Vascular Tube Formation Research Articles

Neuropilin-1 aggravates liver cirrhosis by promoting angiogenesis via VEGFR2-dependent PI3K/Akt pathway in hepatic sinusoidal endothelial cells.

BackgroundWe have revealed that neuropilin-1 (NRP-1) promoted hepatic stellate cell activation and liver fibrosis through its profibrogenic signalling pathways. However, the role of NRP-1 in angiogenesis in hepatic sinusoidal endothelial cells (HSECs) during liver cirrhosis remains unclear.MethodsThe correlation between NRP-1 expression and angiogenesis was evaluated in both human and murine cirrhotic liver tissues by immunohistochemical staining, quantitative real-time PCR, and western blotting. In addition, the role and mechanism of NRP-1 in regulating VEGFR2-dependent angiogenesis was identified in endothelial cells (ECs) in vitro. Moreover, liver histocultures were used to test the therapeutic effect of NRP-1 blocking in liver fibrosis.FindingsHigher expression of NRP-1 in HSECs was detected, which was positively correlated with angiogenesis in liver cirrhosis. In vitro, NRP-1 knockdown suppressed the expression and activation of VEGFR2, accompanied by reduced ability of the vascular tube formation and the migration of ECs. Conversely, NRP-1 overexpression upregulated VEGFR2, promoted tube formation, and the migration of ECs. Mechanistically, NRP-1 modulated the expression of VEGFR2 by regulating FAK and its kinase activity. Furthermore, NRP-1 promoted VEGFR2-dependent angiogenesis via the PI3K/Akt pathway in HSECs. Blocking NRP-1 function reduced intrahepatic angiogenesis and fibrosis-associated factors in the in vitro liver histocultures.InterpretationNRP-1 promotes angiogenesis by upregulating the expression and activation of VEGFR2 through the PI3K/Akt signalling pathway in liver cirrhosis. This study highlights the possibility of therapeutically targeting NRP-1 for the treatment of cirrhosis.FundNational Natural Science Foundation of China (No. 81570551; 81770607; 81600469; 81401868), Key Research project of Shandong Province (No. 2016GSF201008; 2017GSF218053), Natural Science Foundation of Shandong Province (No. ZR2017MH102), National Science and Technology Major Project of China (No. 2018ZX10302206-001-006).

ACE2 inhibits breast cancer angiogenesis via suppressing the VEGFa/VEGFR2/ERK pathway

BackgroundBreast cancer angiogenesis is key for metastasis and predicts a poor prognosis. Angiotensin-converting enzyme 2 (ACE2), as a member of the renin-angiotensin system (RAS), was reported to restrain the progression of hepatocellular carcinoma (HCC) and non-small cell lung cancer (NSCLC) through inhibiting angiogenesis. However, the relationship between ACE2 and breast cancer angiogenesis remains unclear.MethodsThe prognosis and relative gene selection were analysed using the GEPIA, GEO, TCGA and STRING databases. ACE2 expression in breast cancer tissue was estimated by reverse transcription-quantitative polymerase chain reaction (qPCR). Breast cancer cell migration, proliferation and angiogenesis were assessed by Transwell migration, proliferation, tube formation, and wound healing assays. The expression of vascular endothelial growth factor A (VEGFa) was detected by qPCR and Western blotting. The phosphorylation of vascular endothelial growth factor receptor 2 (VEGFR2), mitogen-activated protein kinase 1/2 (MEK1/2), and extracellular signal-regulated protein kinase 1/2 (ERK1/2) was examined by Western blotting. Breast cancer metastasis and angiogenesis in vivo were measured using a zebrafish model.ResultsACE2 was downregulated in breast cancer patients. Patients with higher ACE2 expression had longer relapse-free survival (RFS). In vitro, ACE2 inhibited breast cancer migration. Meanwhile, ACE2 in breast cancer cells inhibited human umbilical vascular endothelial cell (HUVEC) proliferation, tube formation and migration. In the zebrafish model, ACE2 inhibited breast cancer cell metastasis, as demonstrated by analyses of the number of disseminated foci and the metastatic distance. Neo-angiogenesis was also decreased by ACE2. ACE2 downregulated the expression of VEGFa in breast cancer cells. Furthermore, ACE2 in breast cancer cells inactivated the phosphorylation of VEGFR2, MEK1/2, and ERK1/2 in HUVECs.ConclusionsOur findings suggest that ACE2, as a potential resister to breast cancer, might inhibit breast cancer angiogenesis through the VEGFa/VEGFR2/ERK pathway.Trial registrationRetrospectively registered.

