesquiterpenoids seem to be a promising resource of natural angiogenic modulators, as it can be attested by the significant number of recent publications in this subject. On the other hand, other isoprenoids, such as the triterpenoid ursolic acid, are still under-explored and further studies are needed to understand their role within angiogenic process. Further insights into isoprenoids mode of action in angiogenesis will hopefully pave the way towards their successful clinical use.

Assessment of CD-105 as an Angiogenic Modulator in Odontogenic Myxomas and Dental Follicles.

Aim Odontogenic myxoma is a benign intraosseous neoplasm of the jaws, with a locally aggressive behavior and a high recurrence rate. CD-105 is a homodimeric cell membrane glycoprotein and is a component of the TGF-β1 growth factor receptor complex that modulates angiogenesis by regulating the proliferation, differentiation and cellular migration. The aim of this study is to quantify the microvascular density of the odontogenic myxoma based on the expression of CD-105. Materials and Methods The analysis included 18 odontogenic myxoma and 18 dental follicles as controls. A standard immunohistochemical procedure was performed with the CD-105 antibody. Five representative fields (40×) of the odontogenic myxoma and the dental follicles were selected to determine the microvascular density, which was then followed by a descriptive and comparative statistical analysis. Results Dental follicles presented a significantly higher microvascular density compared with odontogenic myxoma (P = .001). The odontogenic myxoma smaller than 3 cm showed a greater microvascular density than those larger than 3 cm in size (P > .05), and the microvascular density was lower in large odontogenic myxomas as compared with the dental follicles (P = .003). Conclusion A weaker expression of CD-105 in odontogenic myxoma might indicate a lower angiogenic activity, suggesting that vascular proliferation has a limited role in the growth mechanisms and in the aggressive behavior of this neoplasm.

Abstract B11: A dual PI3K/mTOR inhibitor, BEZ235 blocks tumor-induced angiogenesis: Evidence for an effect on the tumor and endothelial compartment

Abstract Background: Tumor progression including HER2+ breast tumor, particularly in aggressive and malignant tumors, is associated with the angiogenesis. To investigate the effects of BEZ235 on angiogenesis, we evaluated its effects on the hypoxic stabilization or the PI3K-AKT-mTOR pathway-mediated synthesis of hypoxia-inducible factor -1α (HIF-1α) and tumor-induced angiogenesis. Methodology: Here we have studied the in vitro and in vivo effects of BEZ235 in HER2+/T-sensitive (BT474), HER2+/T-resistant (BT474HerR) and HER2+/PIK3CA (HCC1954 & UACC893) mutated models. We assessed in vitro activation status of the PI3K-AKT-mTOR signaling pathway following BEZ235 treatment in HER2+ BC cell lines. We next evaluated the impact of BEZ235 on hypoxia-induced or the PI3K-AKT-mTOR pathway ⊠mediated HIF1α stabilization/synthesis which is a master regulator of angiogenesis and also tumor-induced angiogenesis using xenograft models. Results: The results show that 1) BEZ235 inhibited downstream activation of the PI3K-AKT-mTOR signaling pathway effectors, p-AKT (Ser473, The308), p-P70S6K, p-S6RP and p-4EBP1, 2) unlike other pan-PI3Kinase inhibitor (e.g. LY294002, SF1126), BEZ235 has no effect on hypoxia-mediated stabilization of HIF1α however, heregulin-induced HIF1α synthesis which is an important angiogenic modulator in breast cancer cells, was significantly decreased following the treatment of BEZ235 in HER2+ and HER2+/PIK3CA mutated breast cancer cells, 3) interestingly, BEZ235 mediated abrogation of HIF1α synthesis is more pronounced than mTORC1 inhibition alone, 4) integrin-directed endothelial cell migration is one of the critical steps for tumor-induced angiogenesis. BEZ235 abrogates endothelial cells functions especially integrin-directed HUVEC cells migration on vitronectin (avb3/ avb5) and significantly abrogates endothelial cells tube formation on Matrigel and 5) furthermore, we evaluated the effects of BEZ235 on the capacity of HER2+ and HER2+/PIK3CA mutated breast tumor cells to recruit a blood supply in vivo during thrice weekly treatments with BEZ235. A microvessel density (MVD) in control versus BEZ235-treated tumors showed a significant decrease in MVD (CD31+) and also VEGFR immunostaining in BEZ235 treated tumors. Conclusion: Our data suggest that BEZ235 has potent antiangiogenic activity in three different xenograft models probably via mTOR-HIF1α-VEGF signaling axis. Citation Format: Pradip De, Yuliang Sun, Jennifer H. Carlson, Xiaoqian Lin, Nandini Dey, Brian Leyland-Jones. A dual PI3K/mTOR inhibitor, BEZ235 blocks tumor-induced angiogenesis: Evidence for an effect on the tumor and endothelial compartment. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Breast Cancer Research; Oct 17-20, 2015; Bellevue, WA. Philadelphia (PA): AACR; Mol Cancer Res 2016;14(2_Suppl):Abstract nr B11.

