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  1. Microvascular endothelial cells of the corpus luteum

    Directory of Open Access Journals (Sweden)

    Spanel-Borowski Katherina

    2003-11-01

    Full Text Available Abstract The cyclic nature of the capillary bed in the corpus luteum offers a unique experimental model to examine the life cycle of endothelial cells, involving discrete physiologically regulated steps of angiogenesis, blood vessel maturation and blood vessel regression. The granulosa cells and theca cells of the developing antral follicle and the steroidogenic cells of the corpus luteum produce and respond to angiogenic factors and vasoactive peptides. Following ovulation the neovascularization during the early stages of corpus luteum development has been compared to the rapid angiogenesis observed during tumor formation. On the other end of the spectrum, the microvascular endothelial cells are the first cells to undergo apoptosis at the onset of corpus luteum regression. Important insights on the morphology and function of luteal endothelial cells have been gained from a combination of in vitro and in vivo studies on endothelial cells. Endothelial cells communicate with cells comprising the functional unit of the corpus luteum, i.e., other vascular cells, steroidogenic cells, and immune cells. This review is designed to provide an overview of the types of endothelial cells present in the corpus luteum and their involvement in corpus luteum development and regression. Available evidence indicates that microvascular endothelial cells of the corpus luteum are not alike, and may differ during the process of angiogenesis and angioregression. The contributions of vasoactive peptides generated by the luteal endothelin-1 and the renin-angiotensin systems are discussed in context with the function of endothelial cells during corpus luteum formation and regression. The ability of two cytokines, tumor necrosis factor alpha and interferon gamma, are evaluated as paracrine mediators of endothelial cell function during angioregression. Finally, chemokines are discussed as a vital endothelial cell secretory products that contribute to the recruitment of

  2. Rapid homogeneous endothelialization of high aspect ratio microvascular networks.

    Science.gov (United States)

    Naik, Nisarga; Hanjaya-Putra, Donny; Haller, Carolyn A; Allen, Mark G; Chaikof, Elliot L

    2015-08-01

    Microvascularization of an engineered tissue construct is necessary to ensure the nourishment and viability of the hosted cells. Microvascular constructs can be created by seeding the luminal surfaces of microfluidic channel arrays with endothelial cells. However, in a conventional flow-based system, the uniformity of endothelialization of such an engineered microvascular network is constrained by mass transfer of the cells through high length-to-diameter (L/D) aspect ratio microchannels. Moreover, given the inherent limitations of the initial seeding process to generate a uniform cell coating, the large surface-area-to-volume ratio of microfluidic systems demands long culture periods for the formation of confluent cellular microconduits. In this report, we describe the design of polydimethylsiloxane (PDMS) and poly(glycerol sebacate) (PGS) microvascular constructs with reentrant microchannels that facilitates rapid, spatially homogeneous endothelial cell seeding of a high L/D (2 cm/35 μm; > 550:1) aspect ratio microchannels. MEMS technology was employed for the fabrication of a monolithic, elastomeric, reentrant microvascular construct. Isotropic etching and PDMS micromolding yielded a near-cylindrical microvascular channel array. A 'stretch - seed - seal' operation was implemented for uniform incorporation of endothelial cells along the entire microvascular area of the construct yielding endothelialized microvascular networks in less than 24 h. The feasibility of this endothelialization strategy and the uniformity of cellularization were established using confocal microscope imaging.

  3. Differentiation state determines neural effects on microvascular endothelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Muffley, Lara A., E-mail: muffley@u.washington.edu [University of Washington, Campus Box 359796, 300 9th Avenue, Seattle, WA 98104 (United States); Pan, Shin-Chen, E-mail: pansc@mail.ncku.edu.tw [University of Washington, Campus Box 359796, 300 9th Avenue, Seattle, WA 98104 (United States); Smith, Andria N., E-mail: gnaunderwater@gmail.com [University of Washington, Campus Box 359796, 300 9th Avenue, Seattle, WA 98104 (United States); Ga, Maricar, E-mail: marga16@uw.edu [University of Washington, Campus Box 359796, 300 9th Avenue, Seattle, WA 98104 (United States); Hocking, Anne M., E-mail: ahocking@u.washington.edu [University of Washington, Campus Box 359796, 300 9th Avenue, Seattle, WA 98104 (United States); Gibran, Nicole S., E-mail: nicoleg@u.washington.edu [University of Washington, Campus Box 359796, 300 9th Avenue, Seattle, WA 98104 (United States)

    2012-10-01

    Growing evidence indicates that nerves and capillaries interact paracrinely in uninjured skin and cutaneous wounds. Although mature neurons are the predominant neural cell in the skin, neural progenitor cells have also been detected in uninjured adult skin. The aim of this study was to characterize differential paracrine effects of neural progenitor cells and mature sensory neurons on dermal microvascular endothelial cells. Our results suggest that neural progenitor cells and mature sensory neurons have unique secretory profiles and distinct effects on dermal microvascular endothelial cell proliferation, migration, and nitric oxide production. Neural progenitor cells and dorsal root ganglion neurons secrete different proteins related to angiogenesis. Specific to neural progenitor cells were dipeptidyl peptidase-4, IGFBP-2, pentraxin-3, serpin f1, TIMP-1, TIMP-4 and VEGF. In contrast, endostatin, FGF-1, MCP-1 and thrombospondin-2 were specific to dorsal root ganglion neurons. Microvascular endothelial cell proliferation was inhibited by dorsal root ganglion neurons but unaffected by neural progenitor cells. In contrast, microvascular endothelial cell migration in a scratch wound assay was inhibited by neural progenitor cells and unaffected by dorsal root ganglion neurons. In addition, nitric oxide production by microvascular endothelial cells was increased by dorsal root ganglion neurons but unaffected by neural progenitor cells. -- Highlights: Black-Right-Pointing-Pointer Dorsal root ganglion neurons, not neural progenitor cells, regulate microvascular endothelial cell proliferation. Black-Right-Pointing-Pointer Neural progenitor cells, not dorsal root ganglion neurons, regulate microvascular endothelial cell migration. Black-Right-Pointing-Pointer Neural progenitor cells and dorsal root ganglion neurons do not effect microvascular endothelial tube formation. Black-Right-Pointing-Pointer Dorsal root ganglion neurons, not neural progenitor cells, regulate

  4. Antiproliferative effect of elevated glucose in human microvascular endothelial cells

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    Kamal, K.; Du, W.; Mills, I.; Sumpio, B. E.

    1998-01-01

    Diabetic microangiopathy has been implicated as a fundamental feature of the pathological complications of diabetes including retinopathy, neuropathy, and diabetic foot ulceration. However, previous studies devoted to examining the deleterious effects of elevated glucose on the endothelium have been performed largely in primary cultured cells of macrovessel origin. Difficulty in the harvesting and maintenance of microvascular endothelial cells in culture have hindered the study of this relevant population. Therefore, the objective of this study was to characterize the effect of elevated glucose on the proliferation and involved signaling pathways of an immortalized human dermal microvascular endothelial cell line (HMEC-1) that possess similar characteristics to their in vivo counterparts. Human dermal microvascular endothelial cells (HMEC-1) were grown in the presence of normal (5 mM) or high D-glucose (20 mM) for 14 days. The proliferative response of HMEC-1 was compared under these conditions as well as the cAMP and PKC pathways by in vitro assays. Elevated glucose significantly inhibited (P diabetic microangiopathy.

  5. Microvascular endothelial cell heterogeneity : general concepts and pharmacological consequences for anti-angiogenic therapy of cancer

    NARCIS (Netherlands)

    Langenkamp, Elise; Molema, Grietje

    2009-01-01

    Microvascular endothelial cells display a large degree of heterogeneity in function depending on their location in the vascular tree. The existence of organ-specific, microvascular-bed-specific, and even intravascular variations in endothelial cell gene expression emphasizes their high cell-to-cell

  6. Translational Medicine Study on Cardiac Microvascular Endothelial Barrier Function and Myocardial Ischemia/Re-perfusion Injury

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    Yeong Yeh Lee

    2015-09-01

    Full Text Available Vascular endothelial barrier is defined as the ability of endothelial cells and their components that make up the microvascular wall structure in controlling the cellular components and marco-molecular substances in blood from penetrating vascular walls. It is the place for the selective exchange of oxygen, nutrients and metabolites, and has kernel effect in maintaining myocardial micro-environmental homeostasis. In clinic, microvascular permeability is commonly used as the index for evaluating endothelial barrier function. Myocardial microvascular endothelial cells, inter-endothelial connexin and basilar membrane (BM interact synergically to constitute the basis for barrier function, which has a selective permeability effect on interaction between nutrient substances and other myocardial cell molecules. Increase of microvascular permeability is closely associated with cardiovascular events like coronary heart disease (CHD and myocardial ischemia, and is the risk factor for CHD attack. And deep exploration of the mechanism of endothelial permeability and positive selection of new-type re-perfusion complementary drugs for alleviating endothelial permeability can be beneficial in improving the prognosis of patients with acute myocardial infarction (AMI. Therefore, from the view of translational medicine, this study mainly summarized the increase of microvascular permeability and its pathological significance after AMI, physiological and pathological mechanisms of regulating microvascular permeability and complementary therapies for AMI re-perfusion as well as microvascular endothelial barrier function, hoping to provide a basis for improving the prognosis of patients with AMI.

  7. Translational Medicine Study on Cardiac Microvascular Endothelial Barrier Function and Myocardial Ischemia/Re-perfusion Injury

    Institute of Scientific and Technical Information of China (English)

    Yeong Yeh Lee

    2015-01-01

    Vascular endothelial barrier is defined as the ability of endothelial cells and their components that make up the microvascular wall structure in controlling the cellular components and marco-molecular substances in blood from penetrating vascular walls. It is the place for the selective exchange of oxygen, nutrients and metabolites, and has kernel effect in maintaining myocardial micro-environmental homeostasis. In clinic, microvascular permeability is commonly used as the index for evaluating endothelial barrier function. Myocardial microvascular endothelial cells, inter-endothelial connexin and basilar membrane (BM) interact synergically to constitute the basis for barrier function, which has a selective permeability effect on interaction between nutrient substances and other myocardial cell molecules. Increase of microvascular permeability is closely associated with cardiovascular events like coronary heart disease (CHD) and myocardial ischemia, and is the risk factor for CHD attack. And deep exploration of the mechanism of endothelial permeability and positive selection of new-type re-perfusion complementary drugs for alleviating endothelial permeability can be beneifcial in improving the prognosis of patients with acute myocardial infarction (AMI). Therefore, from the view of translational medicine, this study mainly summarized the increase of microvascular permeability and its pathological signiifcance after AMI, physiological and pathological mechanisms of regulating microvascular permeability and complementary therapies for AMI re-perfusion as well as microvascular endothelial barrier function, hoping to provide a basis for improving the prognosis of patients with AMI.

  8. Brain microvascular endothelial cell transplantation ameliorates ischemic white matter damage.

    Science.gov (United States)

    Puentes, Sandra; Kurachi, Masashi; Shibasaki, Koji; Naruse, Masae; Yoshimoto, Yuhei; Mikuni, Masahiko; Imai, Hideaki; Ishizaki, Yasuki

    2012-08-21

    Ischemic insults affecting the internal capsule result in sensory-motor disabilities which adversely affect the patient's life. Cerebral endothelial cells have been reported to exert a protective effect against brain damage, so the transplantation of healthy endothelial cells might have a beneficial effect on the outcome of ischemic brain damage. In this study, endothelin-1 (ET-1) was injected into the rat internal capsule to induce lacunar infarction. Seven days after ET-1 injection, microvascular endothelial cells (MVECs) were transplanted into the internal capsule. Meningeal cells or 0.2% bovine serum albumin-Hank's balanced salt solution were injected as controls. Two weeks later, the footprint test and histochemical analysis were performed. We found that MVEC transplantation improved the behavioral outcome based on recovery of hind-limb rotation angle (P<0.01) and induced remyelination (P<0.01) compared with the control groups. Also the inflammatory response was repressed by MVEC transplantation, judging from fewer ED-1-positive activated microglial cells in the MVEC-transplanted group than in the other groups. Elucidation of the mechanisms by which MVECs ameliorate ischemic damage of the white matter may provide important information for the development of effective therapies for white matter ischemia.

  9. Polylactic Acid Nanoparticles Targeted to Brain Microvascular Endothelial Cells

    Institute of Scientific and Technical Information of China (English)

    WANG Huafang; HU Yu; SUN Wangqiang; XIE Changsheng

    2005-01-01

    In this work, blank polylactic acid (PLA) nanoparticles with unstained surface were prepared by the nano-deposition method. On the basis of the preparation, the effect of surface modification on brain microvascular endothelial cells (BMECs) targeting was examined by in vivo experiments and fluorescence microscopy. The results showed that PLA nanoparticles are less toxic than PACA nanoparticles but their BMECs targeting is similar to PACA nanoparticles. The experiments suggest that drugs can be loaded onto the particles and become more stable through adsorption on the surface of PLA nanoparticles with high surface activity. The surface of PLA nanoparticles was obviously modified and the hydrophilicity was increased as well in the presence of non-ionic surfactants on PLA nanoparticles. As a targeting moiety, polysobate 80 (T-80) can facilitate BMECs targeting of PLA nanoparticles.

  10. Barrier stabilizing mediators in regulation of microvascular endothelial permeability

    Institute of Scientific and Technical Information of China (English)

    HUANG Qiao-bing

    2012-01-01

    Increase of microvascular permeability is one of the most important pathological events in the pathogenesis of trauma and bum injury.Massive leakage of fluid from vascular space leads to lose of blood plasma and decrease of effective circulatory blood volume,resulting in formation of severe tissue edema,hypotension or even shock,especially in severe bum injury.Fluid resuscitation has been the only valid approach to sustain patient's blood volume for a long time,due to the lack of overall and profound understanding of the mechanisms of vascular hyperpenneability response.There is an emerging concept in recent years that some so-called barrier stabilizing mediators play a positive role in preventing the increase of vascular permeability.These mediators may be released in response to proinflammatory mediators and serve to restore endothelial barrier function.Some of these stabilizing mediators are important even in quiescent state because they preserve basal vascular permeability at low levels.This review introduces some of these mediators and reveals their underlying signaling mechanisms during endothelial barrier enhancing process.

  11. Prognosis of invasive breast cancer after adjuvant therapy evaluated with VEGF microvessel density and microvascular imaging.

    Science.gov (United States)

    Li, Ying; Wei, Xi; Zhang, Sheng; Zhang, Jin

    2015-11-01

    The aim of this study was to investigate the role of ultrasonographic microvascular imaging in the evaluation of prognosis of patients with invasive breast cancer treated by adjuvant therapies. A total of 121 patients with invasive breast cancer underwent ultrasonographic contrast-enhanced imaging, vascular endothelial growth factor (VEGF) staining, and microvessel density (MVD) counts. The parameters of microvascular imaging and the expression of VEGF and MVD in primary breast cancer were calculated. The correlation between these factors and the overall and progression-free survival rate were analyzed using the Kaplan-Meier method. Among 121 cases, the positive VEGF cases were 75 and negative ones were 46. The cut point of 52.3 was calculated by the regressive curve for MVD counts. The data showed the mean intensity (MI) was positively associated with both the MVD counts (r = .51, p prognosis of patients, high VEGF expression and MVD counts were associated with reduced progressive and survival times (PFS, p = .032 and p = .034; OS, p = .041 and p = .038, respectively). The correlation between parameters of microvascular imaging, VEGF expressive status, and the MVD counts were established. The cut point of mean intensity (MI = 40) was used to investigate as an independent predictor for PFS (p = .021) and OS (p = .025), respectively, due to a strong correlation between MVD counts and VEGF expression in patients with invasive breast cancer. The microvascular imaging could be a visual and helpful tool to predict the prognosis of patients with invasive breast cancer treated by adjuvant therapies.

  12. Metformin improves endothelial function in aortic tissue and microvascular endothelial cells subjected to diabetic hyperglycaemic conditions.

    Science.gov (United States)

    Ghosh, Suparna; Lakshmanan, Arun P; Hwang, Mu Ji; Kubba, Haidar; Mushannen, Ahmed; Triggle, Chris R; Ding, Hong

    2015-12-01

    The cellular mechanisms whereby metformin, the first line drug for type 2 diabetes (T2DM), mediates its antidiabetic effects remain elusive, particularly as to whether metformin has a direct protective action on the vasculature. This study was designed to determine if a brief 3-h exposure to metformin protects endothelial function against the effects of hyperglycaemia. We investigated the protective effects of metformin on endothelial-dependent vasodilatation (EDV) in thoracic aortae from T2DM db/db mice and on high glucose (HG, 40 mM) induced changes in endothelial nitric oxide synthase (eNOS) signaling in mouse microvascular endothelial cells (MMECs) in culture. Exposure of aortae from db+/? non-diabetic control mice to high glucose (HG, 40 mM) containing Krebs for 3-h significantly (Pmetformin; metformin also improved ACh-induced EDV in aortae from diabetic db/db mice. Immunoblot analysis of MMECs cultured in HG versus NG revealed a significant reduction of the ratio of phosphorylated (p-eNOS)/eNOS and p-Akt/Akt, but not the expression of total eNOS or Akt. The 3-h exposure of MMECs to metformin significantly (Pmetformin can reverse/reduce the impact of HG on endothelial function, via mechanisms linked to increased phosphorylation of eNOS and Akt.

  13. Sirtinol treatment reduces inflammation in human dermal microvascular endothelial cells.

    Directory of Open Access Journals (Sweden)

    Angela Orecchia

    Full Text Available Histone deacetylases (HDAC are key enzymes in the epigenetic control of gene expression. Recently, inhibitors of class I and class II HDAC have been successfully employed for the treatment of different inflammatory diseases such as rheumatoid arthritis, colitis, airway inflammation and asthma. So far, little is known so far about a similar therapeutic effect of inhibitors specifically directed against sirtuins, the class III HDAC. In this study, we investigated the expression and localization of endogenous sirtuins in primary human dermal microvascular endothelial cells (HDMEC, a cell type playing a key role in the development and maintenance of skin inflammation. We then examined the biological activity of sirtinol, a specific sirtuin inhibitor, in HDMEC response to pro-inflammatory cytokines. We found that, even though sirtinol treatment alone affected only long-term cell proliferation, it diminishes HDMEC inflammatory responses to tumor necrosis factor (TNFα and interleukin (IL-1β. In fact, sirtinol significantly reduced membrane expression of adhesion molecules in TNFã- or IL-1β-stimulated cells, as well as the amount of CXCL10 and CCL2 released by HDMEC following TNFα treatment. Notably, sirtinol drastically decreased monocyte adhesion on activated HDMEC. Using selective inhibitors for Sirt1 and Sirt2, we showed a predominant involvement of Sirt1 inhibition in the modulation of adhesion molecule expression and monocyte adhesion on activated HDMEC. Finally, we demonstrated the in vivo expression of Sirt1 in the dermal vessels of normal and psoriatic skin. Altogether, these findings indicated that sirtuins may represent a promising therapeutic target for the treatment of inflammatory skin diseases characterized by a prominent microvessel involvement.

  14. Optically measured microvascular blood flow contrast of malignant breast tumors.

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    Regine Choe

    Full Text Available Microvascular blood flow contrast is an important hemodynamic and metabolic parameter with potential to enhance in vivo breast cancer detection and therapy monitoring. Here we report on non-invasive line-scan measurements of malignant breast tumors with a hand-held optical probe in the remission geometry. The probe employs diffuse correlation spectroscopy (DCS, a near-infrared optical method that quantifies deep tissue microvascular blood flow. Tumor-to-normal perfusion ratios are derived from thirty-two human subjects. Mean (95% confidence interval tumor-to-normal ratio using surrounding normal tissue was 2.25 (1.92-2.63; tumor-to-normal ratio using normal tissues at the corresponding tumor location in the contralateral breast was 2.27 (1.94-2.66, and using normal tissue in the contralateral breast was 2.27 (1.90-2.70. Thus, the mean tumor-to-normal ratios were significantly different from unity irrespective of the normal tissue chosen, implying that tumors have significantly higher blood flow than normal tissues. Therefore, the study demonstrates existence of breast cancer contrast in blood flow measured by DCS. The new, optically accessible cancer contrast holds potential for cancer detection and therapy monitoring applications, and it is likely to be especially useful when combined with diffuse optical spectroscopy/tomography.