β2-Adrenergic receptor expression in human endometrial endothelial cells potentially mediates 2-hydroxyestradiol-induced microvascular stabilization: implications for endometriosis

Catecholestrogens are biologically active metabolites of 17β-estradiol (E2) synthesized by cytochrome P450 enzymes forming 2-Hydroxyestradiol (OHE2) and 4-OHE2. Although catecholestrogens bind estrogen receptors (ERs) with low affinity, their high affinity for β2-adrenergic receptors (β2-AR) regulates uterine endothelial cell proliferation in gestation1. Women with endometriosis have aberrant E2 metabolizing enzymes potentially inducing catecholestrogen production2. We hypothesize that ectopic and eutopic endometrial microvascular endothelial cells express β2-AR and that, under stimulation by 2-OHE2, endometrial endothelial cells have increased cell viability and an effect on endothelial tube stabilization. Paired eutopic/ectopic endometrial sections from women with endometriosis in the proliferative (n=5) or secretory (n=4) phases were immunostained for β2-AR and evaluated semi-quantitatively by HSCORE. Confluent human endometrial endothelial cells (HEEC) grown in 96-well plates were pretreated with control or with β-AR antagonist propranolol (2x10-6 M) followed by treatment with vehicle (control) or with 10-8 M E2 or 2-OHE2. Then, vascular tube formation (15x103 cells/well, n=3) and endothelial proliferation (5x103 cells/well, n=3) were evaluated. Tube formation was analyzed at 2, 4, 8, 16, and 24h post-treatment. Images were utilized to score endothelial tube formation and degradation based on the following scores: initial tube coalescence= 1, initiation of tube networks= 2, completed tube networks= 3, vascular destabilization= 4, and complete loss of vascular network= 5. XTT assays measured cell viability as an indicator of proliferation at 24h post-treatment. Statistical analysis used a t-test or One-way ANOVA and post hoc Tukey test. Immunohistochemistry analysis revealed increased β2-AR staining in vascular endothelial cells in ectopic compared to eutopic tissue during the proliferative phase (P<0.05) and showed a trend toward increased β2-AR staining in eutopic compared to ectopic tissue in the secretory phase; however, it did not achieve significance (P=0.055). In vitro endothelial tube formation analyses revealed no difference in scores at 2, 4, and 8h post-treatment with both E2 and 2-OHE2 compared with control. However, a difference at 16 and 24h post-treatment existed in that treatment with E2 and 2-OHE2 led to lower scores (mean = 3) compared to control (mean = 5) indicating a stabilization of endothelial tube structure in the treatment groups. XTT analyses revealed that compared to control, there were no significant differences in proliferative index in the treatment groups compared to control (P>0.05).Figure 2Endothelial Tube Formation Assay showing control compared to E2 10-8 M or 2-OHE2 10-8 M at hours 2, 4, 8, 16, and 24h post-treatment.View Large Image Figure ViewerDownload Hi-res image Download (PPT) The increased β2-AR expression in endothelial cells in ectopic endometriotic tissue during the proliferative phase suggests it plays a role in angiogenesis during implantation of ectopic endometrial tissue. Our in vitro data, that stabilization of newly formed endothelial tube network by 2-OHE2, suggest that the effect of 2-OHE2 may be mediated by β2-AR on endometriotic endothelial cells to maturate newly formed vasculatures during angiogenesis, thereby contributing to endometriosis pathogenesis.

Ameliorating effects of Cuscuta chinensis Lamak extract on hind‑limb ischemia, and angiogenic‑ or inflammatory associated factors in ovariectomized mice.