Effect of Coronary Artery Bypass Grafting on the Plasma Levels of VEGF, bFGF, Copper and NO as Angiogenic Biomarkers in Chronic Ischemic Heart Disease Patients

Objectives: Myocardial ischemia offers paradoxical protection from ischemic necrosis by the development of coronary collateral circulation through the process of angiogenesis. The current study aimed to assess the plasma levels of vascular endothelial growth factor basic fibroblast growth factor nitric oxide and copper as angiogenic biomarkers to evaluate their role in the development of coronary collateral circulation in patients with chronic ischemic heart disease. Also, to evaluate the angiogenic modulator effects of coronary artery bypass grafting (CABG) via repeated measurements of such biomarkers 3 months postoperative. Methods: This case control and observational study involved 25 ischemic heart disease male patients candidate for elective coronary artery bypass grafting who admitted at the department of cardiothoracic surgery, faculty of medicine, Assiut University and 25 healthy age matched males as control group. Spectrophotometric assay of NO and copper and ELISA assay of VEGF and bFGF plasma levels were done. Results: Significant higher plasma VEGF, bFGF, copper and NO levels preoperatively when compared with the control group (p<0.01 for all) with significant lowering of their plasma levels when repeatedly measured three months following the CABG (p<0.01 for all). Conclusions: Our findings revealed that plasma levels of VEGF, bFGF, copper and NO have been significantly decreased in patients with chronic myocardial ischemia following CABG, thus, proving the proper coronary reperfusion effect of CABG with decrease the demand for the angiogenesis in the ischemic myocardium. There was a positive correlation between the plasma levels of VEGF and NO.

Lipid Raft Integrity Is Required for Survival of Triple Negative Breast Cancer Cells.

PurposeLipid rafts are cholesterol enriched microdomains that colocalize signaling pathways involved in cell proliferation, metastasis, and angiogenesis. We examined the effect of methyl-β-cyclodextrin (MβCD)-mediated cholesterol extraction on the proliferation, adhesion, invasion, and angiogenesis of triple negative breast cancer (TNBC) cells.MethodsWe measured cholesterol and estimated cell toxicity. Detergent resistant membrane (DRM) and non-DRM fractions were separated using the OptiPrep gradient method. Cell cycles stages were analyzed by flow cytometry, apoptosis was assessed using the TdT-mediated dUTP nick end-labeling assay, and metastasis was determined using a Matrigel invasion assay. Neo-vessel pattern and levels of angiogenic modulators were determined using an in vitro angiogenesis assay and an angiogenesis array, respectively.ResultsThe present study found that the cholesterol-depleting agent MβCD, efficiently depleted membrane cholesterol and caused concentration dependent (0.1–0.5 mM) cytotoxicity compared to nystatin and filipin III in TNBC cell lines, MDA-MB 231 and MDA-MB 468. A reduced proportion of caveolin-1 found in DRM fractions indicated a cholesterol extraction-induced disruption of lipid raft integrity. MβCD inhibited 52% of MDA-MB 231 cell adhesion on fibronectin and 56% of MDA-MB 468 cell adhesion on vitronectin, while invasiveness of these cells was decreased by 48% and 52% respectively, following MβCD treatment (48 hours). MβCD also caused cell cycle arrest at the G2M phase and apoptosis in MDA-MB 231 cells (25% and 58% cells, respectively) and in MDA-MB 468 cells (30% and 38% cells, respectively). We found that MβCD treated cells caused a 52% and 58% depletion of neovessel formation in both MDA-MB 231 and MDA-MB 468 cell lines, respectively. This study also demonstrated that MβCD treatment caused a respective 2.6- and 2.5-fold depletion of tyrosine protein kinase receptor (TEK) receptor tyrosine kinase levels in both TNBC cell lines.ConclusionMβCD-induced cholesterol removal enhances alterations in lipid raft integrity, which reduces TNBC cell survival.