  15. Microvascular Coronary Artery Spasm Presents Distinctive Clinical Features With Endothelial Dysfunction as Nonobstructive Coronary Artery Disease

    Science.gov (United States)

    Ohba, Keisuke; Sugiyama, Seigo; Sumida, Hitoshi; Nozaki, Toshimitsu; Matsubara, Junichi; Matsuzawa, Yasushi; Konishi, Masaaki; Akiyama, Eiichi; Kurokawa, Hirofumi; Maeda, Hirofumi; Sugamura, Koichi; Nagayoshi, Yasuhiro; Morihisa, Kenji; Sakamoto, Kenji; Tsujita, Kenichi; Yamamoto, Eiichiro; Yamamuro, Megumi; Kojima, Sunao; Kaikita, Koichi; Tayama, Shinji; Hokimoto, Seiji; Matsui, Kunihiko; Sakamoto, Tomohiro; Ogawa, Hisao

    2012-01-01

    events over 47.8±27.5 months. Conclusions Microvascular CAS causes distinctive clinical features and endothelial dysfunction that are important to recognize as nonobstructive coronary artery disease so that optimal care with calcium channel blockers can be provided. Clinical Trial Registration URL: www.umin.ac.jp/ctr. Unique identifier: UMIN000003839. PMID:23316292

  16. Alternative venous outflow vessels in microvascular breast reconstruction.

    Science.gov (United States)

    Mehrara, Babak J; Santoro, Timothy; Smith, Andrew; Arcilla, Eric A; Watson, James P; Shaw, William W; Da Lio, Andrew L

    2003-08-01

    The lack of adequate recipient vessels often complicates microvascular breast reconstruction in patients who have previously undergone mastectomy and irradiation. In addition, significant size mismatch, particularly in the outflow veins, is an important contributor to vessel thrombosis and flap failure. The purpose of this study was to review the authors' experience with alternative venous outflow vessels for microvascular breast reconstruction. In a retrospective analysis of 1278 microvascular breast reconstructions performed over a 10-year period, the authors identified all patients in whom the external jugular or cephalic veins were used as the outflow vessels. Patient demographics, flap choice, the reasons for the use of alternative venous drainage vessels, and the incidence of microsurgical complications were analyzed. The external jugular was used in 23 flaps performed in procedures with 22 patients. The superior gluteal and transverse rectus abdominis musculocutaneous (TRAM) flaps were used in the majority of the cases in which the external jugular vein was used (72 percent gluteal, 20 percent TRAM flap). The need for alternative venous outflow vessels was usually due to a significant vessel size mismatch between the superior gluteal and internal mammary veins (74 percent). For three of the external jugular vein flaps (13 percent), the vein was used for salvage after the primary draining vein thrombosed, and two of three flaps in these cases were eventually salvaged. In three patients, the external jugular vein thrombosed, resulting in two flap losses, while the third was salvaged using the cephalic vein. A total of two flaps were lost in the external jugular vein group. The cephalic vein was used in 11 flaps (TRAM, 64.3 percent; superior gluteal, 35.7 percent) performed in 11 patients. In five patients (54.5 percent), the cephalic vein was used to salvage a flap after the primary draining vein thrombosed; the procedure was successful in four cases. In three

  17. Propionyl-L-Carnitine Enhances Wound Healing and Counteracts Microvascular Endothelial Cell Dysfunction.

    Directory of Open Access Journals (Sweden)

    Maria Giovanna Scioli

    Full Text Available Impaired wound healing represents a high cost for health care systems. Endothelial dysfunction characterizes dermal microangiopathy and contributes to delayed wound healing and chronic ulcers. Endothelial dysfunction impairs cutaneous microvascular blood flow by inducing an imbalance between vasorelaxation and vasoconstriction as a consequence of reduced nitric oxide (NO production and the increase of oxidative stress and inflammation. Propionyl-L-carnitine (PLC is a natural derivative of carnitine that has been reported to ameliorate post-ischemic blood flow recovery.We investigated the effects of PLC in rat skin flap and cutaneous wound healing. A daily oral PLC treatment improved skin flap viability and associated with reactive oxygen species (ROS reduction, inducible nitric oxide synthase (iNOS and NO up-regulation, accelerated wound healing and increased capillary density, likely favoring dermal angiogenesis by up-regulation for iNOS, vascular endothelial growth factor (VEGF, placental growth factor (PlGF and reduction of NADPH-oxidase 4 (Nox4 expression. In serum-deprived human dermal microvascular endothelial cell cultures, PLC ameliorated endothelial dysfunction by increasing iNOS, PlGF, VEGF receptors 1 and 2 expression and NO level. In addition, PLC counteracted serum deprivation-induced impairment of mitochondrial β-oxidation, Nox4 and cellular adhesion molecule (CAM expression, ROS generation and leukocyte adhesion. Moreover, dermal microvascular endothelial cell dysfunction was prevented by Nox4 inhibition. Interestingly, inhibition of β-oxidation counteracted the beneficial effects of PLC on oxidative stress and endothelial dysfunction.PLC treatment improved rat skin flap viability, accelerated wound healing and dermal angiogenesis. The beneficial effects of PLC likely derived from improvement of mitochondrial β-oxidation and reduction of Nox4-mediated oxidative stress and endothelial dysfunction. Antioxidant therapy and

  18. Transport and regulation mechanism of the colloidal gold liposomes in the brain microvascular endothelial cells

    Institute of Scientific and Technical Information of China (English)

    WANG Lipeng; CHANG Yanzhong

    2015-01-01

    Objective:Blood-brain barrier is the key barrier of brain in the innate immune. It can prevent the harmful substances from the blood into the brain. In order to keep the brain in a relatively stable environment and maintain the normal function of the nervous system, it can also pump harmful substances or excess substances outside the brain selectively. Among them, brain microvascular endothelial cell tissue is a key part in the blood-brain barrier's function. The number of the patients with central nervous system ( CNS) diseases increased year by year. The therapeutic drug is usually inhibited by the blood-brain barrier and is difficult to work. Therefore, how to modify the drug and to make it easier to cross the blood brain barrier is the key point to cure CNS. At present, more than 95% research focus only on how nano drugs can enter the cell, the way and efficiency to enter the cell and the research of effect of nano drug etc. For the process of drug carrier in endocytosis, intracellular transport and release and regulation of research are rarely reported. Clathrin and P-glycoprotein are related protein in endo-cytosis and exocytosis with nano drug. Clathrin is located on the plasma membrane. It participates in endocytosis of some nutrients, and maybe the entry into the cell of some drugs. P-glycoprotein is located in the membrane of cer-ebral capillary endothelial cells. It can efflux drugs relying on ATP. Although there is a certain understanding of the cell in the inner swallow and efflux. But the process of the liposome drug is not clear. To solve the above prob-lems, using colloidal gold liposome nano materials to trace liposome's transport and regulation mechanism in brain microvascular endothelial cells, and study endocytosis, release, distribution and regulation mechanism of nano lipo-somes in brain microvascular. The solution of this problem can guide to construct reasonable drug carrier, and look forward to clarifing the molecular basis and mechanism of

  19. ID3 contributes to the acquisition of molecular stem cell-like signature in microvascular endothelial cells: its implication for understanding microvascular diseases.

    Science.gov (United States)

    Das, Jayanta K; Voelkel, Norbert F; Felty, Quentin

    2015-03-01

    While significant progress has been made to advance our knowledge of microvascular lesion formation, yet the investigation of how stem-like cells may contribute to the pathogenesis of microvascular diseases is still in its infancy. We assessed whether the inhibitor of DNA binding and differentiation 3 (ID3) contributes to the acquisition of a molecular stem cell-like signature in microvascular endothelial cells. The effects of stable ID3 overexpression and SU5416 treatment - a chemical inducer of microvascular lesions, had on the stemness signature were determined by flow cytometry, immunoblot, and immunohistochemistry. Continuous ID3 expression produced a molecular stemness signature consisting of CD133(+) VEGFR3(+) CD34(+) cells. Cells exposed to SU5416 showed positive protein expression of ID3, VEGFR3, CD34 and increased expression of pluripotent transcription factors Oct-4 and Sox-2. ID3 overexpressing cells supported the formation of a 3-D microvascular lesion co-cultured with smooth muscle cells. In addition, in vivo microvascular lesions from SuHx rodent model showed an increased expression of ID3, VEGFR3, and Pyk2 similar to SU5416 treated human endothelial cells. Further investigations into how normal and stem-like cells utilize ID3 may open up new avenues for a better understanding of the molecular mechanisms which are underlying the pathological development of microvascular diseases.

  20. Demonstration of actin filament stress fibers in microvascular endothelial cells in situ.

    Science.gov (United States)

    Nehls, V; Drenckhahn, D

    1991-07-01

    We have developed a method for immunostaining the microvascular tree of rat mesenteric windows in situ. The procedure consists of three steps, i.e., mild fixation with formaldehyde, controlled proteolytic digestion of the mesothelial layer, and permeabilization with acetone. Discrimination between different microvascular segments was possible by double-fluorescent staining with antibodies to the smooth muscle isoform of alpha-actin and to nonmuscle myosin from platelets. Antibodies to nonmuscle myosin labeled numerous longitudinally oriented cables in endothelial cells of all microvascular segments (arterioles, metarterioles, pre-, mid-, and postcapillaries, small venules). Occasionally, the myosin-containing cables displayed the interrupted sarcomere-like staining pattern that is diagnostic for stress fibers. In contrast, staining of actin filaments with phalloidin-rhodamin resulted in a noninterrupted, continuous fluorescence of the stress fibers. A possible functional role of microvascular endothelial stress fibers is to serve as a tensile cytoskeletal scaffold that stabilizes the tubular, three-dimensional geometry of microvessels and, in addition, to help the endothelium resist the shear forces created by blood flow and by collision with red and white blood cells.

  1. Cathepsin S Cleavage of Protease-Activated Receptor-2 on Endothelial Cells Promotes Microvascular Diabetes Complications.

    Science.gov (United States)

    Kumar Vr, Santhosh; Darisipudi, Murthy N; Steiger, Stefanie; Devarapu, Satish Kumar; Tato, Maia; Kukarni, Onkar P; Mulay, Shrikant R; Thomasova, Dana; Popper, Bastian; Demleitner, Jana; Zuchtriegel, Gabriele; Reichel, Christoph; Cohen, Clemens D; Lindenmeyer, Maja T; Liapis, Helen; Moll, Solange; Reid, Emma; Stitt, Alan W; Schott, Brigitte; Gruner, Sabine; Haap, Wolfgang; Ebeling, Martin; Hartmann, Guido; Anders, Hans-Joachim

    2016-06-01

    Endothelial dysfunction is a central pathomechanism in diabetes-associated complications. We hypothesized a pathogenic role in this dysfunction of cathepsin S (Cat-S), a cysteine protease that degrades elastic fibers and activates the protease-activated receptor-2 (PAR2) on endothelial cells. We found that injection of mice with recombinant Cat-S induced albuminuria and glomerular endothelial cell injury in a PAR2-dependent manner. In vivo microscopy confirmed a role for intrinsic Cat-S/PAR2 in ischemia-induced microvascular permeability. In vitro transcriptome analysis and experiments using siRNA or specific Cat-S and PAR2 antagonists revealed that Cat-S specifically impaired the integrity and barrier function of glomerular endothelial cells selectively through PAR2. In human and mouse type 2 diabetic nephropathy, only CD68(+) intrarenal monocytes expressed Cat-S mRNA, whereas Cat-S protein was present along endothelial cells and inside proximal tubular epithelial cells also. In contrast, the cysteine protease inhibitor cystatin C was expressed only in tubules. Delayed treatment of type 2 diabetic db/db mice with Cat-S or PAR2 inhibitors attenuated albuminuria and glomerulosclerosis (indicators of diabetic nephropathy) and attenuated albumin leakage into the retina and other structural markers of diabetic retinopathy. These data identify Cat-S as a monocyte/macrophage-derived circulating PAR2 agonist and mediator of endothelial dysfunction-related microvascular diabetes complications. Thus, Cat-S or PAR2 inhibition might be a novel strategy to prevent microvascular disease in diabetes and other diseases.

  2. Inhibition of microvascular endothelial cell apoptosis by angiopoietin-1 and the involvement of cytochrome C

    Institute of Scientific and Technical Information of China (English)

    SHI Lian-guo; ZHANG Guo-ping; JIN Hui-ming

    2006-01-01

    Background Angiopoietin-1 (Ang-1) is an endothelial-specific growth factor that can promote angiogenesis.Studies demonstrated that Ang-1 can inhibit apoptosis of umbilical endothelial cells, but so far little is known about its effects on apoptosis of microvascular endothelial cells. With the apoptotic model of murinecerebral-derived microvascular endothelial cells (bEnd.3) induced by serum-free culture,we attempted to clarify the molecular mechanism of bEnd.3 apoptosis, particularly its relation to cytochrome C (Cyt C).Methods The cultured microvascular endothelial cell strain, bEnd.3 cell, was employed. An apoptotic model of bEnd.3 was established by serum-free culture. Flow cytometry after Annexin labeling and PI staining were used to assess the apoptotic effects of Ang-1 on bEnd.3, and the expression of Bax/Bcl-2, caspase 8, caspase 3, and Cyt C were detected with Western blotting and ELISA.Results The apoptotic rate of bEnd.3 cells after stimulation with Ang-1 (100 ng/L) in serum-free medium was significantly higher than that in control group. Ang-1 inhibited early-stage apoptosis more than late-stage apoptosis provided by propidium iodide (PI) and AnnexinV double staining. The inhibition of Ang-1 on bEnd.3cell apoptosis was strengthened with the increase in concentration (0-400 ng/ml). Ang-1 could decrease the expression of Bax, caspase3 and 8, and increase that of Bcl-2. The results of ELISA indicated that Ang-1significantly decreased CytC content in cytoplasm and increase that in mitochondria.Conclusions Ang-1 could inhibit bEnd.3 apoptosis induced by serum-free medium culture. The apoptosis was associated with decreased Bax expression, increased Bcl-2 expression, which result in Cyt C transferring from mitochondria to cytoplasm, and then caspases activation are reduced and cell apoptosis is suppressed.

  3. Glutathione in Cerebral Microvascular Endothelial Biology and Pathobiology: Implications for Brain Homeostasis

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    Wei Li

    2012-01-01

    Full Text Available The integrity of the vascular endothelium of the blood-brain barrier (BBB is central to cerebrovascular homeostasis. Given the function of the BBB as a physical and metabolic barrier that buffers the systemic environment, oxidative damage to the endothelial monolayer will have significant deleterious impact on the metabolic, immunological, and neurological functions of the brain. Glutathione (GSH is a ubiquitous major thiol within mammalian cells that plays important roles in antioxidant defense, oxidation-reduction reactions in metabolic pathways, and redox signaling. The existence of distinct GSH pools within the subcellular organelles supports an elegant mode for independent redox regulation of metabolic processes, including those that control cell fate. GSH-dependent homeostatic control of neurovascular function is relatively unexplored. Significantly, GSH regulation of two aspects of endothelial function is paramount to barrier preservation, namely, GSH protection against oxidative endothelial cell injury and GSH control of postdamage cell proliferation in endothelial repair and/or wound healing. This paper highlights our current insights and hypotheses into the role of GSH in cerebral microvascular biology and pathobiology with special focus on endothelial GSH and vascular integrity, oxidative disruption of endothelial barrier function, GSH regulation of endothelial cell proliferation, and the pathological implications of GSH disruption in oxidative stress-associated neurovascular disorders, such as diabetes and stroke.

  4. Uptake of Single-Walled Carbon Nanotubes Conjugated with DNA by Microvascular Endothelial Cells

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    Joseph Harvey

    2012-01-01

    Full Text Available Single-walled carbon nanotubes (SWCNTs have been proposed to have great therapeutic potential. SWCNTs conjugated with drugs or genes travel in the systemic circulation to reach target cells or tissues following extravasation from microvessels although the interaction between SWCNT conjugates and the microvascular endothelial cells (ECs remains unknown. We hypothesized that SWCNT-DNA conjugates would be taken up by microvascular ECs and that this process would be facilitated by SWCNTs compared to facilitation by DNA alone. ECs were treated with various concentrations of SWCNT-DNA-FITC conjugates, and the uptake and intracellular distribution of these conjugates were determined by a confocal microscope imaging system followed by quantitative analysis of fluorescence intensity. The uptake of SWCNT-DNA-FITC conjugates (2 μg/mL by microvascular ECs was significantly greater than that of DNA-FITC (2 μg/mL, observed at 6 hrs after treatment. For the intracellular distribution, SWCNT-DNA-FITC conjugates were detected in the nucleus of ECs, while DNA-FITC was restricted to the cytoplasm. The fluorescence intensity and distribution of SWCNTs were concentration and time independent. The findings demonstrate that SWCNTs facilitate DNA delivery into microvascular ECs, thus suggesting that SWCNTs serving as drug and gene vehicles have therapeutic potential.

  5. Internal Mammary Recipient Site Breast Cancer Recurrence Following Delayed Microvascular Breast Reconstruction

    Science.gov (United States)

    Rosich-Medina, Anais; Wang, Susan; Erel, Ertan; Malata, Charles M.

    2013-01-01

    Objective: The internal mammary vessels are a popular recipient site for microsurgical anastomoses of free flap breast reconstructions. We, however, observed 3 patients undergoing internal mammary vessel delayed free flap breast reconstruction that subsequently developed tumor recurrence at this site. We reviewed their characteristics to determine whether there was a correlation between delayed microsurgical reconstruction and local recurrence. Methods: A retrospective review of a single surgeon's delayed free flap breast reconstructions using the internal mammary vessels was conducted over a 7-year period to identify the time intervals between mastectomy and delayed breast reconstruction and between delayed breast reconstruction and recurrence. Results: Three patients developed local recurrence at the site of the microvascular anastomoses following delayed breast reconstruction. All patients had been disease-free following mastectomy. The median time interval between mastectomy and delayed breast reconstruction was 28 months (range = 20-120 months) while that between delayed breast reconstruction and local recurrence was 7 months (range = 4-10 months). Two patients died from metastatic disease, 36 and 72 months following their local recurrence. One patient remains alive 44 months after reconstruction. Conclusions: Local tumor recurrence at the internal mammary vessel dissection site following delayed breast reconstruction raises the question whether these 2 events may be related. Specifically, could internal mammary vessel dissection undertaken for delayed microsurgical reconstruction predispose to recurrence in the internal mammary lymph nodes? Further research is needed to ascertain whether delayed breast reconstruction increases the risk of local recurrence in this patient group. PMID:23383360

  6. Differential effects of Bartonella henselae on human and feline macro- and micro-vascular endothelial cells.

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    Berrich, Moez; Kieda, Claudine; Grillon, Catherine; Monteil, Martine; Lamerant, Nathalie; Gavard, Julie; Boulouis, Henri Jean; Haddad, Nadia

    2011-01-01

    Bartonella henselae, a zoonotic agent, induces tumors of endothelial cells (ECs), namely bacillary angiomatosis and peliosis in immunosuppressed humans but not in cats. In vitro studies on ECs represent to date the only way to explore the interactions between Bartonella henselae and vascular endothelium. However, no comparative study of the interactions between Bartonella henselae and human (incidental host) ECs vs feline (reservoir host) ECs has been carried out because of the absence of any available feline endothelial cell lines.To this purpose, we have developed nine feline EC lines which allowed comparing the effects of Bartonella strains on human and feline micro-vascular ECs representative of the infection development sites such as skin, versus macro-vascular ECs, such as umbilical vein.Our model revealed intrinsic differences between human (Human Skin Microvascular ECs -HSkMEC and Human Umbilical Vein ECs - iHUVEC) and feline ECs susceptibility to Bartonella henselae infection.While no effect was observed on the feline ECs upon Bartonella henselae infection, the human ones displayed accelerated angiogenesis and wound healing.Noticeable differences were demonstrated between human micro- and macro-vasculature derived ECs both in terms of pseudo-tube formation and healing. Interestingly, Bartonella henselae effects on human ECs were also elicited by soluble factors.Neither Bartonella henselae-infected Human Skin Microvascular ECs clinically involved in bacillary angiomatosis, nor feline ECs increased cAMP production, as opposed to HUVEC.Bartonella henselae could stimulate the activation of Vascular Endothelial Growth Factor Receptor-2 (VEGFR-2) in homologous cellular systems and trigger VEGF production by HSkMECs only, but not iHUVEC or any feline ECs tested.These results may explain the decreased pathogenic potential of Bartonella henselae infection for cats as compared to humans and strongly suggest that an autocrine secretion of VEGF by human skin

  7. Differential effects of Bartonella henselae on human and feline macro- and micro-vascular endothelial cells.

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    Moez Berrich

    Full Text Available Bartonella henselae, a zoonotic agent, induces tumors of endothelial cells (ECs, namely bacillary angiomatosis and peliosis in immunosuppressed humans but not in cats. In vitro studies on ECs represent to date the only way to explore the interactions between Bartonella henselae and vascular endothelium. However, no comparative study of the interactions between Bartonella henselae and human (incidental host ECs vs feline (reservoir host ECs has been carried out because of the absence of any available feline endothelial cell lines.To this purpose, we have developed nine feline EC lines which allowed comparing the effects of Bartonella strains on human and feline micro-vascular ECs representative of the infection development sites such as skin, versus macro-vascular ECs, such as umbilical vein.Our model revealed intrinsic differences between human (Human Skin Microvascular ECs -HSkMEC and Human Umbilical Vein ECs - iHUVEC and feline ECs susceptibility to Bartonella henselae infection.While no effect was observed on the feline ECs upon Bartonella henselae infection, the human ones displayed accelerated angiogenesis and wound healing.Noticeable differences were demonstrated between human micro- and macro-vasculature derived ECs both in terms of pseudo-tube formation and healing. Interestingly, Bartonella henselae effects on human ECs were also elicited by soluble factors.Neither Bartonella henselae-infected Human Skin Microvascular ECs clinically involved in bacillary angiomatosis, nor feline ECs increased cAMP production, as opposed to HUVEC.Bartonella henselae could stimulate the activation of Vascular Endothelial Growth Factor Receptor-2 (VEGFR-2 in homologous cellular systems and trigger VEGF production by HSkMECs only, but not iHUVEC or any feline ECs tested.These results may explain the decreased pathogenic potential of Bartonella henselae infection for cats as compared to humans and strongly suggest that an autocrine secretion of VEGF by human

  8. Stathmin expression in glioma-derived microvascular endothelial cells: a novel therapeutic target.

    Science.gov (United States)

    Dong, Baijing; Mu, Luyan; Qin, Xiangying; Qiao, Wanchen; Liu, Xiaodong; Yang, Liming; Xue, Li; Rainov, Nikolai G; Liu, Xiaoqian

    2012-03-01

    The purpose of this study was to investigate stathmin expression and its mechanisms of action in GDMEC. Microvascular endothelial cells were isolated from human gliomas (n=68) and normal brain specimans (n=20), and purified by magnetic beads coated with anti-CD105 antibody. The expression of stathmin mRNA and protein were detected by RT-PCR and western blotting, respectively. Stathmin expression was silenced by application of specific siRNA in high grade GDMEC. The proliferation, apoptosis and invasion behavior of GDMEC were investigated. The stathmin positive rate of endothelial cells in normal brain, grade I-II glioma and grade III-IV glioma was 20, 66 and 95.5%, respectively (Pstathmin, cell viability was reduced, the apoptosis rate increased and the migration of vascular endothelial cells was suppressed significantly (Pstathmin suppressed neoangiogenesis of glioma and provides a potential target for glioma treatment.