Cuscutachinensis Lamak (CCL) has traditionally been used in Korea to treat sexual disorders and skin problems. The aim of the present study was to investigate the effects of CCL extract on surgical injury‑induced ischemia in the hind limbs of mice. Specifically, female C57BL/6 mice were ovariectomized, and their hind‑limb vessels were ligated with surgical silk (6‑0) and excised. CCL (150or450mg/kg/BW) was then administered to the mice for 3weeks, and the blood flow rate was evaluated using a laser Doppler system at ‑7,0,7,14 and21days following hind‑limb ischemia. The serum expression profiles of angiogenic and inflammatory mediators were measured using an antibody array, and the transcript levels were reverse transcription‑quantitative polymerase chain reaction. The rate of hind limb blood flow was normalized to non‑ischemic lesions and revealed to be markedly elevated at 14 and 21days following ischemia when compared with the vehicle group. The density of capillaries in the hind limbs was also significantly increased following treatment with CCL in a dose‑dependent manner. In addition, the transcriptional expression of angiogenetic factors were upregulated, whereas that of inflammatory cytokines were downregulated. Finally, vascular endothelial cell migration and tube formation were evaluated invitro using human umbilical vein endothelial cells (HUVECs) and identified to be significantly increased following treatment with CCL. Overall, the results of the present study indicate that CCL extract exhibits therapeutic potential for the treatment of hind‑limb ischemia as it promotes peripheral angiogenic and anti‑inflammatory effects in mice.

CAENORHABDITIS ELEGANS AND ANGIOGENESIS

Abstract: Angiogenesis is sprouting of new capillaries from already existing ones. It is a dynamic process that can be seen in every phase of human life. It is among the dynamic mechanisms of both physiological and pathological processes. Vascular endothelial growth factor is one of the many molecules that play a role in angiogenesis. Vascular endothelial growth factor is released specifically to the endothelium. It regulates mitogenesis, vascular tone, vascular permeability and vasodilatation in the vascular endothelium. Caenorhabditis elegans is a nematode used to detect and screen the developmental processes and genetic mutations. It is appropriate to study at the organism level to isolate cells and to demonstrate intercellular interactions in vivo. Polyvinyl fluoride-1 is a molecule that plays a role in the neural development of Caenorhabditis elegans. In addition, the polyvinyl fluoride-1 molecule is told to be effective in angiogenesis. Studies have shown that polyvinyl fluoride-1 binds to vascular endothelial growth factor receptor-1 and vascular endothelial growth factor receptor-2, but not to vascular endothelial growth factor receptor-3 and platelete derivated growth factor receptor β. In the research of human umbilical vein endothelial lines, it was observed that polyvinyl fluoride-1 induced angiogenesis and vascular tube formation. These results suggest that Caenorhabditis elegans may have a very important role in vascular endothelial growth factor studies. Caenorhabditis elegans model is used in many scientific areas such as aging, nervous system and genetic changes. However, only a few laboratories around the world studied the Caenorhabditis elegans angiogenesis model. Besides, this model is not currently used in Turkey. This provides a great advantage in terms of the utilization of this model in angiogenesis.

Protective effect of troxerutin and cerebroprotein hydrolysate injection on cerebral ischemia through inhibition of oxidative stress and promotion of angiogenesis in rats.

Brain ischemia, including cerebral ischemia and cerebrovascular ischemia, leads to poor oxygen supply or cerebral hypoxia, and causes brain tissue death or cerebral infarction/ischemic stroke. The troxerutin and cerebroprotein hydrolysate injection (TCHI), is widely applied in China to improve blood supply in ischemic brain tissues and to enhance neuroprotective effects in clinical practice. However, the benefits and detailed underlying mechanism elaborating the effectiveness of TCHI in cerebrovascular diseases require further investigation. Therefore, in the present study, experimental in vivo and in vitro models were employed to investigate the potential mechanisms of TCHI on cerebral ischemic injury. The results demonstrated that TCHI increased the lactate dehydrogenase levels in the brain homogenate and conversely decreased lactic acid levels. TCHI was further observed to significantly increase superoxide dismutase activity and decrease malondialdehyde levels in ischemic brain tissues. In addition, TCHI significantly induced vascular maturation processes, including proliferation, adhesion, migration and tube formation in cultured human umbilical vein endothelial cells. Additionally, TCHI significantly stimulated microvessel formation in the rat aortic ring and chick chorioallantoic membrane assays. Taken together, these results provided strong evidence that TCHI stimulated angiogenesis at multiple steps, and indicated that TCHI attenuated cerebral ischemic damage through the amelioration of oxidative stress and promotion of angiogenesis.