Role of the microRNA, miR-206, and its target PIK3C2α in endothelial progenitor cell function – potential link with coronary artery disease.

Coronary artery disease is a major cause of morbidity and mortality worldwide. Impaired endothelial function and integrity are major contributory factors to coronary artery disease. MicroRNAs have been proposed to play an important role in coronary artery disease pathogenesis. In the present study, the expression of miR-206 was found to be significantly upregulated in peripheral blood endothelial progenitor cells from patients with coronary artery disease compared to healthy donors. MiR-206 was found to regulate endothelial progenitor cell activities by targeting the protein kinase PIK3C2α, which showed decreased expression in coronary artery disease endothelial progenitor cells. Knockdown of miR-206 in coronary artery disease endothelial progenitor cells rescued their angiogenic and vasculogenic abilities both in vitro and in vivo in a mouse ischemic hindlimb model. Furthermore, knockdown of miR-206 activated not only PIK3C2α, but also the angiogenic signal modulators Akt and endothelial nitric oxide synthase. It is therefore proposed that repression of the phosphoinositide 3-kinase/Akt/endothelial nitric oxide synthase signal transduction pathway by miR-206 downregulates angiogenesis contributing to the pathophysiology of coronary artery disease.

Angiogenesis-regulating microRNAs and Ischemic Stroke.

Stroke is a leading cause of death and disability worldwide. Ischemic stroke is the dominant subtype of stroke and results from focal cerebral ischemia due to occlusion of major cerebral arteries. Thus, the restoration or improvement of reduced regional cerebral blood supply in a timely manner is very critical for improving stroke outcomes and poststroke functional recovery. The recovery from ischemic stroke largely relies on appropriate restoration of blood flow via angiogenesis. Newly formed vessels would allow increased cerebral blood flow, thus increasing the amount of oxygen and nutrients delivered to affected brain tissue. Angiogenesis is strictly controlled by many key angiogenic factors in the central nervous system, and these molecules have been well-documented to play an important role in the development of angiogenesis in response to various pathological conditions. Promoting angiogenesis via various approaches that target angiogenic factors appears to be a useful treatment for experimental ischemic stroke. Most recently, microRNAs (miRs) have been identified as negative regulators of gene expression in a post-transcriptional manner. Accumulating studies have demonstrated that miRs are essential determinants of vascular endothelial cell biology/angiogenesis as well as contributors to stroke pathogenesis. In this review, we summarize the knowledge of stroke-associated angiogenic modulators, as well as the role and molecular mechanisms of stroke-associated miRs with a focus on angiogenesis-regulating miRs. Moreover, we further discuss their potential impact on miR-based therapeutics in stroke through targeting and enhancing post-ischemic angiogenesis.

Heparin/Heparan sulfate proteoglycans glycomic interactome in angiogenesis: biological implications and therapeutical use.

Angiogenesis, the process of formation of new blood vessel from pre-existing ones, is involved in various intertwined pathological processes including virus infection, inflammation and oncogenesis, making it a promising target for the development of novel strategies for various interventions. To induce angiogenesis, angiogenic growth factors (AGFs) must interact with pro-angiogenic receptors to induce proliferation, protease production and migration of endothelial cells (ECs). The action of AGFs is counteracted by antiangiogenic modulators whose main mechanism of action is to bind (thus sequestering or masking) AGFs or their receptors. Many sugars, either free or associated to proteins, are involved in these interactions, thus exerting a tight regulation of the neovascularization process. Heparin and heparan sulfate proteoglycans undoubtedly play a pivotal role in this context since they bind to almost all the known AGFs, to several pro-angiogenic receptors and even to angiogenic inhibitors, originating an intricate network of interaction, the so called “angiogenesis glycomic interactome”. The decoding of the angiogenesis glycomic interactome, achievable by a systematic study of the interactions occurring among angiogenic modulators and sugars, may help to design novel antiangiogenic therapies with implications in the cure of angiogenesis-dependent diseases.