  9. IL-1α-induced microvascular endothelial cells promote neutrophil killing by increasing MMP-9 concentration and lysozyme activity.

    Science.gov (United States)

    Liu, Xiaoye; Dong, Hong; Wang, Mingming; Gao, Ying; Zhang, Tao; Hu, Ge; Duan, Huiqing; Mu, Xiang

    2016-02-01

    The recruitment of neutrophils by endothelial cells during infection has been extensively studied, but little is known about the regulation of neutrophils activity by endothelial cells. To examine the role of microvascular endothelial cells in neutrophil killing, we established a transmigration model using rat intestinal microvascular endothelial cells (RIMVECs) and measured the extracellular and intracellular killing of Escherichia coli, Lactobacillus acidophilus, and Staphylococcus aureus by transendothelial neutrophils. We observed that blood neutrophils engulfed bacteria but did not kill them, and lipopolysaccharide- or hemolysin-injured RIMVECs inhibited the extracellular and intracellular bactericidal activity of transendothelial neutrophils. In comparison, interleukin-1α-induced RIMVECs promoted the extracellular and intracellular killing activity of transendothelial neutrophils and significantly increased MMP-9 concentration and lysozyme activity in transendothelial neutrophils (p neutrophils and bacterial toxin damage of endothelial cells led to reduction in bactericidal activity of transendothelial neutrophils. These findings offered new insight into the role of endothelial cells in the bactericidal activity of neutrophils.

  10. Pseudomonas aeruginosa exotoxin Y-mediated tau hyperphosphorylation impairs microtubule assembly in pulmonary microvascular endothelial cells.

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    Ron Balczon

    Full Text Available Pseudomonas aeruginosa uses a type III secretion system to introduce the adenylyl and guanylyl cyclase exotoxin Y (ExoY into the cytoplasm of endothelial cells. ExoY induces Tau hyperphosphorylation and insolubility, microtubule breakdown, barrier disruption and edema, although the mechanism(s responsible for microtubule breakdown remain poorly understood. Here we investigated both microtubule behavior and centrosome activity to test the hypothesis that ExoY disrupts microtubule dynamics. Fluorescence microscopy determined that infected pulmonary microvascular endothelial cells contained fewer microtubules than control cells, and further studies demonstrated that the microtubule-associated protein Tau was hyperphosphorylated following infection and dissociated from microtubules. Disassembly/reassembly studies determined that microtubule assembly was disrupted in infected cells, with no detectable effects on either microtubule disassembly or microtubule nucleation by centrosomes. This effect of ExoY on microtubules was abolished when the cAMP-dependent kinase phosphorylation site (Ser-214 on Tau was mutated to a non-phosphorylatable form. These studies identify Tau in microvascular endothelial cells as the target of ExoY in control of microtubule architecture following pulmonary infection by Pseudomonas aeruginosa and demonstrate that phosphorylation of tau following infection decreases microtubule assembly.

  11. Short- and long-term effects of silver nanoparticles on human microvascular endothelial cells

    Institute of Scientific and Technical Information of China (English)

    Sara; Castiglioni; Clelia; Caspani; Alessandra; Cazzaniga; Jeanette; AM; Maier

    2014-01-01

    AIM: To study the response to silver nanoparticles(Ag NP) of human microvascular endothelial cells, protagonists of angiogenesis. METHODS: We cultured human microvascular endothelial cells and endothelial colony-forming cells in their corresponding growth medium. Stock solutions of Ag NP were prepared in culture medium and sonicated before use. They were added at different concentrations and for different times to culture media. The toxicity of Ag NP was investigated by measuring the reduction of yellow tetrazolium salt to dark purple formazan(MTT assay) at 575 nm. After staining with trypan blue, cell proliferation was assessed by counting viable cells. The lactate dehydrogenase leakage assay was performed on culture media by following the oxidation of NADH to NAD+ and monitoring the reaction kinetically at 340 nm. Reactive oxygen species production was quantified using 2’-7’-dichlorofluorescein diacetate. The alkaline comet assay was performed after mixing the cells with low melting-point agarose. Electrophoresis was then conducted and the samples were stained with ethidium bromide and analyzed with a fluorescence microscope.RESULTS: Ag NP are cytotoxic in a dose and time dependent fashion for HMEC. At high concentrations, Ag NP determine loss of membrane integrity as demonstrated by the increased activity of lactate dehydrogenase in the culture medium. Ag NP rapidly stimulate the formation of free radicals. However, pre-incubation with Trolox, apocynin, or N-acetyl-L-cysteine, antioxidants which have different structure and act through different mechanisms, is not sufficient to prevent cytotoxicity. Ag NP also induce DNA damage dose-dependently, as shown by comet assay. When exposed to sublethal concentrations of Ag NP for long times, the cells remain viable but are growth retarded. Interestingly, removal of Ag NP partially rescues cell growth. Also genotoxicity is reversible upon removal of Ag NP from culture medium, suggesting that no permanent

  12. Magnetic particle spectroscopy allows precise quantification of nanoparticles after passage through human brain microvascular endothelial cells

    Science.gov (United States)

    Gräfe, C.; Slabu, I.; Wiekhorst, F.; Bergemann, C.; von Eggeling, F.; Hochhaus, A.; Trahms, L.; Clement, J. H.

    2016-06-01

    Crossing the blood-brain barrier is an urgent requirement for the treatment of brain disorders. Superparamagnetic iron oxide nanoparticles (SPIONs) are a promising tool as carriers for therapeutics because of their physical properties, biocompatibility, and their biodegradability. In order to investigate the interaction of nanoparticles with endothelial cell layers in detail, in vitro systems are of great importance. Human brain microvascular endothelial cells are a well-suited blood-brain barrier model. Apart from generating optimal conditions for the barrier-forming cell units, the accurate detection and quantification of SPIONs is a major challenge. For that purpose we use magnetic particle spectroscopy to sensitively and directly quantify the SPION-specific iron content. We could show that SPION concentration depends on incubation time, nanoparticle concentration and location. This model system allows for further investigations on particle uptake and transport at cellular barriers with regard to parameters including particles’ shape, material, size, and coating.

  13. Effect of acetylsalicylic acid on microvascular thrombosis in autologous breast reconstruction.

    Science.gov (United States)

    Enajat, Morteza; Aziz Mohammadi, Mujzgan; Debeij, Jan; van der Hulst, Rene R W J; Mureau, Marc A M

    2014-01-01

    Although advances in microsurgery have increased success rates of autologous breast reconstruction, microvascular thrombosis still remains a major concern as a cause of flap failure. At present, no evidence-based guidelines on pharmacological prevention of microvascular thrombosis exist. This study investigates the effect of acetylsalicylic acid on the incidence of microvascular complications in patients undergoing autologous breast reconstruction. Patients undergoing deep inferior epigastric artery perforator or free transverse rectus abdominis myocutaneous flap breast reconstruction at two academic centers in the Netherlands between 2005 and 2011 were included. Patients at one center received once daily 0.6 mL of nadroparine and 40 mg acetylsalicylic acid, while patients at the other center received 0.6 mL nadroparine only. A total of 430 consecutive patients underwent 592 breast reconstructions. No statistically significant differences were found between the two groups in the incidence of flap failure (2.8 and 2.5%), microvascular thromboembolic complications (2.6 and 3.8%), venous congestion (3.4 and 2.8%), or overall complications (28.0 and 32.3%). Hematoma tended to occur more often in the group receiving acetylsalicylic acid (9.2 and 4.7%).It was found that no protective effect of acetylsalicylic acid on microvascular complications was present. Given its known risks and the somewhat increased occurrence of hematoma in the present study, we stopped to routinely administer acetylsalicylic acid after autologous breast reconstruction.

  14. Pericyte abundance affects sucrose permeability in cultures of rat brain microvascular endothelial cells.

    Science.gov (United States)

    Parkinson, Fiona E; Hacking, Cindy

    2005-07-05

    The blood-brain barrier is a physical and metabolic barrier that restricts diffusion of blood-borne substances into brain. In vitro models of the blood-brain barrier are used to characterize this structure, examine mechanisms of damage and repair and measure permeability of test substances. The core component of in vitro models of the blood-brain barrier is brain microvascular endothelial cells. We cultured rat brain microvascular endothelial cells (RBMEC) from isolated rat cortex microvessels. After 2-14 days in vitro (DIV), immunohistochemistry of these cells showed strong labeling for zona occludens 1 (ZO-1), a tight junction protein expressed in endothelial cells. Pericytes were also present in these cultures, as determined by expression of alpha-actin. The present study was performed to test different cell isolation methods and to compare the resulting cell cultures for abundance of pericytes and for blood-brain barrier function, as assessed by 14C-sucrose flux. Two purification strategies were used. First, microvessels were preabsorbed onto uncoated plastic for 4 h, then unattached microvessels were transferred to coated culture ware. Second, microvessels were incubated with an antibody to platelet-endothelial cell adhesion molecule 1 (PECAM-1; CD31) precoupled to magnetic beads, and a magnetic separation procedure was performed. Our results indicate that immunopurification, but not preadsorption, was an effective method to purify microvessels and reduce pericyte abundance in the resulting cultures. This purification significantly reduced 14C-sucrose fluxes across cell monolayers. These data indicate that pericytes can interfere with the development of blood-brain barrier properties in in vitro models that utilize primary cultures of RBMECs.

  15. Human brain microvascular endothelial cells resist elongation due to shear stress.

    Science.gov (United States)

    Reinitz, Adam; DeStefano, Jackson; Ye, Mao; Wong, Andrew D; Searson, Peter C

    2015-05-01

    Endothelial cells in straight sections of vessels are known to elongate and align in the direction of flow. This phenotype has been replicated in confluent monolayers of bovine aortic endothelial cells and human umbilical vein endothelial cells (HUVECs) in cell culture under physiological shear stress. Here we report on the morphological response of human brain microvascular endothelial cells (HBMECs) in confluent monolayers in response to shear stress. Using a microfluidic platform we image confluent monolayers of HBMECs and HUVECs under shear stresses up to 16 dyne cm(-2). From live-cell imaging we quantitatively analyze the cell morphology and cell speed as a function of time. We show that HBMECs do not undergo a classical transition from cobblestone to spindle-like morphology in response to shear stress. We further show that under shear stress, actin fibers are randomly oriented in the cells indicating that there is no cytoskeletal remodeling. These results suggest that HBMECs are programmed to resist elongation and alignment under shear stress, a phenotype that may be associated with the unique properties of the blood-brain barrier.

  16. Application of a clinical grade CD34-mediated method for the enrichment of microvascular endothelial cells from fat tissue.

    NARCIS (Netherlands)

    Arts, C.H.; Groot, P. de; Heijnen-Snyder, G.J.; Blankensteijn, J.D.; Eikelboom, B.C.; Slaper-Cortenbach, I.C.M.

    2004-01-01

    BACKGROUND: Microvascular endothelial cells (MVEC) derived from s.c. fat are seeded on vascular grafts to prevent early occlusion. We have demonstrated the presence of contaminating cells contributing to MVEC seeding-related intimal hyperplasia in MVEC isolates from fat tissue. We found that cell is

  17. Application of a clinical grade CD34-mediated method for the enrichment of microvascular endothelial cells from fat tissue

    NARCIS (Netherlands)

    Arts, CHP; de Groot, PG; Heijnen-Snyder, GJ; Blankensgteijn, JD; Eikelboom, BC; Slaper-Cortenbach, ICM

    2004-01-01

    Background Microvascular endothelial cells (MVEC) derived from s.c. fat are seeded on vascular grafts to prevent early occlusion. We have demonstrated the presence of contaminating cells contributing to MVEC seeding-related intimal hyperplasia in MVEC isolates from fat tissue. We found that cell iso

  18. Action of shiga toxin type-2 and subtilase cytotoxin on human microvascular endothelial cells.

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    María M Amaral

    Full Text Available The hemolytic uremic syndrome (HUS associated with diarrhea is a complication of Shiga toxin (Stx-producing Escherichia coli (STEC infection. In Argentina, HUS is endemic and responsible for acute and chronic renal failure in children younger than 5 years old. The human kidney is the most affected organ due to the presence of very Stx-sensitive cells, such as microvascular endothelial cells. Recently, Subtilase cytotoxin (SubAB was proposed as a new toxin that may contribute to HUS pathogenesis, although its action on human glomerular endothelial cells (HGEC has not been described yet. In this study, we compared the effects of SubAB with those caused by Stx2 on primary cultures of HGEC isolated from fragments of human pediatric renal cortex. HGEC were characterized as endothelial since they expressed von Willebrand factor (VWF and platelet/endothelial cell adhesion molecule 1 (PECAM-1. HGEC also expressed the globotriaosylceramide (Gb3 receptor for Stx2. Both, Stx2 and SubAB induced swelling and detachment of HGEC and the consequent decrease in cell viability in a time-dependent manner. Preincubation of HGEC with C-9 -a competitive inhibitor of Gb3 synthesis-protected HGEC from Stx2 but not from SubAB cytotoxic effects. Stx2 increased apoptosis in a time-dependent manner while SubAB increased apoptosis at 4 and 6 h but decreased at 24 h. The apoptosis induced by SubAB relative to Stx2 was higher at 4 and 6 h, but lower at 24 h. Furthermore, necrosis caused by Stx2 was significantly higher than that induced by SubAB at all the time points evaluated. Our data provide evidence for the first time how SubAB could cooperate with the development of endothelial damage characteristic of HUS pathogenesis.

  19. Slit2-Robo4 receptor responses inhibit ANDV directed permeability of human lung microvascular endothelial cells.

    Science.gov (United States)

    Gorbunova, Elena E; Gavrilovskaya, Irina N; Mackow, Erich R

    2013-08-01

    Hantaviruses nonlytically infect human endothelial cells (ECs) and cause edematous and hemorrhagic diseases. Andes virus (ANDV) causes hantavirus pulmonary syndrome (HPS), and Hantaan virus (HTNV) causes hemorrhagic fever with renal syndrome (HFRS). Hantaviruses enhance vascular endothelial growth factor directed EC permeability resulting in the disassembly of inter-endothelial cell adherens junctions (AJs). Recent studies demonstrate that Slit2 binding to Robo1/Robo4 receptors on ECs has opposing effects on AJ disassembly and vascular fluid barrier functions. Here we demonstrate that Slit2 inhibits ANDV and HTNV induced permeability and AJ disassembly of pulmonary microvascular ECs (PMECs) by interactions with Robo4. In contrast, Slit2 had no effect on the permeability of ANDV infected human umbilical vein ECs (HUVECs). Analysis of Robo1/Robo4 expression determined that PMECs express Robo4, but not Robo1, while HUVECs expressed both Robo4 and Robo1 receptors. SiRNA knockdown of Robo4 in PMECs prevented Slit2 inhibition of ANDV induced permeability demonstrating that Robo4 receptors determine PMEC responsiveness to Slit2. Collectively, this data demonstrates a selective role for Slit2/Robo4 responses within PMECs that inhibits ANDV induced permeability and AJ disassembly. These findings suggest Slit2s utility as a potential HPS therapeutic that stabilizes the pulmonary endothelium and antagonizes ANDV induced pulmonary edema.

  20. Inhibition of Murine Pulmonary Microvascular Endothelial Cell Apoptosis Promotes Recovery of Barrier Function under Septic Conditions

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    Lefeng Wang

    2017-01-01

    Full Text Available Sepsis is characterized by injury of the pulmonary microvasculature and the pulmonary microvascular endothelial cells (PMVEC, leading to barrier dysfunction and acute respiratory distress syndrome (ARDS. Our recent work identified a strong correlation between PMVEC apoptosis and microvascular leak in septic mice in vivo, but the specific role of apoptosis in septic PMVEC barrier dysfunction remains unclear. Thus, we hypothesize that PMVEC apoptosis is likely required for PMVEC barrier dysfunction under septic conditions in vitro. Septic stimulation (mixture of tumour necrosis factor α, interleukin 1β, and interferon γ [cytomix] of isolated murine PMVEC resulted in a significant loss of barrier function as early as 4 h after stimulation, which persisted until 24 h. PMVEC apoptosis, as reflected by caspase activation, DNA fragmentation, and loss of membrane polarity, was first apparent at 8 h after cytomix. Pretreatment of PMVEC with the pan-caspase inhibitor Q-VD significantly decreased septic PMVEC apoptosis and was associated with reestablishment of PMVEC barrier function at 16 and 24 h after stimulation but had no effect on septic PMVEC barrier dysfunction over the first 8 h. Collectively, our data suggest that early septic murine PMVEC barrier dysfunction driven by proinflammatory cytokines is not mediated through apoptosis, but PMVEC apoptosis contributes to late septic PMVEC barrier dysfunction.

  1. Circulating endothelial progenitor cells, microvascular density and fibrosis in obesity before and after bariatric surgery.

    Science.gov (United States)

    De Ciuceis, Carolina; Rossini, Claudia; Porteri, Enzo; La Boria, Elisa; Corbellini, Claudia; Mittempergher, Francesco; Di Betta, Ernesto; Petroboni, Beatrice; Sarkar, Annamaria; Agabiti-Rosei, Claudia; Casella, Claudio; Nascimbeni, Riccardo; Rezzani, Rita; Rodella, Luigi F; Bonomini, Francesca; Agabiti-Rosei, Enrico; Rizzoni, Damiano

    2013-06-01

    It is not known whether, in obesity, the capillary density or the number of circulating endothelial progenitor cells (EPCs) are reduced, or whether fibrosis of small vessels is also present. In addition, possible effects of weight reduction on these parameters have never been evaluated. Therefore, we investigated EPCs and capillary density in 25 patients with severe obesity, all submitted to bariatric surgery, and in 18 normotensive lean subjects and 12 hypertensive lean patients as controls. All patients underwent a biopsy of subcutaneous fat during bariatric surgery. In five patients, a second biopsy was obtained after consistent weight loss, about 1 year later, during a surgical intervention for abdominoplasty. EPCs and capillary density were reduced in obesity, and EPCs were significantly increased after weight reduction. Vascular collagen content was clearly increased in obese patients. No significant difference in vascular collagen was observed between normotensive obese patients and hypertensive obese patients. After pronounced weight reduction, collagen content was nearly normalized. No difference in stress-strain relation was observed among groups or before and after weight loss. In conclusion, our data suggest that microvascular rarefaction occurs in obesity. EPCs were significantly reduced in obese patients. Pronounced weight loss induced by bariatric surgery seems to induce a significant improvement of EPC number, but not of capillary rarefaction. A pronounced fibrosis of subcutaneous small resistance arteries is present in obese patients, regardless of the presence of increased blood pressure values. Consistent weight loss induced by bariatric surgery may induce an almost complete regression of microvascular fibrosis.

  2. Rescue of Brain Function Using Tunneling Nanotubes Between Neural Stem Cells and Brain Microvascular Endothelial Cells.

    Science.gov (United States)

    Wang, Xiaoqing; Yu, Xiaowen; Xie, Chong; Tan, Zijian; Tian, Qi; Zhu, Desheng; Liu, Mingyuan; Guan, Yangtai

    2016-05-01

    Evidence indicates that neural stem cells (NSCs) can ameliorate cerebral ischemia in animal models. In this study, we investigated the mechanism underlying one of the neuroprotective effects of NSCs: tunneling nanotube (TNT) formation. We addressed whether the control of cell-to-cell communication processes between NSCs and brain microvascular endothelial cells (BMECs) and, particularly, the control of TNT formation could influence the rescue function of stem cells. In an attempt to mimic the cellular microenvironment in vitro, a co-culture system consisting of terminally differentiated BMECs from mice in a distressed state and NSCs was constructed. Additionally, engraftment experiments with infarcted mouse brains revealed that control of TNT formation influenced the effects of stem cell transplantation in vivo. In conclusion, our findings provide the first evidence that TNTs exist between NSCs and BMECs and that regulation of TNT formation alters cell function.