Induction of Mesoderm and Neural Crest-Derived Pericytes from Human Pluripotent Stem Cells to Study Blood-Brain Barrier Interactions

SummaryIn the CNS, perivascular cells (“pericytes”) associate with endothelial cells to mediate the formation of tight junctions essential to the function of the blood-brain barrier (BBB). The BBB protects the CNS by regulating the flow of nutrients and toxins into and out of the brain. BBB dysfunction has been implicated in the progression of Alzheimer's disease (AD), but the role of pericytes in BBB dysfunction in AD is not well understood. In the developing embryo, CNS pericytes originate from two sources: mesoderm and neural crest. In this study, we report two protocols using mesoderm or neural crest intermediates, to generate brain-specific pericyte-like cells from induced pluripotent stem cell (iPSC) lines created from healthy and AD patients. iPSC-derived pericytes display stable expression of pericyte surface markers and brain-specific genes and are functionally capable of increasing vascular tube formation and endothelial barrier properties.

Mesoporous silica nanoparticles as a delivery system for improving antiangiogenic therapy.

PurposeAntiangiogenic drugs usually have short-acting efficacy and poor treatment compliance. The purpose of this study was to determine whether mesoporous silica nanoparticles (MSNs) could be utilized as a nanodrug delivery system for improving antiangiogenic therapy.Materials and methodsMSN-encapsulated bevacizumab nanoparticles were prepared by the nanocasting strategy and characterized by Fourier transform infrared, transmission electron microscopy, and Brunauer–Emmett–Teller method. Encapsulation efficiency and drug loading efficiency of MSN-encapsulated bevacizumab nanoparticles were calculated. The pharmacokinetics, cytotoxicity, and tissue toxicity were evaluated in vitro and in vivo. The antiangiogenic effects of MSN-bevacizumab nanoparticles were evaluated in vitro and in vivo.ResultsMSN encapsulation could prolong the residency of bevacizumab in vitreous/aqueous humor and maintain the long-lasting drug concentration. MSN-encapsulated bevacizumab nanoparticles did not show any obvious cytotoxicity and tissue toxicity. MSN-encapsulated bevacizumab nanoparticles were more effective than bevacizumab in suppressing vascular endothelial growth factor-induced endothelial cell proliferation, migration, and tube formation in vitro. MSN-encapsulated bevacizumab nanoparticles showed sustained inhibitory effects on corneal neovascularization and retinal neovascularization in vivo.ConclusionThis study provides a novel strategy of encapsulating bevacizumab to protect and deliver it, which could increase the time between administration and formulation shelf-life. MSN-encapsulated bevacizumab is a promising drug delivery alternative of antiangiogenic therapy.

Multifunctional nanoparticles based on a polymeric copper chelator for combination treatment of metastatic breast cancer

Copper plays an important role in tumor growth and metastasis. Copper chelation has been confirmed to be an effective strategy for breast cancer therapy through antiangiogenesis. In this work, a copper chelating coil-comb block copolymer RGD-PEG-b-PGA-g-(TETA-DTC-PHis) (RPTDH) was synthesized and used to prepare nanoparticles for loading resiquimod (R848), a TLR7 and TLR8 agonist, thus to combine antiangiogenesis and immune activation to treat breast cancer. RPTDH has strong copper-chelating ability and could self-assemble to form spherical nanoparticles with significant pH-sensitivity. R848 was efficiently loaded into RPTDH nanoparticles and exhibited greatly accelerated releases in weakly acid media simulating tumor microenvironment. RPTDH/R848 nanoparticles significantly inhibited the mobility, invasion and vascular tube formation of HUVECs via copper chelation, demonstrating their strong antiangiogenic activity in vitro. Furthermore, RPTDH/R848 nanoparticles remarkably induced the maturation and activation of human plasmacytoid dendritic CAL-1 cells, indicating their immune-activation ability. In breast tumor-bearing mice, RPTDH/R848 nanoparticles displayed excellent targeting ability for both primary breast tumor and lung metastases, and furthermore dramatically suppressed tumor growth and metastasis through copper deficiency-triggered antiangiogenesis and R848-induced immune activation. In summary, RPTDH/R848 nanoparticles can be used as an therapeutic agent against metastatic breast cancer through combining antiangiogenesis and immune activation.

Cancer-Associated Fibroblasts Promote Angiogenesis of Hepatocellular Carcinoma by VEGF-Mediated EZH2/VASH1 Pathway.

Background:Hepatocellular carcinoma is a highly vascularized tumor, so it is critical to study its angiogenesis. Cancer-associated fibroblasts and enhancer of zeste homolog 2 play an important role in tumor angiogenesis and became significant hallmarks of cancer. But the relationship between enhancer of zeste homolog-2 and cancer-associated fibroblasts in response to angiogenesis and its precise mechanism remains unclear.Methods:The separation of cancer-associated fibroblasts was identified by immunofluorescence. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide analysis was used to reveal the proliferation of human umbilical vein endothelial cells. Vascular endothelial growth factor level was quantified by enzyme-linked immunosorbent assay. The wound healing, transwell, and vascular tube formation assays were used to identify the capability of migration, invasion, and tube formation of human umbilical vein endothelial cells in vitro. The detection of tumor angiogenesis was also performed in vivo. Finally, the level of enhancer of zeste homolog-2 and vasohibin 1 were determined by quantitative real-time polymerase chain reaction and Western blotting.Results:In comparison to control and condition medium noncancerous fibroblasts groups, the condition medium cancer-associated fibroblasts could significantly promote the proliferation, migration, invasion, and angiogenesis of human umbilical vein endothelial cells. We found that cancer-associated fibroblasts promoted angiogenesis of human umbilical vein endothelial cells via vascular endothelial growth factor secretion in vitro and in vivo. The upregulation of enhancer of zeste homolog 2 by vascular endothelial growth factor inhibited the expression of vasohibin 1, thus promoting the proliferation and angiogenesis of human umbilical vein endothelial cells. Taken together, the cancer-associated fibroblasts of hepatocellular carcinoma regulate the enhancer of zeste homolog-2/vasohibin 1 pathway via vascular endothelial growth factor secretion, thereby promoting the proliferation and angiogenesis of human umbilical vein endothelial cells.Conclusion:Our study identified the relationship between cancer-associated fibroblasts and enhancer of zeste homolog-2 and confirmed the pivotal role of cancer-associated fibroblasts in angiogenesis of hepatocellular carcinoma. Cancer-associated fibroblasts promote angiogenesis of hepatocellular carcinoma by vascular endothelial growth factor–mediated enhancer of zeste homolog-2/vasohibin 1 pathway and may be a potentially useful therapeutic target for hepatocellular carcinoma.

(-)-Epigallocatechin-3-gallate protects against uric acid-induced endothelial dysfunction in human umbilical vein endothelial cells

Background: Hyperuricemia is recently reported to be associated with hypertension, metabolic syndrome, and vascular damage. (-)-Epigallocatechin gallate (EGCG) is a major polyphenol component of green tea involving in potent anti-inflammatory and antioxidant effects. The study assessed the effect of EGCG on uric acid (UA)-induced vascular damage. Materials and Methods: The cell viability and angiogenic formation of human umbilical vein endothelial cells (HUVECs) treated with UA and EGCG were determined by methylthiazol tetrazolium and tube formation assays, respectively. The expression level of inflammatory cytokine and vascular factor, including nuclear factor-kappa B (NF-κB), monocyte chemotactic protein-1 (MCP-1), intercellular adhesion molecule 1 (ICAM-1), cycloxygenase-2, tumor necrosis factor (TNF-α), induced nitric oxide synthetase (iNOS), endothelin-1 (ET-1), and von Willebrand factor, were examined by real-time quantitative reverse transcription polymerase chain reaction. The expression of P65 in HUVECs treated with UA in different time points or different concentrations was detected by Western blotting. Results: EGCG suppressed UA-inducing HUVEC cells death. UA treatment of HUVEC significantly increases the expression of P65. EGCG significantly inhibited the UA-induced mRNA expression of NF-κB, MCP-1, ICAM-1, TNF-α, iNOS, and ET-1 in HUVECs. Functional analysis of angiogenic inhibition showed that pretreatment with EGCG improved the vascular tube formation. Conclusion: Our results suggested that UA-induced inflammatory cytokine production and impaired endothelial cell function in HUVECs were attenuated by EGCG. These data indicated that EGCG has a therapeutic potential for UA-mediated endothelial damage.