  3. Isolation and Culture of Human Microvascular endothelium for comparison of the morphological and molecular characteristics of Microvascular endothelial cells under normal gravity against simulated micro gravity

    Directory of Open Access Journals (Sweden)

    Tholcopiyan L

    2010-01-01

    Full Text Available BACKGROUND: Vascular endothelial cells play a major role in wound healing and also in growth of the tumors. Angiogenesis can be a target for treating diseases that are due to either poor vascularisation or decreased blood supply as in stroke, ulcers, heart disease, etc or abnormal and increased vasculature like in tumours. Application of specific compounds that may inhibit or induce the creation of new blood vessels in the body may help in the treatment of such diseases (1. Ex vivo generation of blood vessels may offer an excellent alternative to the synthetic valves that are being currently used in cardiology. Micro gravity also referred to, as weightlessness is not essentially zero gravity but rather minimal gravity. According to cell type, micro gravity causes variety of changes in proliferation and differentiation of cells while also affecting the migration of cells and cellular functions (2, 3. Siamwala et al from AUKBC have already studied the effects of microgravity on the microvascular endothelial cells from bovine lung and macrovascular endothelial cells from the bovine pulmonary artery. It was observed that the proliferation and migration of macrovascular endothelial cells were increased in microgravity (4, 5. Nitric oxide production was also studied and observed that microgravity treatment did not change nitric oxide production by microvascular endothelial cells (4OBJECTIVE: Isolation and Comparison of culture characteristics of Human microvascular endothelium cultured conventionally and in novel nanomaterial scaffold and further study the morphological and molecular characteristics of microvascular endothelial cells under normal gravity against simulated micro gravityMATERIALS AND METHODS: The human Omentum samples were obtained using surgical procedures after informed consent. The microvascular endothelial cells were isolated following the protocol described by Scott et al (6.The isolated cells were seeded in two groups; Group I

  4. Cultivation of Human Microvascular Endothelial Cells on Topographical Substrates to Mimic the Human Corneal Endothelium

    Directory of Open Access Journals (Sweden)

    Jie Shi Chua

    2013-03-01

    Full Text Available Human corneal endothelial cells have a limited ability to replicate in vivo and in vitro. Allograft transplantation becomes necessary when an accident or trauma results in excessive cell loss. The reconstruction of the cornea endothelium using autologous cell sources is a promising alternative option for therapeutic or in vitro drug testing applications. The native corneal endothelium rests on the Descemet’s membrane, which has nanotopographies of fibers and pores. The use of synthetic topographies mimics the native environment, and it is hypothesized that this can direct the behavior and growth of human microvascular endothelial cells (HMVECs to resemble the corneal endothelium. In this study, HMVECs are cultivated on substrates with micron and nano-scaled pillar and well topographies. Closely packed HMVEC monolayers with polygonal cells and well-developed tight junctions were formed on the topographical substrates. Sodium/potassium (Na+/K+ adenine triphosphatase (ATPase expression was enhanced on the microwells substrate, which also promotes microvilli formation, while more hexagonal-like cells are found on the micropillars samples. The data obtained suggests that the use of optimized surface patterning, in particular, the microtopographies, can induce HMVECs to adopt a more corneal endothelium-like morphology with similar barrier and pump functions. The mechanism involved in cell contact guidance by the specific topographical features will be of interest for future studies.

  5. Adhesive properties of Enterobacter sakazakii to human epithelial and brain microvascular endothelial cells

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    Pospischil Andreas

    2006-06-01

    Full Text Available Abstract Background Enterobacter sakazakii is an opportunistic pathogen that has been associated with sporadic cases and outbreaks causing meningitis, necrotizing enterocolitis and sepsis especially in neonates. However, up to now little is known about the mechanisms of pathogenicity in E. sakazakii. A necessary state in the successful colonization, establishment and ultimately production of disease by microbial pathogens is the ability to adhere to host surfaces such as mucous membranes, gastric and intestinal epithelial or endothelial tissue. This study examined for the first time the adherence ability of 50 E. sakazakii strains to the two epithelial cell lines HEp-2 and Caco-2, as well as the brain microvascular endothelial cell line HBMEC. Furthermore, the effects of bacterial culture conditions on the adherence behaviour were investigated. An attempt was made to characterize the factors involved in adherence. Results Two distinctive adherence patterns, a diffuse adhesion and the formation of localized clusters of bacteria on the cell surface could be distinguished on all three cell lines. In some strains, a mixture of both patterns was observed. Adherence was maximal during late exponential phase, and increased with higher MOI. The adhesion capacity of E. sakazakii to HBMEC cells was affected by the addition of blood to the bacteria growth medium. Mannose, hemagglutination, trypsin digestion experiments and transmission electron microscopy suggested that the adhesion of E. sakazakii to the epithelial and endothelial cells is mainly non-fimbrial based. Conclusion Adherence experiments show heterogeneity within different E. sakazakii strains. In agreement with studies on E. cloacae, we found no relationship between the adhesive capacities in E. sakazakii and the eventual production of specific fimbriae. Further studies will have to be carried out in order to determine the adhesin(s involved in the interaction of E. sakazakii with cells and to

  6. Roles of cyclooxygenase-2 in microvascular endothelial cell proliferation induced by basic fibroblast growth factor

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    Background The level of basic fibroblast growth factor (bFGF) increases rapidly after cerebral ischemia. However, the molecular mechanisms for the effects of bFGF on cerebral microvascular endothelial cells (cMVECs) have not yet been fully elucidated. In this study, a murine cMVEC line, bend.3, was employed to study the effects of bFGF on cyclooxygenase (COX) expression and its downstream effects in cMVECs. Methods After treatment with bFGF, RT-PCR and Western blotting analyses were carried out to evaluate the changes in COX-2 mRNA and protein expression, respectively. Ml-r assays were performed to measure cell proliferation. The prostaglandin E2 (PGE2) and vascular endothelial growth factor (VEGF) concentrations in the culture medium were measured by enzyme-linked immunosorbent assay (ELISA). Results COX-2 mRNA and protein expressions in bEnd.3 cells were induced by bFGF in time- and dose-dependent manners. The bFGF-induced COX-2 upregulation led to enhanced PGE2 production by bEnd.3 cells, and this effect was abolished by the selective COX-2 inhibitor NS-398. bFGF also increased VEGF production by bend.3 cells, and this effect was blocked by NS-398 and the EP1/2 (PGE2 receptors) antagonist AH6809. Furthermore, exogenous PGE2 increased VEGF production in bend.3 cells, and AH6809 blocked this effect. Conclusion bFGF increases VEGF production in an autocrine manner by increasing COX-2-generated PGE2 in cMVECs and subsequently stimulates MVEC proliferation and angiogenesis.

  7. Transplanted microvascular endothelial cells promote oligodendrocyte precursor cell survival in ischemic demyelinating lesions.

    Science.gov (United States)

    Iijima, Keiya; Kurachi, Masashi; Shibasaki, Koji; Naruse, Masae; Puentes, Sandra; Imai, Hideaki; Yoshimoto, Yuhei; Mikuni, Masahiko; Ishizaki, Yasuki

    2015-11-01

    We previously showed that transplantation of brain microvascular endothelial cells (MVECs) greatly stimulated remyelination in the white matter infarct of the internal capsule (IC) induced by endothelin-1 injection and improved the behavioral outcome. In the present study, we examined the effect of MVEC transplantation on the infarct volume using intermittent magnetic resonance image and on the behavior of oligodendrocyte lineage cells histochemically. Our results in vivo show that MVEC transplantation reduced the infarct volume in IC and apoptotic death of oligodendrocyte precursor cells (OPCs). These results indicate that MVECs have a survival effect on OPCs, and this effect might contribute to the recovery of the white matter infarct. The conditioned-medium from cultured MVECs reduced apoptosis of cultured OPCs, while the conditioned medium from cultured fibroblasts did not show such effect. These results suggest a possibility that transplanted MVECs increased the number of OPCs through the release of humoral factors that prevent their apoptotic death. Identification of such humoral factors may lead to the new therapeutic strategy against ischemic demyelinating diseases.

  8. Brain microvascular endothelial cell association and distribution of a 5 nm ceria engineered nanomaterial

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    Dan M

    2012-07-01

    with the capillary fraction. Electron microscopy showed the ceria ENM located on the endothelial cell luminal surface.Conclusion: Ceria ENM association with brain capillary endothelial cells saturated between 20 and 60 seconds and ceria ENM brain uptake was not diffusion-mediated. During the 120-second ceria ENM perfusion, ceria ENM predominately associated with the surface of the brain capillary cells, providing the opportunity for its cell uptake or redistribution back into circulating blood.Keywords: ceria engineered nanomaterial, brain microvascular endothelial cell association, in situ brain perfusion, capillary depletion

  9. Cardiotoxic drugs Herceptin and doxorubicin inhibit cardiac microvascular endothelial cell barrier formation resulting in increased drug permeability

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    Emma L. Wilkinson

    2016-10-01

    Full Text Available Cardiotoxicity induced by anti-cancer therapeutics is a severe, and potentially fatal, adverse reaction of the heart in response to certain drugs. Current in vitro approaches to assess cardiotoxicity have focused on analysing cardiomyocytes. More recently it has become apparent that non-cardiomyocyte cells of the heart can potentially contribute to cardiotoxicity. Herceptin and doxorubicin are known to induce cardiotoxicity in the clinic. The effect of these drugs on the endothelial tight junction barrier was tested by analysing tight junction formation and zona occludens-1 (ZO-1 levels, revealing that Herceptin and doxorubicin are able to induce barrier perturbment and decrease barrier function in human cardiac microvascular endothelial cells (HCMECs leading to increased permeability. Herceptin treatment had no effect on the tight junction barrier function in human dermal and human brain microvascular endothelial cells. HCMECs showed detectable levels of HER2 compared with the other endothelial cells suggesting that Herceptin binding to HER2 in these cells may interfere with tight junction formation. Our data suggests that doxorubicin and Herceptin can affect tight junction formation in the cardiac microvasculature leading to increased drug permeability and adverse effects on the cardiac myocytes.

  10. Cocaine inhibits store-operated Ca2+ entry in brain microvascular endothelial cells: critical role for sigma-1 receptors.

    Science.gov (United States)

    Brailoiu, G Cristina; Deliu, Elena; Console-Bram, Linda M; Soboloff, Jonathan; Abood, Mary E; Unterwald, Ellen M; Brailoiu, Eugen

    2016-01-01

    Sigma-1 receptor (Sig-1R) is an intracellular chaperone protein with many ligands, located at the endoplasmic reticulum (ER). Binding of cocaine to Sig-1R has previously been found to modulate endothelial functions. In the present study, we show that cocaine dramatically inhibits store-operated Ca(2+) entry (SOCE), a Ca(2+) influx mechanism promoted by depletion of intracellular Ca(2+) stores, in rat brain microvascular endothelial cells (RBMVEC). Using either Sig-1R shRNA or pharmacological inhibition with the unrelated Sig-1R antagonists BD-1063 and NE-100, we show that cocaine-induced SOCE inhibition is dependent on Sig-1R. In addition to revealing new insight into fundamental mechanisms of cocaine-induced changes in endothelial function, these studies indicate an unprecedented role for Sig-1R as a SOCE inhibitor.

  11. Human microvascular endothelial cell toxicity caused by Brazilian purpuric fever-associated strains of Haemophilus influenzae biogroup aegyptius.

    Science.gov (United States)

    Weyant, R S; Quinn, F D; Utt, E A; Worley, M; George, V G; Candal, F J; Ades, E W

    1994-02-01

    An in vitro cytotoxicity model that uses an immortalized human microvascular endothelial cell line (HMEC-1) differentiates Brazilian purpuric fever (BPF)-associated Haemophilus influenzae biogroup aegyptius (HAE) strains from non-BPF-associated HAE strains. Toxic strains produced a characteristic HMEC-1 phenotype at an MOI of 1000 bacteria/TCC to produce an observable effect. The cytotoxic phenotype was characterized by the presence of large clumps of HMEC-1 cells, which detached from the monolayer within 48 h of inoculation by HAE cells. The cytotoxic phenotype was observed with 100% of BPF-associated HAE (40/40) and 14% of non-BPF-associated HAE (8/57; P < .001). The ability to study a BPF-associated phenotype in vitro using human microvascular cells should enhance our knowledge of BPF pathogenesis.

  12. Effect of curcumin on the adhesion of platelets to brain microvascular endothelial cells in vitro

    Institute of Scientific and Technical Information of China (English)

    Li ZHANG; Zhen-lun GU; Zheng-hong QIN; Zhong-qin LIANG

    2008-01-01

    Aim: To determine whether curcumin prevents the adhesion of platelets to brain microvascular endothelial cells (BMECs) cultured in vitro. Methods: [3H]Ad-chine-labeled platelets were incubated with BMECs to investigate the role of curcumin in the adhesion of platelets to BMECs. The number of platelets adher-ing to the BMECs monolayer was determined by liquid scintillation spectroscopy. The thrombin-induced expression of platelets P-selectin, glycoprotein Ⅱb (GPⅡb), and glycoprotein Ⅲa (GPⅢa) on the cell surface, was measured by flow cytometry. P-selectin mRNA levels of BMECs were determined by RT-PCR. The TNF-α-induced expressions of P-selectin and E-selectin on the surface of BMECs were determined by Western blotting. Results: The adhesion between thrombin-acti-vated platelets and normal BMECs, and that of TNF-α-activated BMECs and normal platelets were significantly increased, and this increase could be inhibited by curcumin (30-240 μmol/L) in a concentration-dependant manner. The platelets activated with thrombin and BMECs stimulated by TNF-α demonstrated an upregulated expressions of P-selectin and E-selectin, and this increase, when pretreated with curcumin for 30 min, could be restrained dose dependently. Curcumin also inhibited the increase of the GPⅡb/GPⅢa expression of thrombin-activated platelets in a concentration-dependent manner. Conclusion: Curcumin can inhibit the platelets to BMECs. This effect may be related to the decreased expressions of P-selectin, E-selectin, and GPⅡb/GPⅢa on platelets and BMECs.

  13. Protease activated receptor signaling is required for African trypanosome traversal of human brain microvascular endothelial cells.

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    Dennis J Grab

    Full Text Available BACKGROUND: Using human brain microvascular endothelial cells (HBMECs as an in vitro model for how African trypanosomes cross the human blood-brain barrier (BBB we recently reported that the parasites cross the BBB by generating calcium activation signals in HBMECs through the activity of parasite cysteine proteases, particularly cathepsin L (brucipain. In the current study, we examined the possible role of a class of protease stimulated HBMEC G protein coupled receptors (GPCRs known as protease activated receptors (PARs that might be implicated in calcium signaling by African trypanosomes. METHODOLOGY/PRINCIPAL FINDINGS: Using RNA interference (RNAi we found that in vitro PAR-2 gene (F2RL1 expression in HBMEC monolayers could be reduced by over 95%. We also found that the ability of Trypanosoma brucei rhodesiense to cross F2RL1-silenced HBMEC monolayers was reduced (39%-49% and that HBMECs silenced for F2RL1 maintained control levels of barrier function in the presence of the parasite. Consistent with the role of PAR-2, we found that HBMEC barrier function was also maintained after blockade of Galpha(q with Pasteurella multocida toxin (PMT. PAR-2 signaling has been shown in other systems to have neuroinflammatory and neuroprotective roles and our data implicate a role for proteases (i.e. brucipain and PAR-2 in African trypanosome/HBMEC interactions. Using gene-profiling methods to interrogate candidate HBMEC pathways specifically triggered by brucipain, several pathways that potentially link some pathophysiologic processes associated with CNS HAT were identified. CONCLUSIONS/SIGNIFICANCE: Together, the data support a role, in part, for GPCRs as molecular targets for parasite proteases that lead to the activation of Galpha(q-mediated calcium signaling. The consequence of these events is predicted to be increased permeability of the BBB to parasite transmigration and the initiation of neuroinflammation, events precursory to CNS disease.

  14. BMPRII influences the response of pulmonary microvascular endothelial cells to inflammatory mediators.

    Science.gov (United States)

    Vengethasamy, Leanda; Hautefort, Aurélie; Tielemans, Birger; Belge, Catharina; Perros, Frédéric; Verleden, Stijn; Fadel, Elie; Van Raemdonck, Dirk; Delcroix, Marion; Quarck, Rozenn

    2016-11-01

    Mutations in the bone morphogenetic protein receptor (BMPR2) gene have been observed in 70 % of patients with heritable pulmonary arterial hypertension (HPAH) and in 11-40 % with idiopathic PAH (IPAH). However, carriers of a BMPR2 mutation have only 20 % risk of developing PAH. Since inflammatory mediators are increased and predict survival in PAH, they could act as a second hit inducing the development of pulmonary hypertension in BMPR2 mutation carriers. Our specific aim was to determine whether inflammatory mediators could contribute to pulmonary vascular cell dysfunction in PAH patients with and without a BMPR2 mutation. Pulmonary microvascular endothelial cells (PMEC) and arterial smooth muscle cells (PASMC) were isolated from lung parenchyma of transplanted PAH patients, carriers of a BMPR2 mutation or not, and from lobectomy patients or lung donors. The effects of CRP and TNFα on mitogenic activity, adhesiveness capacity, and expression of adhesion molecules were investigated in PMECs and PASMCs. PMECs from BMPR2 mutation carriers induced an increase in PASMC mitogenic activity; moreover, endothelin-1 secretion by PMECs from carriers was higher than by PMECs from non-carriers. Recruitment of monocytes by PMECs isolated from carriers was higher compared to PMECs from non-carriers and from controls, with an elevated ICAM-1 expression. CRP increased adhesion of monocytes to PMECs in carriers and non-carriers, and TNFα only in carriers. PMEC from BMPR2 mutation carriers have enhanced adhesiveness for monocytes in response to inflammatory mediators, suggesting that BMPR2 mutation could generate susceptibility to an inflammatory insult in PAH.

  15. Glucocorticoids Significantly Influence the Transcriptome of Bone Microvascular Endothelial Cells of Human Femoral Head

    Institute of Scientific and Technical Information of China (English)

    Qing-Sheng Yu; Wan-Shou Guo; Li-Ming Cheng; Yu-Feng Lu; Jian-Ying Shen; Ping Li

    2015-01-01

    Background:Appropriate expression and regulation of the transcriptome,which mainly comprise ofmRNAs and lncRNAs,are important for all biological and cellular processes including the physiological activities of bone microvascular endothelial cells (BMECs).Through an intricate intracellular signaling systems,the transcriptome regulates the pharmacological response of the cells.Although studies have elucidated the impact of glucocorticoids (GCs) cell-specific gene expression signatures,it remains necessary to comprehensively characterize the impact of lncRNAs to transcriptional changes.Methods:BMECs were divided into two groups.One was treated with GCs and the other left untreated as a paired control.Differential expression was analyzed with GeneSpring software V12.0 (Agilent,Santa Clara,CA,USA) and hierarchical clustering was conducted using Cluster 3.0 software.The Gene Ontology (GO) analysis was performed with Molecular Annotation System provided by CapitalBio Corporation.Results:Our results highlight the involvement of genes implicated in development,differentiation and apoptosis following GC stimulation.Elucidation of differential gene expression emphasizes the importance of regulatory gene networks induced by GCs.We identified 73 up-regulated and 166 down-regulated long noncoding RNAs,the expression of 107 of which significantly correlated with 172 mRNAs induced by hydrocortisone.Conclusions:Transcriptome analysis of BMECs from human samples was performed to identify specific gene networks induced by GCs.Our results identified complex RNA crosstalk underlying the pathogenesis of steroid-induced necrosis of femoral head.

  16. Human Brain Microvascular Endothelial Cells and Umbilical Vein Endothelial Cells Differentially Facilitate Leukocyte Recruitment and Utilize Chemokines for T Cell Migration

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    Shumei Man

    2008-01-01

    Full Text Available Endothelial cells that functionally express blood brain barrier (BBB properties are useful surrogates for studying leukocyte-endothelial cell interactions at the BBB. In this study, we compared two different endothelial cellular models: transfected human brain microvascular endothelial cells (THBMECs and human umbilical vein endothelial cells (HUVECs. With each grow under optimal conditions, confluent THBMEC cultures showed continuous occludin and ZO-1 immunoreactivity, while HUVEC cultures exhibited punctate ZO-1 expression at sites of cell-cell contact only. Confluent THBMEC cultures on 24-well collagen-coated transwell inserts had significantly higher transendothelial electrical resistance (TEER and lower solute permeability than HUVECs. Confluent THBMECs were more restrictive for mononuclear cell migration than HUVECs. Only THBMECs utilized abluminal CCL5 to facilitate T-lymphocyte migration in vitro although both THBMECs and HUVECs employed CCL3 to facilitate T cell migration. These data establish baseline conditions for using THBMECs to develop in vitro BBB models for studying leukocyte-endothelial interactions during neuroinflammation.

  17. Galectin-1 suppresses methamphetamine induced neuroinflammation in human brain microvascular endothelial cells: Neuroprotective role in maintaining blood brain barrier integrity.

    Science.gov (United States)

    Parikh, Neil U; Aalinkeel, R; Reynolds, J L; Nair, B B; Sykes, D E; Mammen, M J; Schwartz, S A; Mahajan, S D

    2015-10-22

    Methamphetamine (Meth) abuse can lead to the breakdown of the blood-brain barrier (BBB) integrity leading to compromised CNS function. The role of Galectins in the angiogenesis process in tumor-associated endothelial cells (EC) is well established; however no data are available on the expression of Galectins in normal human brain microvascular endothelial cells and their potential role in maintaining BBB integrity. We evaluated the basal gene/protein expression levels of Galectin-1, -3 and -9 in normal primary human brain microvascular endothelial cells (BMVEC) that constitute the BBB and examined whether Meth altered Galectin expression in these cells, and if Galectin-1 treatment impacted the integrity of an in-vitro BBB. Our results showed that BMVEC expressed significantly higher levels of Galectin-1 as compared to Galectin-3 and -9. Meth treatment increased Galectin-1 expression in BMVEC. Meth induced decrease in TJ proteins ZO-1, Claudin-3 and adhesion molecule ICAM-1 was reversed by Galectin-1. Our data suggests that Galectin-1 is involved in BBB remodeling and can increase levels of TJ proteins ZO-1 and Claudin-3 and adhesion molecule ICAM-1 which helps maintain BBB tightness thus playing a neuroprotective role. Galectin-1 is thus an important regulator of immune balance from neurodegeneration to neuroprotection, which makes it an important therapeutic agent/target in the treatment of drug addiction and other neurological conditions.

  18. Quercetin protects human brain microvascular endothelial cells from fibrillar β-amyloid1–40-induced toxicity

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    Yongjie Li

    2015-01-01

    Full Text Available Amyloid beta-peptides (Aβ are known to undergo active transport across the blood-brain barrier, and cerebral amyloid angiopathy has been shown to be a prominent feature in the majority of Alzheimer׳s disease. Quercetin is a natural flavonoid molecule and has been demonstrated to have potent neuroprotective effects, but its protective effect on endothelial cells under Aβ-damaged condition is unclear. In the present study, the protective effects of quercetin on brain microvascular endothelial cells injured by fibrillar Aβ1–40 (fAβ1–40 were observed. The results show that fAβ1–40-induced cytotoxicity in human brain microvascular endothelial cells (hBMECs can be relieved by quercetin treatment. Quercetin increases cell viability, reduces the release of lactate dehydrogenase, and relieves nuclear condensation. Quercetin also alleviates intracellular reactive oxygen species generation and increases superoxide dismutase activity. Moreover, it strengthens the barrier integrity through the preservation of the transendothelial electrical resistance value, the relief of aggravated permeability, and the increase of characteristic enzyme levels after being exposed to fAβ1–40. In conclusion, quercetin protects hBMECs from fAβ1–40-induced toxicity.

  19. Exosomal signaling during hypoxia mediates microvascular endothelial cell migration and vasculogenesis.

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    Carlos Salomon

    Full Text Available Vasculogenesis and angiogenesis are critical processes in fetal circulation and placental vasculature development. Placental mesenchymal stem cells (pMSC are known to release paracrine factors (some of which are contained within exosomes that promote angiogenesis and cell migration. The aims of this study were: to determine the effects of oxygen tension on the release of exosomes from pMSC; and to establish the effects of pMSC-derived exosomes on the migration and angiogenic tube formation of placental microvascular endothelial cells (hPMEC. pMSC were isolated from placental villi (8-12 weeks of gestation, n = 6 and cultured under an atmosphere of 1%, 3% or 8% O2. Cell-conditioned media were collected and exosomes (exo-pMSC isolated by differential and buoyant density centrifugation. The dose effect (5-20 µg exosomal protein/ml of pMSC-derived exosomes on hPMEC migration and tube formation were established using a real-time, live-cell imaging system (Incucyte™. The exosome pellet was resuspended in PBS and protein content was established by mass spectrometry (MS. Protein function and canonical pathways were identified using the PANTHER program and Ingenuity Pathway Analysis, respectively. Exo-pMSC were identified, by electron microscopy, as spherical vesicles, with a typical cup-shape and diameters around of 100 nm and positive for exosome markers: CD63, CD9 and CD81. Under hypoxic conditions (1% and 3% O2 exo-pMSC released increased by 3.3 and 6.7 folds, respectively, when compared to the controls (8% O2; p<0.01. Exo-pMSC increased hPMEC migration by 1.6 fold compared to the control (p<0.05 and increased hPMEC tube formation by 7.2 fold (p<0.05. MS analysis identified 390 different proteins involved in cytoskeleton organization, development, immunomodulatory, and cell-to-cell communication. The data obtained support the hypothesis that pMSC-derived exosomes may contribute to placental vascular adaptation to low oxygen tension under both

  20. Effects of Neospora caninum infection on brain microvascular endothelial cells bioenergetics

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    Elsheikha Hany M

    2013-01-01

    Full Text Available Abstract Background The brain is the most commonly affected organ during Neospora caninum infection but the mechanisms utilized by this protozoan parasite for traversal of the blood–brain barrier (BBB are not yet understood. Herein, we investigate the cellular pathogenicity of N. caninum infection on bioenergetics of human brain microvascular endothelial cells (HBMECs, a fundamental component of the BBB. Methods We tracked the growth kinetics of N. caninum in HBMECs. Focusing on cell bioenergetics, oxygen consumption rate (OCR was determined using Clark electrode system and mitochondrial membrane potential (ΔΨm was evaluated using DePsipher staining by fluorescence microscopy in the presence and absence of infection. Results HBMECs provided a receptive environment for parasite proliferation. N. caninum tachyzoites were able to invade and replicate within HBMECs without significantly altering cell proliferation rate, as measured with the MTT assay, up to 24 hr post infection (pi. The oxygen consumption rate (OCR was significantly inhibited (p 6 cell min-1 and from −0.29±0.09 to −0.16±0.1 nmol 106 cell min-1 for uninfected HBMECs and free N. Caninum tachyzoites, respectively]. After normalization for DNA content the basal OCR did not differ between two host cell types: HBMECs and K562. The OCR of HBMECs was significantly elevated 24 hr pi in the absence of substrate, in 10 mM glucose and in the presence of a tetramethyl-p-phenylenediamine (TMPD/ascorbate redox shuttle. Although quantitatively similar results were observed for uninfected K562 cells, there was no effect on their OCR 24 hr pi with N. caninum under any of the above substrate conditions. 6mM azide abolished OCR in all situations. Mitochondrial staining with DePsipher indicated no change in their membrane potential (Δψm up to 24 hr pi. Conclusions N. caninum is able to grow in HBMECs without markedly disrupting their normal proliferation or mitochondrial integrity. However

  1. Influenza H5N1 virus infection of polarized human alveolar epithelial cells and lung microvascular endothelial cells

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    Yuen Kit M

    2009-10-01

    Full Text Available Abstract Background Highly pathogenic avian influenza (HPAI H5N1 virus is entrenched in poultry in Asia and Africa and continues to infect humans zoonotically causing acute respiratory disease syndrome and death. There is evidence that the virus may sometimes spread beyond respiratory tract to cause disseminated infection. The primary target cell for HPAI H5N1 virus in human lung is the alveolar epithelial cell. Alveolar epithelium and its adjacent lung microvascular endothelium form host barriers to the initiation of infection and dissemination of influenza H5N1 infection in humans. These are polarized cells and the polarity of influenza virus entry and egress as well as the secretion of cytokines and chemokines from the virus infected cells are likely to be central to the pathogenesis of human H5N1 disease. Aim To study influenza A (H5N1 virus replication and host innate immune responses in polarized primary human alveolar epithelial cells and lung microvascular endothelial cells and its relevance to the pathogenesis of human H5N1 disease. Methods We use an in vitro model of polarized primary human alveolar epithelial cells and lung microvascular endothelial cells grown in transwell culture inserts to compare infection with influenza A subtype H1N1 and H5N1 viruses via the apical or basolateral surfaces. Results We demonstrate that both influenza H1N1 and H5N1 viruses efficiently infect alveolar epithelial cells from both apical and basolateral surface of the epithelium but release of newly formed virus is mainly from the apical side of the epithelium. In contrast, influenza H5N1 virus, but not H1N1 virus, efficiently infected polarized microvascular endothelial cells from both apical and basolateral aspects. This provides a mechanistic explanation for how H5N1 virus may infect the lung from systemic circulation. Epidemiological evidence has implicated ingestion of virus-contaminated foods as the source of infection in some instances and our

  2. Dysfunction of microvascular endothelial cells induced by TNFα and its molecular mechanism

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    Microvascular endothelial cell (MVEC) is one of the target cells of TNFα (TNF effect). The dysfunction of MVEC induced by TNF plays an important role in some cardio-cerebral vascular diseases. ① Cell proliferation kinetic: Using flow cytometry, we found cell count [(4.30±0.34)×107/L)] in TNF group (4×105 U/L) was obviously less than that in control [(5.23±0.50)×107/L, P<0.01]. The cells of G1 phase were more than those of the control, while the cells of G2, S and M phase became less (P<0.05). ② Coagulant and anticoagulant: 72 h after MVEC cultued in the media, the content of 6-keto-PGF1α (RIA) and activity of PAI decreased significantly in TNF (4×105 U/L) group (P<0.01, vs control). The difference between TXB2 content and t-PA activity in groups was not significant (P>0.05). ③ Adhesive molecule: The effect of low concentration TNF (<4×105 U/L) on adhesion between cultured MVEC and leukocytes was not signficant, but when the concentration of TNF reached 8×105 U/L or more, 12 h after culture the adhesion rate between MVEC and neutrophil increased 30.8%±4.5%. If adding monoclonal antibody of ICAM-1/CD11 into media, the adhesion rate of leukocytes decreased significantly (from 31.2% to 63.4%). ④ NO: The level of nitrite in culture media (Griess reaction) was higher than that of control (P<0.05) after pretreatment of TNF (2×106 U/L) for 6 h. Adding L-NMMA, Dexamethasone or Cycloheximide in media could block the increase of nitrite induced by TNF, while L-Arg could enhance it. The expression of iNOS mRNA of PMVEC increased significantly after treated with TNF (2×106 U/L) for 24 h (quantitative RT/PCR). Pretreatment with Dexamethasone or Cycloheximide could block the increase (P<0.05). Meanwhile, the expression of eNOS mRNA decreased significantly compared with control, the decrease can be blocked by Cycloheximide but not by Dexamethasone. So that TNF can induce the expression of iNOS mRNA in PMVEC, but inhibited the

  3. Hypoxia inducible factor-1alpha mediates protection of DL-3-n-butylphthalide in brain microvascular endothelial cells against oxygen glucose deprivation-induced injury

    Institute of Scientific and Technical Information of China (English)

    Weihong Yang; Ling Li; Ruxun Huang; Zhong Pei; Songjie Liao; Jinsheng Zeng

    2012-01-01

    Studies have demonstrated that DL-3-n-butylphthalide can significantly alleviate oxygen glucose deprivation-induced injury of human umbilical vein endothelial cells at least partly associated with its enhancement on oxygen glucose deprivation -induced hypoxia inducible factor-1α expression. In this study, we hypothesized that DL-3-n-butylphthalide can protect against oxygen glucose deprivation-induced injury of newborn rat brain microvascular endothelial cells by means of upregulating hypoxia inducible factor-1α expression. MTT assay and Hoechst staining results showed that DL-3-n-butylphthalide protected brain microvascular endothelial cells against oxygen glucose deprivation-induced injury in a dose-dependent manner. Western blot and immunofluorescent staining results further confirmed that the protective effect was related to upregulation of hypoxia inducible factor-1α. Real-time RT-PCR reaction results showed that DL-3-n-butylphthalide reduced apoptosis by inhibiting downregulation of pro-apoptotic gene caspase-3 mRNA expression and upregulation of apoptosis-executive protease bcl-2 mRNA expression; however, DL-3-n-butylphthalide had no protective effects on brain microvascular endothelial cells after knockdown of hypoxia inducible factor-1α by small interfering RNA. These findings suggest that DL-3-n-butylphthalide can protect brain microvascular endothelial cells against oxygen glucose deprivation-induced injury by upregulating bcl-2 expression and downregulating caspase-3 expression though hypoxia inducible factor-1α pathway.

  4. Comparison of skin microvascular reactivity with hemostatic markers of endothelial dysfunction and damage in type 2 diabetes

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    Sandra Beer

    2008-12-01

    Full Text Available Sandra Beer1,2, François Feihl1, Juan Ruiz2, Irène Juhan-Vague3, Marie-Françoise Aillaud3, Sandrine Golay Wetzel1, Lucas Liaudet4, Rolf C Gaillard2, Bernard Waeber1Centre Hospitalier Universitaire Vaudois, Division de Physiopathologie Clinique, Lausanne, Suisse1Division de Physiopathologie Clinique, Centre Hospitalier Universitaire Vaudois et Université de Lausanne, Lausanne, Suisse; 2Service d’Endocrinologie, de Diabétologie et de Métabolisme, Centre Hospitalier Universitaire Vaudois et Université de Lausanne, Lausanne, Suisse; 3Laboratoire d’hématologie, Centre Hospitalier Universitaire de Marseille; Inserm UMR 626, Marseille, France; 4Service de Médecine Intensive de l’Adulte, Centre Hospitalier Universitaire Vaudois et Université de Lausanne, Lausanne, SuisseAim: Patients with non-insulin-dependent diabetes mellitus (NIDDM are at increased cardiovascular risk due to an accelerated atherosclerotic process. The present study aimed to compare skin microvascular function, pulse wave velocity (PWV, and a variety of hemostatic markers of endothelium injury [von Willebrand factor (vWF, plasminogen activator inhibitor-1 (PAI-1, tissue plasminogen activator (t-PA, tissue factor pathway inhibitor (TFPI, and the soluble form of thrombomodulin (s-TM] patients with NIDDM.Methods: 54 patients with NIDDM and 38 sex- and age-matched controls were studied. 27 diabetics had no overt micro- and/or macrovascular complications, while the remainder had either or both. The forearm skin blood flow was assessed by laser-Doppler imaging, which allowed the measurement of the response to iontophoretically applied acetylcholine (endotheliumdependent vasodilation and sodium nitroprusside (endothelium-independent vasodilation, as well as the reactive hyperemia triggered by the transient occlusion of the circulation.Results: Both endothelial and non-endothelial reactivity were significantly blunted in diabetics, regardless of the presence or the absence of

  5. Decreased expression of serum and microvascular vascular endothelial growth factor receptor-2 in meningococcal sepsis*.

    NARCIS (Netherlands)

    Flier, M. van der; Baerveldt, E.M.; Miedema, A.; Hartwig, N.G.; Hazelzet, J.A.; Emonts, M.; Groot, R. de; Prens, E.P.; Vught, A.J. van; Jansen, N.J.

    2013-01-01

    OBJECTIVES: To determine the skin microvessel expression of vascular endothelial growth factor receptor 2 and serum-soluble vascular endothelial growth factor receptor 2 levels in children with meningococcal sepsis. DESIGN: Observational study. SETTING: Two tertiary academic children hospital PICUs.

  6. Human Brain Microvascular Endothelial Cells Derived from the BC1 iPS Cell Line Exhibit a Blood-Brain Barrier Phenotype

    OpenAIRE

    Katt, Moriah E.; Xu, Zinnia S.; Gerecht, Sharon; Searson, Peter C.

    2016-01-01

    The endothelial cells that form capillaries in the brain are highly specialized, with tight junctions that minimize paracellular transport and an array of broad-spectrum efflux pumps that make drug delivery to the brain extremely challenging. One of the major limitations in blood-brain barrier research and the development of drugs to treat central nervous system diseases is the lack of appropriate cell lines. Recent reports indicate that the derivation of human brain microvascular endothelial...

  7. Anti inflammatory and anti angiogenic effect of black raspberry extract on human esophageal and intestinal microvascular endothelial cells.

    Science.gov (United States)

    Medda, Rituparna; Lyros, Orestis; Schmidt, Jamie L; Jovanovic, Nebojsa; Nie, Linghui; Link, Benjamin J; Otterson, Mary F; Stoner, Gary D; Shaker, Reza; Rafiee, Parvaneh

    2015-01-01

    Polyphenolic compounds (anthocyanins, flavonoid glycosides) in berries prevent the initiation, promotion, and progression of carcinogenesis in rat's digestive tract and esophagus, in part, via anti-inflammatory pathways. Angiogenesis has been implicated in the pathogenesis of chronic inflammation and tumorigenesis. In this study, we investigated the anti-inflammatory and anti-angiogenic effects of black raspberry extract (BRE) on two organ specific primary human intestinal microvascular endothelial cells, (HIMEC) and human esophageal microvascular endothelial cells (HEMEC), isolated from surgically resected human intestinal and donor discarded esophagus, respectively. HEMEC and HIMEC were stimulated with TNF-α/IL-1β with or without BRE. The anti-inflammatory effects of BRE were assessed based upon COX-2, ICAM-1 and VCAM-1 gene and protein expression, PGE2 production, NFκB p65 subunit nuclear translocation as well as endothelial cell-leukocyte adhesion. The anti-angiogenic effects of BRE were assessed on cell migration, proliferation and tube formation following VEGF stimulation as well as on activation of Akt, MAPK and JNK signaling pathways. BRE inhibited TNF-α/IL-1β-induced NFκB p65 nuclear translocation, PGE2 production, up-regulation of COX-2, ICAM-1 and VCAM-1 gene and protein expression and leukocyte binding in HEMEC but not in HIMEC. BRE attenuated VEGF-induced cell migration, proliferation and tube formation in both HEMEC and HIMEC. The anti-angiogenic effect of BRE is mediated by inhibition of Akt, MAPK and JNK phosphorylations. BRE exerted differential anti-inflammatory effects between HEMEC and HIMEC following TNF-α/IL-1β activation whereas demonstrated similar anti-angiogenic effects following VEGF stimulation in both cell lines. These findings may provide more insight into the anti-tumorigenic capacities of BRE in human disease and cancer.

  8. Application of a Novel Microvascular Imaging Technique in Breast Lesion Evaluation.

    Science.gov (United States)

    Yongfeng, Zhao; Ping, Zhou; Wengang, Liu; Yang, Shao; Shuangming, Tian

    2016-09-01

    Conventional power Doppler imaging (PDI) and the novel Superb Microvascular Imaging (SMI) technique were applied to observe the distribution of microvessels in 135 breast lesions, using semi-quantitative grading, penetrating vessel evaluation and flow distribution pattern to evaluate diagnostic efficacy. Compared with PDI, SMI detected more flow signals and details of microvessels. Further, when a centrally distributed branching or diffusing mode was used as a criterion for diagnosing malignancy, SMI improved diagnosis of breast masses. Sensitivity, specificity, positive predictive value and negative predictive value of SMI-assessed flow distribution were 85.4%, 92.6%, 83.3% and 93.5%, respectively, compared with 70.7%, 92.6%, 80.5% and 87.9% for PDI. We also found that flow distribution pattern analysis is superior to semi-quantitative grading and the penetrating vessel method in differentiating malignant breast lesions. Our work here further supports SMI as a novel and promising technique in visualizing microvasculature in breast lesions that may be of paramount use in initial diagnosis as well as follow-up assessment in various treatment regimes.

  9. Morphine induces expression of platelet-derived growth factor in human brain microvascular endothelial cells: implication for vascular permeability.

    Directory of Open Access Journals (Sweden)

    Hongxiu Wen

    Full Text Available Despite the advent of antiretroviral therapy, complications of HIV-1 infection with concurrent drug abuse are an emerging problem. Morphine, often abused by HIV-infected patients, is known to accelerate neuroinflammation associated with HIV-1 infection. Detailed molecular mechanisms of morphine action however, remain poorly understood. Platelet-derived growth factor (PDGF has been implicated in a number of pathological conditions, primarily due to its potent mitogenic and permeability effects. Whether morphine exposure results in enhanced vascular permeability in brain endothelial cells, likely via induction of PDGF, remains to be established. In the present study, we demonstrated morphine-mediated induction of PDGF-BB in human brain microvascular endothelial cells, an effect that was abrogated by the opioid receptor antagonist-naltrexone. Pharmacological blockade (cell signaling and loss-of-function (Egr-1 approaches demonstrated the role of mitogen-activated protein kinases (MAPKs, PI3K/Akt and the downstream transcription factor Egr-1 respectively, in morphine-mediated induction of PDGF-BB. Functional significance of increased PDGF-BB manifested as increased breach of the endothelial barrier as evidenced by decreased expression of the tight junction protein ZO-1 in an in vitro model system. Understanding the regulation of PDGF expression may provide insights into the development of potential therapeutic targets for intervention of morphine-mediated neuroinflammation.

  10. Activation of sonic hedgehog signaling attenuates oxidized low-density lipoprotein-stimulated brain microvascular endothelial cells dysfunction in vitro.