Biological Effects of IL-26 on T Cell–Mediated Skin Inflammation, Including Psoriasis

Psoriasis is a chronic inflammatory skin disease characterized mainly by epidermal hyperplasia, scaling, and erythema; T helper 17 cells have a role in its pathogenesis. Although IL-26, known as a T helper 17 cytokine, is upregulated in psoriatic skin lesions, its precise role is unclear. We investigated the role of IL-26 in the imiquimod-induced psoriasis-like murine model using human IL-26 transgenic mice. Erythema symptoms induced by daily applications of imiquimod increased dramatically in human IL-26 transgenic mice compared with controls. Vascularization and immune cell infiltration were prominent in skin lesions of human IL-26 transgenic mice. Levels of fibroblast growth factor (FGF) 1, FGF2, and FGF7 were significantly upregulated in the skin lesions of imiquimod-treated human IL-26 transgenic mice and psoriasis patients. Invitro analysis demonstrated that FGF1, FGF2, and FGF7 levels were elevated in human keratinocytes and vascular endothelial cells following IL-26 stimulation. Furthermore, IL-26 acted directly on vascular endothelial cells, promoting proliferation and tube formation, possibly through protein kinase B, extracellular signal-regulated kinase, and NF-κB pathways. Moreover, similar effects of IL-26 were observed in the murine contact hypersensitivity model, indicating that these effects are not restricted to psoriasis. Altogether, our data indicate that IL-26 may be a promising therapeutic target in T cell-mediated skin inflammation, including psoriasis.

Biomechanics of Endothelial Tubule Formation Differentially Modulated by Cerebral Cavernous Malformation Proteins

SummaryAt early stages of organismal development, endothelial cells self-organize into complex networks subsequently giving rise to mature blood vessels. The compromised collective behavior of endothelial cells leads to the development of a number of vascular diseases, many of which can be life-threatening. Cerebral cavernous malformation is an example of vascular diseases caused by abnormal development of blood vessels in the brain. Despite numerous efforts to date, enlarged blood vessels (cavernomas) can be effectively treated only by risky and complex brain surgery. In this work, we use a comprehensive simulation model to dissect the mechanisms contributing to an emergent behavior of the multicellular system. By tightly integrating computational and experimental approaches we gain a systems-level understanding of the basic mechanisms of vascular tubule formation, its destabilization, and pharmacological rescue, which may facilitate the development of new strategies for manipulating collective endothelial cell behavior in the disease context.

Autocrine IL-1β mediates the promotion of corneal neovascularization by senescent fibroblasts.

Our previous study has confirmed that senescent fibroblasts promote corneal neovascularization (CNV) partially via the enhanced secretion of matrix metalloproteases (MMPs). However, the regulation of MMP expression in senescent fibroblasts remained unclear. In this study, we identified that the expression and secretion levels of interleukin-1β (IL-1β) were significantly upregulated in senescent human corneal fibroblasts than that in normal fibroblasts. Moreover, compared with vehicle-pretreated senescent fibroblasts, IL-1β pretreatment enhanced the expression of angiogenic factors but reduced the expression of angiostatic factors in senescent fibroblasts. When cocultured with human umbilical vein endothelial cells, IL-1β-pretreated senescent fibroblasts more strongly promoted their proliferation, migration, and tube-formation capacities than the vehicle-controlled senescent fibroblasts. In addition, either interleukin-1 receptor antagonist or anti-IL-1β neutralization completely inhibited the promotion of senescent fibroblasts in vascular tube formation in vitro and CNV in vivo. Therefore, we concluded that autocrine IL-1β mediated the promotion of senescent fibroblasts on corneal neovascularization.

Neonatal Micro-RNA Profile Determines Endothelial Function in Offspring of Hypertensive Pregnancies.

Offspring of hypertensive pregnancies are at increased risk of developing hypertension in adulthood. In the neonatal period they display endothelial cell dysfunction and altered microvascular development. MicroRNAs, as important endothelial cellular regulators, may play a role in this early endothelial dysfunction. Therefore we identified differential microRNA patterns in endothelial cells from offspring of hypertensive pregnancies and determined their role in postnatal vascular cell function. Studies were performed on human umbilical vein endothelial cell (HUVECs) samples from 57 pregnancies. Unbiased RNA-sequencing identified 30 endothelial-related microRNAs differentially expressed in HUVECs from hypertensive compared to normotensive pregnancies. Quantitative reverse transcription PCR (RT-qPCR) confirmed a significant higher expression level of the top candidate, miR-146a. Combined miR-146a targeted gene expression and pathway analysis revealed significant alterations in genes involved in inflammation, angiogenesis and immune response in the same HUVECs. Elevated miR-146a expression level at birth identified cells with reduced ability for in vitro vascular tube formation, which was rescued by miR-146a inhibition. In contrast, miR-146a overexpression significantly reduced vascular tube formation in HUVECs from normotensive pregnancies. Finally, we confirmed that mir146a levels at birth predicted in vivo microvascular development during the first three postnatal months. Offspring of hypertensive pregnancy have a distinct endothelial regulatory microRNA profile at birth, which is related to altered endothelial cell behaviour, and predicts patterns of microvascular development during the first three months of life. Modification of this microRNA profile in vitro can restore impaired vascular cell function.

Molecular signature of progenitor cells isolated from young and adult human hearts

The loss of endogenous cardiac regenerative capacity within the first week of postnatal life has intensified clinical trials to induce cardiac regeneration in the adult mammalian heart using different progenitor cell types. We hypothesized that donor age-related phenotypic and functional characteristics of cardiac progenitor cells (CPC) account for mixed results of cell-based cardiac repair. We compared expression profiles and cell turnover rates of human heart-derived c-kitpos progenitors (c-kitpos CPC) and cardiosphere-derived cells (CDC) from young and adult donor origin and studied their in vitro angiogenic and cardiac differentiation potential, which can be relevant for cardiac repair. We report that 3-dimensional CDC expansion recapitulates a conducive environment for growth factor and cytokine release from adult donor cells (aCDC) that optimally supports vascular tube formation and vessel sprouting. Transdifferentiation capacity of c-kitpos CPCs and CDCs towards cardiomyocyte-like cells was modest, however, most notable in young c-kitpos cells and adult CDCs. Progenitors isolated with different methods thus show cell- and donor-specific characteristics that may account for variable contributions in functional myocardial recovery.

LncRNA PVT1 promotes the angiogenesis of vascular endothelial cell by targeting miR‑26b to activate CTGF/ANGPT2.

Angiogenesis is essential for various biological processes, including tumor blood supply delivery, cancer cell growth, invasion and metastasis. Plasmacytoma variant translocation1 (PVT1) long noncoding RNA (lncRNA) has been previously reported to affect angiogenesis of glioma microvascular endothelial cells by regulating microRNA (miR)‑186 expression level. However, the specific underlying molecular mechanism of PVT1 regulation of angiogenesis in vascular endothelial cells remains to be elucidated. The present study investigated the role of PVT1 in cell proliferation, migration and vascular tube formation of human umbilical vein endothelial cells (HUVECs) using MTT assay, Transwell migration assay and invitro vascular tube formation assay, respectively. In order to determine the effect of miR‑26b on cell proliferation, migration and vascular tube formation of HUVECs, miR‑26 mimic or miR‑26b inhibitor were transfected into HUVECs. Reverse transcription‑quantitative polymerase chain reaction and western blotting were conducted to quantify the mRNA and protein expression levels of target genes. The present study confirmed that miR‑26b bound 3'‑untranslated region (3'‑UTR) and subsequently influenced gene expression level using dual luciferase reporter assay. The current study observed that PVT1 affected cell proliferation, migration and invitro vascular tube formation of HUVECs. In addition, it was determined that PVT1 was able to bind and degrade miR‑26b to promote connective tissue growth factor (CTGF) and angiopoietin2 (ANGPT2) expression. miR‑26b was also identified to have a suppressive role in cell proliferation, migration and invitro vascular tube formation of HUVECs via binding 3'‑UTR regions and downregulating CTGF and ANGPT2 expression levels. The current findings may improve the understanding of the underlying mechanism of PVT1 contributing to angiogenesis of vascular endothelial cells and offer rationale for targeting PVT1 to treat angiogenesis dysfunction‑associated diseases, including cancer metastasis.

MiR-103 regulates the angiogenesis of ischemic stroke rats by targeting vascular endothelial growth factor (VEGF).