    Science.gov (United States)

    Jiang, Xiu-Long; Chen, Ting; Zhang, Xu

    2015-01-01

    The study was performed to investigate the role of sonic hedgehog (SHH) in the oxidized low-density lipoprotein (oxLDL)-induced blood-brain barrier (BBB) disruption. The primary mouse brain microvascular endothelial cells (MBMECs) were exposed to oxLDL. The results indicated that treatment of MBMECs with oxLDL decreased the cell viability, and oxidative stress was involved in oxLDL-induce MBMECs dysfunction with increasing intracellular ROS and MDA formation as well as decreasing NO release and eNOS mRNA expression. In addition, SHH signaling components, such as SHH, Smo and Gli1, mRNA and protein levels were significantly decreased after incubation with increasing concentrations of oxLDL. Treatment with oxLDL alone or SHH loss-of-function significantly increased the permeability of MBMECs, and overexpression of SHH attenuated oxLDL-induced elevation of permeability in MBMECs. Furthermore, SHH gain-of-function could reverse oxLDL-induced apoptosis through inhibition caspase3 and caspase8 levels in MBMECs. Taken together, these results demonstrated that the suppression of SHH in MBMECs might contribute to the oxLDL-induced disruption of endothelial barrier. However, the overexpression of SHH could reverse oxLDL-induced endothelial cells dysfunction in vitro.

  11. Microvascularized 3D Breast Cancer Constructs for Drug Testing and Development

    Science.gov (United States)

    2014-10-01

    of collagen I, either bovine or rat tail , in solid cultures without microchannels/perfusion for 7 days. Viability (A) was assessed by...Buchanan showed that breast carcinoma cells embedded in rat tail collagen I and perfused through a central endothelialized channel (711 µm in...consisting of collagen I and Matrigel (basement membrane material (BM)), a rigid and porous structure to support the viscous ECM and the cells that

  12. Effect of penehyclidine hydrochloride on β-arrestin-1 expression in lipopolysaccharide-induced human pulmonary microvascular endothelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Zhan, J. [Department of Anesthesiology, Zhongnan Hospital, Wuhan University, Wuhan, Hubei (China); Xiao, F. [Department of Osteology, Pu Ai Hospital, Huazhong University of Science and Technology, Wuhan, Hubei (China); Zhang, Z.Z.; Wang, Y.P.; Chen, K.; Wang, Y.L. [Department of Anesthesiology, Zhongnan Hospital, Wuhan University, Wuhan, Hubei (China)

    2013-12-02

    β-arrestins are expressed proteins that were first described, and are well-known, as negative regulators of G protein-coupled receptor signaling. Penehyclidine hydrochloride (PHC) is a new anti-cholinergic drug that can inhibit biomembrane lipid peroxidation, and decrease cytokines and oxyradicals. However, to date, no reports on the effects of PHC on β-arrestin-1 in cells have been published. The aim of this study was to investigate the effect of PHC on β-arrestin-1 expression in lipopolysaccharide (LPS)-induced human pulmonary microvascular endothelial cells (HPMEC). Cultured HPMEC were pretreated with PHC, followed by LPS treatment. Muscarinic receptor mRNAs were assayed by real-time quantitative PCR. Cell viability was assayed by the methyl thiazolyl tetrazolium (MTT) conversion test. The dose and time effects of PHC on β-arrestin-1 expression in LPS-induced HPMEC were determined by Western blot analysis. Cell malondialdehyde (MDA) level and superoxide dismutase (SOD) activity were measured. It was found that the M{sub 3} receptor was the one most highly expressed, and was activated 5 min after LPS challenge. Furthermore, 2 μg/mL PHC significantly upregulated expression of β-arrestin-1 within 10 to 15 min. Compared with the control group, MDA levels in cells were remarkably increased and SOD activities were significantly decreased in LPS pretreated cells, while PHC markedly decreased MDA levels and increased SOD activities. We conclude that PHC attenuated ROS injury by upregulating β-arrestin-1 expression, thereby implicating a mechanism by which PHC may exert its protective effects against LPS-induced pulmonary microvascular endothelial cell injury.

  13. Functional expression of choline transporter like-protein 1 (CTL1) and CTL2 in human brain microvascular endothelial cells.

    Science.gov (United States)

    Iwao, Beniko; Yara, Miki; Hara, Naomi; Kawai, Yuiko; Yamanaka, Tsuyoshi; Nishihara, Hiroshi; Inoue, Takeshi; Inazu, Masato

    2016-02-01

    In this study, we examined the molecular and functional characterization of choline transporter in human brain microvascular endothelial cells (hBMECs). Choline uptake into hBMECs was a saturable process that was mediated by a Na(+)-independent, membrane potential and pH-dependent transport system. The cells have two different [(3)H]choline transport systems with Km values of 35.0 ± 4.9 μM and 54.1 ± 8.1 μM, respectively. Choline uptake was inhibited by choline, acetylcholine (ACh) and the choline analog hemicholinium-3 (HC-3). Various organic cations also interacted with the choline transport system. Choline transporter-like protein 1 (CTL1) and CTL2 mRNA were highly expressed, while mRNA for high-affinity choline transporter 1 (CHT1) and organic cation transporters (OCTs) were not expressed in hBMECs. CTL1 and CTL2 proteins were localized to brain microvascular endothelial cells in human brain cortical sections. Both CTL1 and CTL2 proteins were expressed on the plasma membrane and mitochondria. CTL1 and CTL2 proteins are mainly expressed in plasma membrane and mitochondria, respectively. We conclude that choline is mainly transported via an intermediate-affinity choline transport system, CTL1 and CTL2, in hBMECs. These transporters are responsible for the uptake of extracellular choline and organic cations. CTL2 participate in choline transport mainly in mitochondria, and may be the major site for the control of choline oxidation.

  14. Associations of Macro- and Microvascular Endothelial Dysfunction With Subclinical Ventricular Dysfunction in End-Stage Renal Disease.

    Science.gov (United States)

    Dubin, Ruth F; Guajardo, Isabella; Ayer, Amrita; Mills, Claire; Donovan, Catherine; Beussink, Lauren; Scherzer, Rebecca; Ganz, Peter; Shah, Sanjiv J

    2016-10-01

    Patients with end-stage renal disease (ESRD) suffer high rates of heart failure and cardiovascular mortality, and we lack a thorough understanding of what, if any, modifiable factors contribute to cardiac dysfunction in these high-risk patients. To evaluate endothelial function as a potentially modifiable cause of cardiac dysfunction in ESRD, we investigated cross-sectional associations of macro- and microvascular dysfunction with left and right ventricular dysfunction in a well-controlled ESRD cohort. We performed comprehensive echocardiography, including tissue Doppler imaging and speckle-tracking echocardiography of the left and right ventricle, in 149 ESRD patients enrolled in an ongoing prospective, observational study. Of these participants, 123 also underwent endothelium-dependent flow-mediated dilation of the brachial artery (macrovascular function). Microvascular function was measured as the velocity time integral of hyperemic blood flow after cuff deflation. Impaired flow-mediated dilation was associated with higher left ventricular mass, independently of age and blood pressure: per 2-fold lower flow-mediated dilation, left ventricular mass was 4.1% higher (95% confidence interval, 0.49-7.7; P=0.03). After adjustment for demographics, blood pressure, comorbidities, and medications, a 2-fold lower velocity time integral was associated with 9.5% higher E/e' ratio (95% confidence interval, 1.0-16; P=0.03) and 6.7% lower absolute right ventricular longitudinal strain (95% confidence interval, 2.0-12; P=0.003). Endothelial dysfunction is a major correlate of cardiac dysfunction in ESRD, particularly diastolic and right ventricular dysfunction, in patients whose volume status is well controlled. Future investigations are needed to determine whether therapies targeting the vascular endothelium could improve cardiac outcomes in ESRD.

  15. High glucose, glucose fluctuation and carbonyl stress enhance brain microvascular endothelial barrier dysfunction: Implications for diabetic cerebral microvasculature

    Directory of Open Access Journals (Sweden)

    Wei Li

    2015-08-01

    Full Text Available We previously demonstrated that in normal glucose (5 mM, methylglyoxal (MG, a model of carbonyl stress induced brain microvascular endothelial cell (IHEC dysfunction that was associated with occludin glycation and prevented by N-acetylcysteine (NAC. Herein, we investigated the impact of high glucose and low GSH, conditions that mimicked the diabetic state, on MG-induced IHEC dysfunction. MG-induced loss of transendothelial electrical resistance (TEER was potentiated in IHECs cultured for 7 or 12 days in 25 mM glucose (hyperglycemia; moreover, barrier function remained disrupted 6 h after cell transfer to normal glucose media (acute glycemic fluctuation. Notably, basal occludin glycation was elevated under these glycemic states. TEER loss was exaggerated by inhibition of glutathione (GSH synthesis and abrogated by NAC, which corresponded to GSH decreases and increases, respectively. Significantly, glyoxalase II activity was attenuated in hyperglycemic cells. Moreover, hyperglycemia and GSH inhibition increased MG accumulation, consistent with a compromised capacity for MG elimination. α-Oxoaldehydes (MG plus glyoxal levels were elevated in streptozotocin-induced diabetic rat plasma. Immunohistochemistry revealed a prevalence of MG-positive, but fewer occludin-positive microvessels in the diabetic brain in vivo, and Western analysis confirmed an increase in MG–occludin adducts. These results provide the first evidence that hyperglycemia and acute glucose fluctuation promote MG–occludin formation and exacerbate brain microvascular endothelial dysfunction. Low occludin expression and high glycated-occludin contents in diabetic brain in vivo are factors that would contribute to the dysfunction of the cerebral microvasculature during diabetes.

  16. Effects of the physicochemical properties of titanium dioxide nanoparticles, commonly used as sun protection agents, on microvascular endothelial cells

    Science.gov (United States)

    Strobel, Claudia; Torrano, Adriano A.; Herrmann, Rudolf; Malissek, Marcelina; Bräuchle, Christoph; Reller, Armin; Treuel, Lennart; Hilger, Ingrid

    2014-01-01

    Until now, the potential effects of titanium dioxide (TiO2) nanoparticles on endothelial cells are not well understood, despite their already wide usage. Therefore, the present work characterizes six TiO2 nanoparticle samples in the size range of 19 × 17 to 87 × 13 nm, which are commonly present in sun protection agents with respect to their physicochemical properties (size, shape, ζ-potential, agglomeration, sedimentation, surface coating, and surface area), their interactions with serum proteins and biological impact on human microvascular endothelial cells (relative cellular dehydrogenase activity, adenosine triphosphate content, and monocyte chemoattractant protein-1 release). We observed no association of nanoparticle morphology with the agglomeration and sedimentation behavior and no variations of the ζ-potential (-14 to -19 mV) in dependence on the surface coating. In general, the impact on endothelial cells was low and only detectable at concentrations of 100 μg/ml. Particles containing a rutile core and having rod-like shape had a stronger effect on cell metabolism than those with anatase core and elliptical shape (relative cellular dehydrogenase activity after 72 h: 60 vs. 90 %). Besides the morphology, the nanoparticle shell constitution was found to influence the metabolic activity of the cells. Upon cellular uptake, the nanoparticles were localized perinuclearly. Considering that in the in vivo situation endothelial cells would come in contact with considerably lower nanoparticle amounts than the lowest-observable adverse effects level (100 μg/ml), TiO2 nanoparticles can be considered as rather harmless to humans under the investigated conditions.

  17. The Diagnostic Value of Superb Microvascular Imaging (SMI) in Detecting Blood Flow Signals of Breast Lesions

    Science.gov (United States)

    Ma, Yan; Li, Gang; Li, Jing; Ren, Wei-dong

    2015-01-01

    Abstract The correlation between color Doppler flow imaging (CDFI) and Superb Microvascular Imaging (SMI) for detecting blood flow in breast lesions was investigated, as was the diagnostic value of SMI in differentiating benign from malignant breast lesions. These lesions were evaluated using both CDFI and SMI according to Adler's method. Pathologic examination showed 57 malignant lesions and 66 benign lesions. The number of blood vessels in a single mass was detected by 2 techniques (SMI and CDFI), and the difference between the 2 values (SMI-CDFI) was calculated. The optimal threshold for the diagnosis of malignant neoplasms and the diagnostic performances of SMI, CDFI, and SMI-CDFI were calculated. For the total lesions and malignant lesions alone, the difference between SMI and CDFI for detecting blood flow was significant (P < 0.01), but the difference was not significant for benign lesions (P = 0.15). The area under the receiver operating characteristic curve was 0.73 (95% confidence interval [CI]: 0.64–0.82) for CDFI; 0.81 (95% CI: 0.74–0.89) for SMI; and 0.89 (95% CI: 0.82–0.95) for SMI-CDFI. Furthermore, the modality of “SMI-CDFI” showed the best diagnostic performance. SMI provides further microvessel information in breast lesions. The diagnostic modality of “SMI-CDFI” can improve the diagnostic performance of ultrasound in the differentiation between benign and malignant masses. PMID:26356718

  18. Hsp90 inhibition suppresses NF-κB transcriptional activation via Sirt-2 in human lung microvascular endothelial cells.

    Science.gov (United States)

    Thangjam, Gagan S; Birmpas, Charalampos; Barabutis, Nektarios; Gregory, Betsy W; Clemens, Mary Ann; Newton, Joseph R; Fulton, David; Catravas, John D

    2016-05-15

    The ability of anti-heat shock protein 90 (Hsp90) drugs to attenuate NF-κB-mediated transcription is the major basis for their anti-inflammatory properties. While the molecular mechanisms underlying this effect are not clear, they appear to be distinct in human endothelial cells. We now show for the first time that type 2 sirtuin (Sirt-2) histone deacetylase binds human NF-κB target gene promoter and prevents the recruitment of NF-κB proteins and subsequent assembly of RNA polymerase II complex in human lung microvascular endothelial cells. Hsp90 inhibitors stabilize the Sirt-2/promoter interaction and impose a "transcriptional block," which is reversed by either inhibition or downregulation of Sirt-2 protein expression. Furthermore, this process is independent of NF-κB (p65) Lysine 310 deacetylation, suggesting that it is distinct from known Sirt-2-dependent mechanisms. We demonstrate that Sirt-2 is recruited to NF-κB target gene promoter via interaction with core histones. Upon inflammatory challenge, chromatin remodeling and core histone H3 displacement from the promoter region removes Sirt-2 and allows NF-κB/coactivator recruitment essential for RNA Pol II-dependent mRNA induction. This novel mechanism may have important implications in pulmonary inflammation.

  19. Iron oxide nanoparticles induce human microvascular endothelial cell permeability through reactive oxygen species production and microtubule remodeling

    Directory of Open Access Journals (Sweden)

    Shi Xianglin

    2009-01-01

    Full Text Available Abstract Background Engineered iron nanoparticles are being explored for the development of biomedical applications and many other industry purposes. However, to date little is known concerning the precise mechanisms of translocation of iron nanoparticles into targeted tissues and organs from blood circulation, as well as the underlying implications of potential harmful health effects in human. Results The confocal microscopy imaging analysis demonstrates that exposure to engineered iron nanoparticles induces an increase in cell permeability in human microvascular endothelial cells. Our studies further reveal iron nanoparticles enhance the permeability through the production of reactive oxygen species (ROS and the stabilization of microtubules. We also showed Akt/GSK-3β signaling pathways are involved in iron nanoparticle-induced cell permeability. The inhibition of ROS demonstrate ROS play a major role in regulating Akt/GSK-3β – mediated cell permeability upon iron nanoparticle exposure. These results provide new insights into the bioreactivity of engineered iron nanoparticles which can inform potential applications in medical imaging or drug delivery. Conclusion Our results indicate that exposure to iron nanoparticles induces an increase in endothelial cell permeability through ROS oxidative stress-modulated microtubule remodeling. The findings from this study provide new understandings on the effects of nanoparticles on vascular transport of macromolecules and drugs.

  20. Quantitative in vitro assay to measure neutrophil adhesion to activated primary human microvascular endothelial cells under static conditions.

    Science.gov (United States)

    Wilhelmsen, Kevin; Farrar, Katherine; Hellman, Judith

    2013-08-23

    The vascular endothelium plays an integral part in the inflammatory response. During the acute phase of inflammation, endothelial cells (ECs) are activated by host mediators or directly by conserved microbial components or host-derived danger molecules. Activated ECs express cytokines, chemokines and adhesion molecules that mobilize, activate and retain leukocytes at the site of infection or injury. Neutrophils are the first leukocytes to arrive, and adhere to the endothelium through a variety of adhesion molecules present on the surfaces of both cells. The main functions of neutrophils are to directly eliminate microbial threats, promote the recruitment of other leukocytes through the release of additional factors, and initiate wound repair. Therefore, their recruitment and attachment to the endothelium is a critical step in the initiation of the inflammatory response. In this report, we describe an in vitro neutrophil adhesion assay using calcein AM-labeled primary human neutrophils to quantitate the extent of microvascular endothelial cell activation under static conditions. This method has the additional advantage that the same samples quantitated by fluorescence spectrophotometry can also be visualized directly using fluorescence microscopy for a more qualitative assessment of neutrophil binding.

  1. Molecular and Cellular Characterization of Space Flight Effects on Microvascular Endothelial Cell Function - PreparatoryWork for the SFEF Project

    Science.gov (United States)

    Balsamo, Michele; Barravecchia, Ivana; Mariotti, Sara; Merenda, Alessandra; De Cesari, Chiara; Vukich, Marco; Angeloni, Debora

    2014-12-01

    Exposure to microgravity during space flight (SF) of variable length induces suffering of the endothelium (the cells lining all blood vessels), mostly responsible for health problems found in astronauts and animals returning from space. Of interest to pre-nosological medicine, the effects of microgravity on astronauts are strikingly similar to the consequences of sedentary life, senescence and degenerative diseases on Earth, although SF effects are accelerated and reversible. Thus, microgravity is a significant novel model for better understanding of common pathologies. A comprehensive cell and molecular biology study is needed in order to explain pathophysiological findings after SFs. This project will study the effects of microgravity and cosmic radiation on endothelial cells (ECs) cultured on the International Space Station through analysis of 1) cell transcriptome, 2) DNA methylome, 3) DNA damage and cell senescence, 4) variations in cell cycle and cell morphology. This project has been selected by the European Space Agency and the Italian Space Agency and is presently in preparation. The ground study presented here was performed to determine the biological and engineering requirements that will allow us to retrieve suitable samples after culturing, fixing and storing ECs in space. We expect to identify molecular pathways activated by space microgravity in microvascular ECs, which may shed light on pathogenic molecular mechanisms responsible for endothelial suffering shared by astronauts and individuals affected with aging, degenerative and sedentary life-associated pathologies on Earth.

  2. Prostaglandin F2-alpha receptor (FPr expression on porcine corpus luteum microvascular endothelial cells (pCL-MVECs

    Directory of Open Access Journals (Sweden)

    Forni Monica

    2007-07-01

    Full Text Available Abstract Background The corpus luteum (CL is a transient endocrine gland and prostaglandin F2-alpha is considered to be the principal luteolysin in pigs. In this species, the in vivo administration of prostaglandin F2-alpha induces apoptosis in large vessels as early as 6 hours after administration. The presence of the prostaglandin F2-alpha receptor (FPr on the microvascular endothelial cells (pCL-MVECs of the porcine corpus luteum has not yet been defined. The aim of the study was to assess FPr expression in pCL-MVECs in the early and mid-luteal phases (EL-p, ML-p, and during pregnancy (P-p. Moreover, the effectiveness of prostaglandin F2-alpha treatment in inducing pCL-MVEC apoptosis was tested. Methods Porcine CLs were collected in the EL and ML phases and during P-p. All CLs from each animal were minced together and the homogenates underwent enzymatic digestion. The pCL-MVECs were then positively selected by an immunomagnetic separation protocol using Dynabeads coated with anti-CD31 monoclonal antibody and seeded in flasks in the presence of EGM 2-MV (Microvascular Endothelial Cell Medium-2. After 4 days of culture, the cells underwent additional immunomagnetic selection and were seeded in flasks until the confluent stage. PCR Real time, western blot and immunodetection assays were utilized to assess the presence of FPr on pCL-MVEC primary cultures. Furthermore, the influence of culture time (freshly isolated, cultured overnight and at confluence and hormonal treatment (P4 and E2 on FPr expression in pCL-MVECs was also investigated. Apoptosis was detected by TUNEL assay of pCL-MVECs exposed to prostaglandin F2-alpha. Results We obtained primary cultures of pCL-MVECs from all animals. FPr mRNA and protein levels showed the highest value (ANOVA in CL-MVECs derived from the early-luteal phase. Moreover, freshly isolated MVECs showed a higher FPr mRNA value than those cultured overnight and confluent cells (ANOVA. prostaglandin F2-alpha

  3. Exposure to lipopolysaccharide and/or unconjugated bilirubin impair the integrity and function of brain microvascular endothelial cells.

    Directory of Open Access Journals (Sweden)

    Filipa L Cardoso

    Full Text Available BACKGROUND: Sepsis and jaundice are common conditions in newborns that can lead to brain damage. Though lipopolysaccharide (LPS is known to alter the integrity of the blood-brain barrier (BBB, little is known on the effects of unconjugated bilirubin (UCB and even less on the joint effects of UCB and LPS on brain microvascular endothelial cells (BMEC. METHODOLOGY/PRINCIPAL FINDINGS: Monolayers of primary rat BMEC were treated with 1 µg/ml LPS and/or 50 µM UCB, in the presence of 100 µM human serum albumin, for 4 or 24 h. Co-cultures of BMEC with astroglial cells, a more complex BBB model, were used in selected experiments. LPS led to apoptosis and UCB induced both apoptotic and necrotic-like cell death. LPS and UCB led to inhibition of P-glycoprotein and activation of matrix metalloproteinases-2 and -9 in mono-cultures. Transmission electron microscopy evidenced apoptotic bodies, as well as damaged mitochondria and rough endoplasmic reticulum in BMEC by either insult. Shorter cell contacts and increased caveolae-like invaginations were noticeable in LPS-treated cells and loss of intercellular junctions was observed upon treatment with UCB. Both compounds triggered impairment of endothelial permeability and transendothelial electrical resistance both in mono- and co-cultures. The functional changes were confirmed by alterations in immunostaining for junctional proteins β-catenin, ZO-1 and claudin-5. Enlargement of intercellular spaces, and redistribution of junctional proteins were found in BMEC after exposure to LPS and UCB. CONCLUSIONS: LPS and/or UCB exert direct toxic effects on BMEC, with distinct temporal profiles and mechanisms of action. Therefore, the impairment of brain endothelial integrity upon exposure to these neurotoxins may favor their access to the brain, thus increasing the risk of injury and requiring adequate clinical management of sepsis and jaundice in the neonatal period.

  4. Influence of curvature on the morphology of brain microvascular endothelial cells

    Science.gov (United States)

    Ye, Mao; Yang, Zhen; Wong, Andrew; Searson, Peter; Searson Group Team

    2013-03-01

    There are hundreds or thousands of endothelial cells around the perimeter of a single artery or vein, and hence an individual cell experiences little curvature. In contrast, a single endothelial cell may wrap around itself to form the lumen of a brain capillary. Curvature plays a key role in many biological, chemical and physical processes, however, its role in dictating the morphology and polarization of brain capillary endothelial cells has not been investigated. We hypothesize that curvature and shear flow play a key role in determining the structure and function of the blood-brain barrier (BBB). We have developed the ``rod'' assay to study the influence of curvature on the morphology of confluent monolayers of endothelial cells. In this assay cells are plated onto glass rods pulled down to the desired diameter in the range from 5 - 500 μm and coated with collagen. We show that curvature has a significant influence on the morphology of endothelial cells and may have an important role in blood-brain barrier function.

  5. Internal Mammary Recipient Site Breast Cancer Recurrence Following Delayed Microvascular Breast Reconstruction

    OpenAIRE

    Rosich-Medina, Anais; Wang, Susan; Erel, Ertan; Malata, Charles M.

    2013-01-01

    Objective: The internal mammary vessels are a popular recipient site for microsurgical anastomoses of free flap breast reconstructions. We, however, observed 3 patients undergoing internal mammary vessel delayed free flap breast reconstruction that subsequently developed tumor recurrence at this site. We reviewed their characteristics to determine whether there was a correlation between delayed microsurgical reconstruction and local recurrence. Methods: A retrospective review of a single surg...

  6. Hypoxia-induced reduction of sVEGFR-2 levels in human colonic microvascular endothelial cells in vitro: Comparative study with HUVEC.

    Science.gov (United States)

    Jayasinghe, Caren; Simiantonaki, Nektaria; Michel-Schmidt, Romi; Kirkpatrick, Charles James

    2009-01-01

    The functionality of large-vessel endothelial cells, such as human umbilical vein endothelial cells (HUVEC), may differ significantly from that in the microvasculature. We established a method for the isolation of human colonic microvascular endothelial cells (HCMEC). Since colonic diseases are often accompanied by hypoxia we examined its effects on HCMEC of five individuals in comparison with HUVEC, with respect to the secretion of the soluble form of the two important vascular endothelial growth factor (VEGF) receptors, VEGFR-1 and 2. After dissociation by dispase/collagenase of mucosal and submucosal tissue obtained from normal adult colon, HCMEC were isolated using CD31-coated magnetic beads and cultivated as monolayers. Subsequent characterization studies demonstrated the endothelial phenotype, including VEGFR-1 and 2 mRNA and protein expression. sVEGFR expression analyses were performed using ELISA. Under hypoxic conditions significantly enhanced levels of sVEGFR-1 on HUVEC were observed (pHUVEC were variable, that is, either unchanged or up-regulated. The different secretion profiles of sVEGFR-1 and 2 between HUVEC and HCMEC under normoxia and hypoxia underline the importance of using a functionally adequate and relevant microvasculature for in vitro studies of colonic diseases. The homogeneously reduced sVEGFR-2 levels in hypoxic HCMEC provide evidence for a novel microvascular endothelium-specific biomarker in hypoxia-response processes.

  7. Tumor Microvasculature: Endothelial Leakiness and Endothelial Pore Size Distribution in a Breast Cancer Model

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    E.E. Uzgiris

    2008-01-01

    Full Text Available Tumor endothelial leakiness is quantified in a rat mammary adenocarcinoma model using dynamic contrast enhancement MRI and contrast agents of widely varying sizes. The contrast agents were constructed to be of globular configuration and have their uptake rate into tumor interstitium be driven by the same diffusion process and limited only by the availability of endothelial pores of passable size. It was observed that the endothelial pore distribution has a steep power law dependence on size, r−β, with an exponent of −4.1. The model of large pore dominance in tumor leakiness as reported in some earlier investigation with fluorescent probes and optical chamber methods is rejected for this tumor model and a number of other tumor types including chemically induced tumors. This steep power law dependence on size is also consistent with observations on human breast cancer.

  8. Differential effects of Bartonella henselae on human and feline macro- and micro-vascular endothelial cells

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    Berrich, Moez; Kieda, Claudine; Grillon, Catherine; Monteil, Martine; Lamerant, Nathalie; Gavard, Julie; Haddad, Nadia

    2011-01-01

    Bartonella henselae, a zoonotic agent, induces tumors of endothelial cells (ECs), namely bacillary angiomatosis and peliosis in immunosuppressed humans but not in cats. In vitro studies on ECs represent to date the only way to explore the interactions between Bartonella henselae and vascular endothelium. However, no comparative study of the interactions between Bartonella henselae and human (incidental host) ECs vs feline (reservoir host) ECs has been carried out because of the absence of ...

  9. Differential Effects of Bartonella henselae on Human and Feline Macro- and Micro-Vascular Endothelial Cells

    OpenAIRE

    Moez Berrich; Claudine Kieda; Catherine Grillon; Martine Monteil; Nathalie Lamerant; Julie Gavard; Henri Jean Boulouis; Nadia Haddad

    2011-01-01

    Bartonella henselae, a zoonotic agent, induces tumors of endothelial cells (ECs), namely bacillary angiomatosis and peliosis in immunosuppressed humans but not in cats. In vitro studies on ECs represent to date the only way to explore the interactions between Bartonella henselae and vascular endothelium. However, no comparative study of the interactions between Bartonella henselae and human (incidental host) ECs vs feline (reservoir host) ECs has been carried out because of the absence of any...

  10. Endogenous microRNAs in human microvascular endothelial cells regulate mRNAs encoded by hypertension-related genes.

    Science.gov (United States)

    Kriegel, Alison J; Baker, Maria Angeles; Liu, Yong; Liu, Pengyuan; Cowley, Allen W; Liang, Mingyu

    2015-10-01

    The goal of this study was to systematically identify endogenous microRNAs (miRNAs) in endothelial cells that regulate mRNAs encoded by genes relevant to hypertension. Small RNA deep sequencing was performed in cultured human microvascular endothelial cells. Of the 50 most abundant miRNAs identified, 30 had predicted target mRNAs encoded by genes with known involvement in hypertension or blood pressure regulation. The cells were transfected with anti-miR oligonucleotides to inhibit each of the 30 miRNAs and the mRNA abundance of predicted targets was examined. Of 95 miRNA-target pairs examined, the target mRNAs were significantly upregulated in 35 pairs and paradoxically downregulated in 8 pairs. The result indicated significant suppression of the abundance of mRNA encoded by ADM by endogenous miR-181a-5p, ATP2B1 by the miR-27 family, FURIN by miR-125a-5p, FGF5 by the let-7 family, GOSR2 by miR-27a-3p, JAG1 by miR-21-5p, SH2B3 by miR-30a-5p, miR-98, miR-181a-5p, and the miR-125 family, TBX3 by the miR-92 family, ADRA1B by miR-22-3p, ADRA2A by miR-30a-5p and miR-30e-5p, ADRA2B by miR-30e-5p, ADRB1 by the let-7 family and miR-98, EDNRB by the miR-92 family, and NOX4 by the miR-92 family, miR-100-5p, and miR-99b-5p (n=3-9; Phypertension.

  11. Stimulated mast cells promote maturation of myocardial microvascular endothelial cell neovessels by modulating the angiopoietin-Tie-2 signaling pathway

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    Wang, Z.H. [Division of Cardiology, Shanghai Sixth People' s Hospital, School of Medicine, Shanghai Jiaotong University, Shanghai, China, Division of Cardiology, Shanghai Sixth People’s Hospital, School of Medicine, Shanghai Jiaotong University, Shanghai (China); Yancheng People' s First Hospital, Division of Cardiology, Yancheng, Jiangsu, China, Division of Cardiology, Yancheng People’s First Hospital, Yancheng, Jiangsu (China); Zhu, W.; Tao, J.P.; Zhang, Q.Y.; Wei, M. [Division of Cardiology, Shanghai Sixth People' s Hospital, School of Medicine, Shanghai Jiaotong University, Shanghai, China, Division of Cardiology, Shanghai Sixth People’s Hospital, School of Medicine, Shanghai Jiaotong University, Shanghai (China)

    2013-10-22

    Angiopoietin (Ang)-1 and Ang-2 interact in angiogenesis to activate the Tie-2 receptor, which may be involved in new vessel maturation and regression. Mast cells (MCs) are also involved in formation of new blood vessels and angiogenesis. The present study was designed to test whether MCs can mediate angiogenesis in myocardial microvascular endothelial cells (MMVECs). Using a rat MMVEC and MC co-culture system, we observed that Ang-1 protein levels were very low even though its mRNA levels were increased by MCs. Interestingly, MCs were able to enhance migration, proliferation, and capillary-like tube formation, which were associated with suppressed Ang-2 protein expression, but not Tie-2 expression levels. These MCs induced effects that could be reversed by either tryptase inhibitor [N-tosyl-L-lysine chloromethyl ketone (TLCK)] or chymase inhibitor (N-tosyl-L-phenylalanyl chloromethyl ketone), with TLCK showing greater effects. In conclusion, our data indicated that MCs can interrupt neovessel maturation via suppression of the Ang-2/Tie-2 signaling pathway.

  12. Proteomic Analysis of Human Brain Microvascular Endothelial Cells Reveals Differential Protein Expression in Response to Enterovirus 71 Infection

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    Wenying Luo

    2015-01-01

    Full Text Available 2D DIGE technology was employed on proteins prepared from human brain microvascular endothelial cells (HBMEC, to study the differentially expressed proteins in cells at 0 h, 1 h, 16 h, and 24 h after infection. Proteins found to be differentially expressed were identified with matrix-assisted laser desorption/ionization time-of-flight/time-of-flight mass spectrometry (MALDITOF/TOF MS analysis. We identified 43 spots showing changes of at least 2.5 fold up- or downregulated expressions in EV71-infected cells at different time when comparing to control, and 28 proteins could be successfully identified by MALDI TOF/TOF mass spectrometry analysis. 4 proteins were significantly upregulated, and 6 proteins were downregulated, another 18 proteins were different expression at different incubation time. We identified changes in the expression of 12 cellular metabolism-related proteins, 5 molecules involved in cytoskeleton, 3 molecules involved in energy metabolism, 2 molecules involved in signal transduction, 1 molecule involved in the ubiquitin-proteasome pathway, 1 molecule involved in cell cycle, 1 molecule involved in apoptosis-related protein, 1 molecular chaperone, and 2 unknown proteins. These findings build up a comprehensive profile of the HBMEC proteome and provide a useful basis for further analysis of the pathogenic mechanism that underlies EV71 infections to induce severe neural complications.

  13. EGb 761 protects cardiac microvascular endothelial cells against hypoxia/reoxygenation injury and exerts inhibitory effect on ATM pathway.

    Science.gov (United States)

    Zhang, Chao; Wang, Deng-Feng; Zhang, Zhuang; Han, Dong; Yang, Kan

    2016-12-14

    Ginkgo biloba extract (EGb 761) has been widely clinically used to reduce myocardial ischemia reperfusion injury (MIRI). Microvascular endothelial cells (MVECs) may be a proper cellular model in vitro for the effect and mechanism study against MIRI. However, the effect of EGb 761 on MVECs resisting hypoxia/reoxygenation (H/R) injury is little reported. In this study, H/R-injuried MVECs were treated with EGb 761, then cell viability, apoptosis, ROS production, SOD activity, caspase-3 activity, and the protein level of ATM, γ-H2AX, p53, Bax were measured. ATM siRNA was transfected to study the changes of protein in ATM pathway. EGb 761 presented protective effect on H/R-injuried MVECs with decreasing cell death, apoptosis and ROS, and elevated SOD activity. Next, EGb 761 could inhibit H/R-induced ATM, γ-H2AX, p53, Bax in a dose-dependent manner. Moreover, ATM siRNA also could inhibit H/R-induced ATM, γ-H2AX, p53, Bax. Overall, these findings verify EGb 761 protects cardiac MVECs from H/R injury, and for the first time, illustrate the influence on ATM pathway and apoptosis of EGb 761 via dampening ROS.

  14. Exosomes contribute to the transmission of anti-HIV activity from TLR3-activated brain microvascular endothelial cells to macrophages

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    Sun, Li; Wang, Xu; Zhou, Yu; Zhou, Run-Hong; Ho, Wen-Zhe; Li, Jie-Liang

    2017-01-01

    Human brain microvascular endothelial cells (HBMECs), the major cell type in the blood-brain barrier (BBB), play a key role in maintaining brain homeostasis. However, their role in the BBB innate immunity against HIV invasion of the central nervous system (CNS) remains to be determined. Our early work showed that TLR3 signaling of HBMECs could produce the antiviral factors that inhibit HIV replication in macrophages. The present study examined whether exosomes from TLR3-activated HBMECs mediate the intercellular transfer of antiviral factors to macrophages. Primary human macrophages could take up exosomes from TLR3-activated HBMECs. HBMECs-derived exosomes contained multiple antiviral factors, including several key IFN-stimulated genes (ISGs; ISG15, ISG56, and Mx2) at mRNA and protein levels. The depletion of exosomes from TLR3-activated HBMECs culture supernatant diminished HBMECs-mediated anti-HIV activity in macrophages. In conclusion, we demonstrate that exosomes shed by HBMECs are able to transport the antiviral molecules to macrophages. This finding suggests the possibility that HIV nonpermissive BBB cells (HBMECs) can help to restore the antiviral state in HIV-infected macrophages, which may be a defense mechanism against HIV neuroinvasion. PMID:27496004

  15. Tumour Necrosis Factor α Enhances CCL2 and ICAM-1 Expression in Peripheral Nerve Microvascular Endoneurial Endothelial Cells

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    Kelly A. Langert

    2013-01-01

    Full Text Available Recruitment and trafficking of autoreactive leucocytes across the BNB (blood–nerve barrier is an early pathological insult in GBS (Guillain-Barré syndrome, an aggressive autoimmune disorder of the PNS (peripheral nervous system. Whereas the aetiology and pathogenesis of GBS remain unclear, pro-inflammatory cytokines, including TNFα (tumour necrosis factor α, are reported to be elevated early in the course of GBS and may initiate nerve injury by activating the BNB. Previously, we reported that disrupting leucocyte trafficking in vivo therapeutically attenuates the course of an established animal model of GBS. Here, PNMECs (peripheral nerve microvascular endothelial cells that form the BNB were harvested from rat sciatic nerves, immortalized by SV40 (simian virus 40 large T antigen transduction and subsequently challenged with TNFα. Relative changes in CCL2 (chemokine ligand 2 and ICAM-1 (intercellular adhesion molecule 1 expression were determined. We report that TNFα elicits marked dose- and time-dependent increases in CCL2 and ICAM-1 mRNA and protein content and promotes secretion of functional CCL2 from immortalized and primary PNMEC cultures. TNFα-mediated secretion of CCL2 promotes, in vitro, the transendothelial migration of CCR2-expressing THP-1 monocytes. Increased CCL2 and ICAM-1 expression in response to TNFα may facilitate recruitment and trafficking of autoreactive leucocytes across the BNB in autoimmune disorders, including GBS.

  16. Focal adhesion kinase is involved in type III group B streptococcal invasion of human brain microvascular endothelial cells.

    Science.gov (United States)

    Shin, Sooan; Paul-Satyaseela, Maneesh; Maneesh, Paul-Satyaseela; Lee, Jong-Seok; Romer, Lewis H; Kim, Kwang Sik

    2006-01-01

    Group B streptococcus (GBS), the leading cause of neonatal meningitis, has been shown to invade human brain microvascular endothelial cells (HBMEC), which constitute the blood-brain barrier. GBS invasion of HBMEC has been shown to require the host cell actin cytoskeleton rearrangements. The present study examined the mechanisms underlying actin cytoskeleton rearrangements that are involved in type III GBS invasion of HBMEC. We showed that type III GBS invasion was inhibited by genistein, a general tyrosine kinase inhibitor (mean 54% invasion decrease at 100 microM), and LY294002, a phosphatidylinositol 3 (PI3) kinase inhibitor (mean 70% invasion decrease at 50 microM), but not by PP2, an inhibitor of the Src family tyrosine kinases. We subsequently showed that the focal adhesion kinase (FAK) was the one of the host proteins tyrosine phosphorylated by type III GBS. Over-expression of a dominant negative form of the FAK C-terminal domain significantly decreased type III GBS invasion of HBMEC (mean 51% invasion decrease). In addition, we showed that FAK phosphorylation correlated with its association of paxillin, an adapter protein of actin filament, and PI3-kinase subunit p85. This is the first demonstration that FAK phosphorylation and its association with paxillin and PI3 kinase play a key role in type III GBS invasion of HBMEC.

  17. Effect of baicalin and berberine on transport of nimodipine on primary-cultured, rat brain microvascular endothelial cells

    Institute of Scientific and Technical Information of China (English)

    Dong-mei ZHANG; Hai-yan LIU; Lin XIE; Xiao-dong LIU

    2007-01-01

    Aim: To investigate whether baicalin and berberine affects the transport of nimodipine (NMD) across the blood-brain barrier (BBB). Methods: Primary-cultured, rat brain microvascular endothelial cells (rBMEC) were used as an in vitro model of the BBB. When cells became confluent, the steady-state uptake of NMD by rBMEC with or without baicalin and berberine was measured. The ef-fects of baicalin and berberine on the efflux of NMD from rBMEC were also studied.Results: Baicalin (2-5 μg/mL) increased the uptake of NMD, and baicalin (10-20 μg/mL) decreased the uptake. The steady-state uptake of NMD was higher than that of control group in the presence of 0.01-1 μg/mL berberine, but was lower in the presence of 2-10 μg/mL berberine. Conclusion: The bidirectional effect of baicalin and berberine on the uptake of NMD by rBMEC was found. Higher concentration showed an inhibitory effect, and lower concentration demonstrated an increasing effect.

  18. Culture of mouse cardiac microvascular endothelial cells in vitro%小鼠心肌微血管内皮细胞的体外培养

    Institute of Scientific and Technical Information of China (English)

    陈桂秀; 杨明涛; 刘涛; 王浩宇; 刘康; 冯刚

    2013-01-01

    目的 建立小鼠心肌微血管内皮细胞培养体系.方法 4~6周的清洁级C57小鼠的心室肌,利用胰蛋白酶及Ⅱ型胶原酶消化过滤收集的滤液进行重新悬浮种植于明胶包被的培养瓶中,通过倒置电镜观察细胞的生长形态及生长状态,得出生长曲线,并利用免疫荧光鉴定(心肌微血管内皮细胞特异性抗原vWF)培养出的小鼠心肌微血管内皮细胞.结果 通过形态学观察及免疫荧光鉴定证实为小鼠心肌微血管内皮细胞.培养的小鼠心肌微血管内皮细胞第1、2天生长相对缓慢,而到第3、4天细胞呈对数生长,第6、7天细胞达到融合.结论 采用明胶包被培养瓶,通过机械剪切、蛋白酶消化、过滤方法,并进行了相关鉴定,可获得较纯的小鼠心肌微血管内皮细胞,这为研究心肌微血管内皮细胞的迁移、血管再生等提供了实验来源.%Objective To establish the culture system of mouse cardiac microvascular endothelial cells. Methods The ventricular muscle from 4 to 6 weeks of C57 mice was digested by trypsin and type II collagen, then filtered and collected the filtrate to grow in culture bottle that was used gelatin to envelop. The morphology and growth curve of mouse cardiac microvascular endothelial cells was investigated by using electron microscopy. The immunofluorescence was uesd to identify culture of mouse primary cardiac microvascular endothelial cells by the expression of factor vWF-related antigen. Results The mouse cardiac microvascular endothelial cells were demonstrated by morphological observation and immunofluorescence. Culture of mouse cardiac microvascular endothelial cells grow were relatively slowly in 1 to 2 day, grow ogarithmicly in 3 to 4 day, grow cell fusion in 6 to 7 day. Conclusion In the experiment, used gelatin to envelope culture bottle,We can obtain mouse cardiac microvascular endothelial cells with higher purity by mechanical shearing, protease digestion, filtering, and

  19. Morphological and protein profile comparison of large vessel and microvascular endothelial cells in culture

    Energy Technology Data Exchange (ETDEWEB)

    Beer, D.M.; Kim, J.S.; Carson, M.P.; Haudeuschild, C.C.; Patton, W.F.; Jacobson, B.S.