Objective(s):To investigate the effect of miR-103 on the angiogenesis of ischemic stroke rats via targeting vascular endothelial growth factor (VEGF) at the molecular level.Materials and Methods:Rat models had received the middle cerebral artery occlusion (MCAO) or sham operation before grouping, and cell models of oxygen-glucose deprivation (OGD) were performed. FITC-dextran, matrigel, and Trans-well assays were used to evaluate the vascular density, tube formation, and cell migration respectively. The expression levels of miR-103 and VEGF were detected by quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting. Dual-luciferase assay was used for analyzing the targeting relationship between miR-103 and VEGF.Results:We found the reduced miR-103 in rats after MCAO. Down-regulating miR-103 with the miR-103 inhibitor enhanced VEGF, ameliorated the neurological scores, decreased infarct volume, and increased vascular density in rats after MCAO. Besides, in OGD human umbilical vein endothelial cells (HUVECs), inhibition of miR-103 could promote the increase of tube length and the migration of cells. Additionally, we found that miR-103 could directly target VEGF and thereby lead to the down-expression of VEGF. Meanwhile, si-VEGF could reverse the effect of miR-103 inhibitor on angiogenesis in rats subjected to MCAO.Conclusion:Inhibition of miR-103 could promote ischemic stroke angiogenesis and reduce infarction volume via enhancing VEGF, which provides a new target for the clinical treatment of ischemic stroke.

Defibrotide Stimulates Angiogenesis and Protects Endothelial Cells from Calcineurin Inhibitor-Induced Apoptosis via Upregulation of AKT/Bcl-xL.

Sinusoidal obstruction syndrome is a life-threatening complication that can occur after haematopoietic stem cell transplantation. Defibrotide (DF) has been approved for the treatment of individuals with severe sinusoidal obstruction syndrome following haematopoietic stem cell transplantation in the European Union and the United States. However, the precise mechanisms by which DF protects endothelial cells remain to be elucidated. In this study, we found that DF stimulated angiogenesis in vitro and in vivo as assessed by vascular tube formation, scratch-wound repair and Matrigel plug assays. These effects were associated with an activation of pro-survival signalling pathways, including AKT (protein kinase B), ERK (extracellular signal-regulated kinases) and p38. More importantly, DF alleviated calcineurin inhibitor-induced growth inhibition and apoptosis of human umbilical vein endothelial cells and human hepatic sinusoidal endothelial cells in parallel with upregulation of anti-apoptotic protein B-cell lymphoma-extra-large (Bcl-xL), which was mediated by AKT (protein kinase B). Notably, these effects were abrogated when Bcl-xL was depleted by small interfering RNA (ribonucleic acid). In addition, DF counteracted calcineurin inhibitor-induced activation of nuclear factor-κB and Janus kinase 2 (JAK2)/Signal Transducer and Activator of Transcription 3 (STAT3) signalling and production of cytokines in vascular endothelial cell-derived EA.hy926 cells. Taken together, DF has pro-angiogenic, anti-apoptotic and anti-inflammatory effects on endothelial cells. DF is a potentially useful agent to prevent the development of, and treat individuals with, endothelial cell injury-related complications after haematopoietic stem cell transplantation.

Oxytocin inhibits ovarian cancer metastasis by repressing the expression of MMP-2 and VEGF.

Breastfeeding is associated with a decreased risk of ovarian cancer. However, the mechanism underlying this apparent clinical benefit is unknown. Oxytocin (OXT), a hypothalamic nonapetide, plays a crucial role in many reproductive and behavioural functions. In recent year, OXT acts as a growth regulator in many kind of tumor tissues, through the activation of a specific G-coupled transmembrane receptor, the oxytocin receptor (OXTR). OXT has been proved to inhibit the proliferation, migration and invasion of ovarian cancer SKOV3 cells in vitro. But, the underlying mechanisms remain unknown. Here, we found OXT inhibited proliferation, and critically migration and invasion of human ovarian cancer cells SKOV3 and A2780. Strikingly, OXT inhibited ovarian cancer metastasis by repressing the expressions of MMP-2 and VEGF. Moreover, OXT inhibited vascular endothelial cell tube formation by reducing the VEGF production from ovarian cancer cells. Our findings may provide a possible explanation for breastfeeding-associated protective effects and suggest new therapeutic opportunities for ovarian cancer.