    1986-05-01

    Bovine adrenal medulla (AmMEC) and brain (BrMEC) microvessel endothelial cells, and bovine aortic (BAE) endothelial cells were isolated and cultured under identical conditions using a modification of a technique previously described for BrMEC. The cells were isolated and passaged under conditions minimizing cell surface alterations. Primary cultures were confluent in 4-6 days at a plating density in the region of 10/sup 4/ cells/cm/sup 2/. BAEs maintained a cobblestone morphology and a denser monolayer than MECs in primary and passaged cells whether the cells were passaged using Pancreatin, Trypsin-EDTA, or Collagenase-EDTA. MECs were initially elongate and became more like BAEs with passaging. BAEs and AmMECs were examined for differences in whole cell, Triton extracted cytoskeleton and plasma membrane (PM) protein profiles by two-dimensional gel electrophoresis. Cells were labeled with /sup 35/S-methionine and PM by lactoperoxidase catalyzed iodination. Though for the most part protein patterns were similar, several proteins in the PM and cytoskeletal preparations differed. A significant difference in the isoelectric forms of proteins with the same molecular weight was observed in the PM.

  20. A novel paradigm for heart failure with preserved ejection fraction: comorbidities drive myocardial dysfunction and remodeling through coronary microvascular endothelial inflammation.

    Science.gov (United States)

    Paulus, Walter J; Tschöpe, Carsten

    2013-07-23

    Over the past decade, myocardial structure, cardiomyocyte function, and intramyocardial signaling were shown to be specifically altered in heart failure with preserved ejection fraction (HFPEF). A new paradigm for HFPEF development is therefore proposed, which identifies a systemic proinflammatory state induced by comorbidities as the cause of myocardial structural and functional alterations. The new paradigm presumes the following sequence of events in HFPEF: 1) a high prevalence of comorbidities such as overweight/obesity, diabetes mellitus, chronic obstructive pulmonary disease, and salt-sensitive hypertension induce a systemic proinflammatory state; 2) a systemic proinflammatory state causes coronary microvascular endothelial inflammation; 3) coronary microvascular endothelial inflammation reduces nitric oxide bioavailability, cyclic guanosine monophosphate content, and protein kinase G (PKG) activity in adjacent cardiomyocytes; 4) low PKG activity favors hypertrophy development and increases resting tension because of hypophosphorylation of titin; and 5) both stiff cardiomyocytes and interstitial fibrosis contribute to high diastolic left ventricular (LV) stiffness and heart failure development. The new HFPEF paradigm shifts emphasis from LV afterload excess to coronary microvascular inflammation. This shift is supported by a favorable Laplace relationship in concentric LV hypertrophy and by all cardiac chambers showing similar remodeling and dysfunction. Myocardial remodeling in HFPEF differs from heart failure with reduced ejection fraction, in which remodeling is driven by loss of cardiomyocytes. The new HFPEF paradigm proposes comorbidities, plasma markers of inflammation, or vascular hyperemic responses to be included in diagnostic algorithms and aims at restoring myocardial PKG activity.

  1. Up-regulation of COX-2/PGE2 by endothelin-1 via MAPK-dependent NF-κB pathway in mouse brain microvascular endothelial cells

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    Lin Chih-Chung

    2013-01-01

    Full Text Available Abstract Background Endothelin-1 (ET-1 is a proinflammatory mediator and elevated in the regions of several brain injury and inflammatory diseases. The deleterious effects of ET-1 on endothelial cells may aggravate brain inflammation mediated through the regulation of cyclooxygenase-2 (COX-2/prostaglandin E2 (PGE2 system in various cell types. However, the signaling mechanisms underlying ET-1-induced COX-2 expression in brain microvascular endothelial cells remain unclear. Herein we investigated the effects of ET-1 in COX-2 regulation in mouse brain microvascular endothelial (bEnd.3 cells. Results The data obtained with Western blotting, RT-PCR, and immunofluorescent staining analyses showed that ET-1-induced COX-2 expression was mediated through an ETB-dependent transcriptional activation. Engagement of Gi- and Gq-protein-coupled ETB receptors by ET-1 led to phosphorylation of ERK1/2, p38 MAPK, and JNK1/2 and then activated transcription factor NF-κB. Moreover, the data of chromatin immunoprecipitation (ChIP and promoter reporter assay demonstrated that the activated NF-κB was translocated into nucleus and bound to its corresponding binding sites in COX-2 promoter, thereby turning on COX-2 gene transcription. Finally, up-regulation of COX-2 by ET-1 promoted PGE2 release in these cells. Conclusions These results suggested that in mouse bEnd.3 cells, activation of NF-κB by ETB-dependent MAPK cascades is essential for ET-1-induced up-regulation of COX-2/PGE2 system. Understanding the mechanisms of COX-2 expression and PGE2 release regulated by ET-1/ETB system on brain microvascular endothelial cells may provide rationally therapeutic interventions for brain injury or inflammatory diseases.

  2. Endothelial cells stimulate growth of normal and cancerous breast epithelial cells in 3D culture

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    Magnusson Magnus K

    2010-07-01

    Full Text Available Abstract Background Epithelial-stromal interaction provides regulatory signals that maintain correct histoarchitecture and homeostasis in the normal breast and facilitates tumor progression in breast cancer. However, research on the regulatory role of the endothelial component in the normal and malignant breast gland has largely been neglected. The aim of the study was to investigate the effects of endothelial cells on growth and differentiation of human breast epithelial cells in a three-dimensional (3D co-culture assay. Methods Breast luminal and myoepithelial cells and endothelial cells were isolated from reduction mammoplasties. Primary cells and established normal and malignant breast cell lines were embedded in reconstituted basement membrane in direct co-culture with endothelial cells and by separation of Transwell filters. Morphogenic and phenotypic profiles of co-cultures was evaluated by phase contrast microscopy, immunostaining and confocal microscopy. Results In co-culture, endothelial cells stimulate proliferation of both luminal- and myoepithelial cells. Furthermore, endothelial cells induce a subpopulation of luminal epithelial cells to form large acini/ducts with a large and clear lumen. Endothelial cells also stimulate growth and cloning efficiency of normal and malignant breast epithelial cell lines. Transwell and gradient co-culture studies show that endothelial derived effects are mediated - at least partially - by soluble factors. Conclusion Breast endothelial cells - beside their role in transporting nutrients and oxygen to tissues - are vital component of the epithelial microenvironment in the breast and provide proliferative signals to the normal and malignant breast epithelium. These growth promoting effects of endothelial cells should be taken into consideration in breast cancer biology.

  3. High glucose induced oxidative stress and apoptosis in cardiac microvascular endothelial cells are regulated by FoxO3a.

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    Chaoming Peng

    Full Text Available AIM: Cardiac microvascular endothelial cells (CMECs dysfunction contributes to cardiovascular complications in diabetes, whereas, the underlying mechanism is not fully clarified. FoxO transcription factors are involved in apoptosis and reactive oxygen species (ROS production. Therefore, the present study was designed to elucidate the potential role of FoxO3a on the CMECs injury induced by high glucose. MATERIALS AND METHODS: CMECs were isolated from hearts of adult rats and cultured in normal or high glucose medium for 6 h, 12 h and 24 h respectively. To down-regulate FoxO3a expression, CMECs were transfected with FoxO3a siRNA. ROS accumulation and apoptosis in CMECs were assessed by dihydroethidine (DHE staining and TUNEL assay respectively. Moreover, the expressions of Akt, FoxO3a, Bim and BclxL in CMECs were assessed by Western blotting assay. RESULTS: ROS accumulation in CMECs was significantly increased after high glucose incubation for 6 to 24 h. Meanwhile, high glucose also increased apoptosis in CMECs, correlated with decreased the phosphorylation expressions of Akt and FoxO3a. Moreover, high glucose incubation increased the expression of Bim, whereas increased anti-apoptotic protein BclxL. Furthermore, siRNA target FoxO3a silencing enhanced the ROS accumulation, whereas suppressed apoptosis in CMECs. FoxO3a silencing also abolished the disturbance of Bcl-2 proteins induced by high glucose in CMECs. CONCLUSION: Our data provide evidence that high glucose induced FoxO3a activation which suppressed ROS accumulation, and in parallel, resulted in apoptosis of CMECs.

  4. Insulin-like growth factor-1 secreted by brain microvascular endothelial cells attenuates neuron injury upon ischemia.

    Science.gov (United States)

    Wang, Jun; Tang, Yibo; Zhang, Wei; Zhao, Haiping; Wang, Runjun; Yan, Yangyang; Xu, Liwei; Li, Pengtao

    2013-08-01

    Insulin-like growth factor (IGF)-1 is essential for the development of the nervous system, and is present in many cell types. Relatively little is known about IGF-1 expression in brain microvascular endothelial cells (BMECs). For in vivo studies, we examined the expression of IGF-1 and insulin-like growth factor-binding protein (IGFBP)-2 after focal cerebral ischemia for 12 h, 24 h, 3 days and 7 days, utilizing a permanent middle cerebral artery occlusion (MCAO) model in rats. For in vitro studies, we examined the levels of IGF-1 and IGFBP-2 in the culture medium or primary culture of BMECs injured by oxygen-glucose deprivation (OGD). Then, we elucidated the protective effects of IGF-1 on cortical neurons injured by OGD and the possible mechanism. In addition, we investigated the effect of BMEC-conditioned medium on IGF-1 receptor expression in neurons. The results showed that IGF-1 expression increased in serum and brain tissue, whereas IGFBP-2 expression decreased in brain tissue of MCAO-injured rats. In primary culture of BMECs, the expression levels of IGF-1 and IGFBP-2 were significantly higher under OGD conditions in culture. IGF-1 administration improved neuron viability upon normoxia or OGD, and upregulated p-Akt expression. This effect was reversed by LY294002, a specific inhibitor of the phosphoinositide 3-kinase-Akt signaling pathway. Furthermore, conditioned medium from OGD-treated BMECs substantially suppressed neuron viability and the expression of IGF-1 receptor simultaneously. These data demonstrate that therapeutic strategies that prioritize the early recovery of the IGF-1 system in BMECs might be promising in ischemic injury.

  5. Interactions of Pseudomonas aeruginosa and Corynebacterium spp. with non-phagocytic brain microvascular endothelial cells and phagocytic Acanthamoeba castellanii.

    Science.gov (United States)

    Siddiqui, Ruqaiyyah; Lakhundi, Sahreena; Khan, Naveed Ahmed

    2015-06-01

    Several lines of evidence suggest that Acanthamoeba interact with bacteria, which may aid in pathogenic bacterial transmission to susceptible hosts, and these interactions may have influenced evolution of bacterial pathogenicity. In this study, we tested if Gram-negative Pseudomonas aeruginosa and Gram-positive Corynebacterium spp. can associate/invade and survive inside Acanthamoeba castellanii trophozoites and cysts, as well as non-phagocytic human brain microvascular endothelial cells. The results revealed that both Corynebacterium spp. and P. aeruginosa were able to associate as well as invade and/or taken up by the phagocytic A. castellanii trophozoite. In contrast, P. aeruginosa exhibited higher association as well as invasion of non-phagocytic HBMEC compared with Corynebacterium spp. Notably, P. aeruginosa remained viable during the encystment process and exhibited higher levels of recovery from mature cysts (74.54 bacteria per amoebae) compared with Corynebacterium spp. (2.69 bacteria per amoeba) (P < 0.05). As Acanthamoeba cysts can be airborne, these findings suggest that Acanthamoeba is a potential vector in the transmission of P. aeruginosa to susceptible hosts. When bacterial-ridden amoebae were exposed to favourable (nutrient-rich) conditions, A. castellanii emerged as vegetative trophozoites and remained viable, and likewise viable P. aeruginosa were also observed but rarely any Corynebacterium spp. were observed. Correspondingly, P. aeruginosa but not Corynebacterium spp. exhibited higher cytotoxicity to non-phagocytic HBMEC, producing more than 75% cell death in 24 h, compared to 20% cell death observed with Corynebacterium spp. Additionally, it was observed that the bacterial conditioned medium had no negative effect on A. castellanii growth. Further characterization of amoebal and bacterial interactions will assist in identifying the role of Acanthamoeba in the transmission and evolution of pathogenic bacteria.

  6. LPS Induces Occludin Dysregulation in Cerebral Microvascular Endothelial Cells via MAPK Signaling and Augmenting MMP-2 Levels

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    Lan-hui Qin

    2015-01-01

    Full Text Available Disrupted blood-brain barrier (BBB integrity contributes to cerebral edema during central nervous system infection. The current study explored the mechanism of lipopolysaccharide- (LPS- induced dysregulation of tight junction (TJ proteins. Human cerebral microvascular endothelial cells (hCMEC/D3 were exposed to LPS, SB203580 (p38MAPK inhibitor, or SP600125 (JNK inhibitor, and cell vitality was determined by MTT assay. The proteins expressions of p38MAPK, JNK, and TJs (occludin and zonula occludens- (ZO- 1 were determined by western blot. The mRNA levels of TJ components and MMP-2 were measured with quantitative real-time polymerase chain reaction (qRT-PCR, and MMP-2 protein levels were determined by enzyme-linked immunosorbent assay (ELISA. LPS, SB203580, and SP600125 under respective concentrations of 10, 7.69, or 0.22 µg/mL had no effects on cell vitality. Treatment with LPS decreased mRNA and protein levels of occludin and ZO-1 and enhanced p38MAPK and JNK phosphorylation and MMP-2 expression. These effects were attenuated by pretreatment with SB203580 or SP600125, but not in ZO-1 expression. Both doxycycline hyclate (a total MMP inhibitor and SB-3CT (a specific MMP-2 inhibitor partially attenuated the LPS-induced downregulation of occludin. These data suggest that MMP-2 overexpression and p38MAPK/JNK pathways are involved in the LPS-mediated alterations of occludin in hCMEC/D3; however, ZO-1 levels are not influenced by p38MAPK/JNK.

  7. Lipid raft/caveolae signaling is required for Cryptococcus neoformans invasion into human brain microvascular endothelial cells

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    Long Min

    2012-02-01

    Full Text Available Abstract Background Cryptococcus neoformans has a predilection for central nervous system infection. C. neoformans traversal of the blood brain barrier, composed of human brain microvascular endothelial cells (HBMEC, is the crucial step in brain infection. However, the molecular mechanism of the interaction between Cryptococcus neoformans and HBMEC, relevant to its brain invasion, is still largely unknown. Methods In this report, we explored several cellular and molecular events involving the membrane lipid rafts and caveolin-1 (Cav1 of HBMEC during C. neoformans infection. Immunofluorescence microscopy was used to examine the roles of Cav1. The knockdown of Cav1 by the siRNA treatment was performed. Phosphorylation of Cav1 relevant to its invasion functions was investigated. Results We found that the host receptor CD44 colocalized with Cav1 on the plasma membrane, and knockdown of Cav1 significantly reduced the fungal ability to invade HBMEC. Although the CD44 molecules were still present, HBMEC membrane organization was distorted by Cav1 knockdown. Concomitantly, knockdown of Cav1 significantly reduced the fungal crossing of the HBMEC monolayer in vitro. Upon C. neoformans engagement, host Cav1 was phosphorylated in a CD44-dependent manner. This phosphorylation was diminished by filipin, a disrupter of lipid raft structure. Furthermore, the phosphorylated Cav1 at the lipid raft migrated inward to the perinuclear localization. Interestingly, the phospho-Cav1 formed a thread-like structure and colocalized with actin filaments but not with the microtubule network. Conclusion These data support that C. neoformans internalization into HBMEC is a lipid raft/caveolae-dependent endocytic process where the actin cytoskeleton is involved, and the Cav1 plays an essential role in C. neoformans traversal of the blood-brain barrier.

  8. Effects of Fas, NF-κB and caspases on rat microvascular endothelial cell apoptosis induced by TNFα

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    AIM: To study the apoptotic effect of TNFα on rat pulmonary microvascular endothelial cells (PMVEC) and the influences of Fas, NF-κB in its mechanism. METHODS: Apoptosis of PMVEC was analyzed and quantitated with TUNEL, flow cytometer. The distribution of NF-κB was detected via histoimmunochemical staining in TNF-treated cells and the control. Northern blot was applied to assess the influence of TNF on PMVEC Fas expression. Fas antibody was used to investigate the apoptotic effect of Fas on PMVEC. Activation of caspase-8 was detected with Western blot. Expression of caspase-3 was analyzed with histoimmunochemical staining. RESULTS: After treatment with 5×108 U/L TNF for 24 hours, viable PMVEC significantly diminished. Apoptosis rate was 14.0%±3.1% detected with TUNEL, and 13.1% with flow cytometer. Histoimmunochemical staining showed that NF-κB relocated from cytoplasm to the nuclear. When the cells were co-cultured with TNF and APDC, an NF-κB inhibitor, less cells were viable and more cells were positively stained with TUNEL. Fas expression in PMVEC was elevated treated with TNF. Apoptosis in PMVEC was found aggravated, when the cells were co-cultured with TNF and anti-Fas antibody. The positive rate was 24.1%±1.5% with TUNEL. Increase of caspase-8 activation was manifested by Western blot following TNF stimulation. Caspase-3 expression was found elevated using histoimmunochemical staining. Cell permeable caspase-3 inhibitor significantly ameliorated PMVEC apoptosis induced by TNF. CONCLUSION: 1. Large dose of TNF(5×108 U/L) can induce apoptosis in rat PMVEC. 2. NF-κB has a protective effect on PMVEC apoptosis. 3. TNF up-regulates Fas expression in PMVEC. And the latter takes a part in apoptosis. 4. TNF induced caspase-8 activation in PMVEC, and more caspase-3 was expressed. These may be involved in PMVEC apoptosis induced by TNF.

  9. MiR-466b-1-3p regulates P-glycoprotein expression in rat cerebral microvascular endothelial cells.

    Science.gov (United States)

    Yang, Xiaobo; Ren, Weimin; Shao, Yiye; Chen, Yinghui

    2017-04-03

    Epilepsy is one of the most common neurological disorders, and approximately one-third of epilepsy cases are resistant to treatment with anti-epileptic drug (AED). P-glycoprotein (P-gp) is a multi-drug transporter that is thought to play a pivotal role in multiple drug resistance (MDR) in epilepsy. The regulatory mechanism of P-gp remains largely unknown; however, recent studies have demonstrated that microRNAs (miRNAs) may regulate the chemo-resistance mediated by P-gp. This study investigated the effect of specific miRNAs that regulate P-gp expression in rat cerebral microvascular endothelial cells (RCMECs). Primary cultures of RCMECs were treated with phenobarbital (PB) at various concentrations to induce P-gp overexpression. MiRNA microarrays were used to investigate the expression profiles of miRNAs in the resistant RCMECs induced by PB and corresponding non-resistant cells. Our data demonstrated decreased miR-466b-1-3p expression in the resistant cells compared with the non-resistant cells. Moreover, the recombinant RNA of 466b-1-3p (mimic) and the artificial antisense RNA of miR-466b-1-3p (inhibitor) were constructed and transfected into resistant RCMECs. The expression and function of P-gp were measured by Western blotting, quantitative real-time polymerase chain reaction (qRT-PCR) and flow cytometry using rhodamine efflux. The mRNA and protein levels of P-gp increased as the concentration of PB increased, whereas miR-466b-1-3p levels decreased with increasing PB concentrations (Pp mimic down-regulated P-gp expression, whereas the miR-466b-1-3p inhibitor up-regulated P-gp expression (Pp may regulate PB-induced P-gp expression in RCMECs.

  10. The interaction between circulating complement proteins and cutaneous microvascular endothelial cells in the development of childhood Henoch-Schonlein Purpura.

    Directory of Open Access Journals (Sweden)

    Yao-Hsu Yang

    Full Text Available In addition to IgA, the deposition of complement (C3 in dermal vessels is commonly found in Henoch-Schönlein purpura (HSP. The aim of this study is to elucidate the role of circulating complement proteins in the pathogenesis of childhood HSP.Plasma levels of C3a, C4a, C5a, and Bb in 30 HSP patients and 30 healthy controls were detected by enzyme-linked immunosorbent assay (ELISA. The expression of C3a receptor (C3aR, C5a receptor (CD88, E-selectin, intercellular adhesion molecule 1 (ICAM-1, C3, C5, interleukin (IL-8, monocyte chemotactic protein (MCP-1, and RANTES by human dermal microvascular endothelial cells (HMVEC-d was evaluated either by flow cytometry or by ELISA.At the acute stage, HSP patients had higher plasma levels of C3a (359.5 ± 115.3 vs. 183.3 ± 94.1 ng/ml, p < 0.0001, C5a (181.4 ± 86.1 vs. 33.7 ± 26.3 ng/ml, p < 0.0001, and Bb (3.7 ± 2.6 vs. 1.0 ± 0.6 μg/ml, p < 0.0001, but not C4a than healthy controls. Although HSP patient-derived acute phase plasma did not alter the presentation of C3aR and CD88 on HMVEC-d, it enhanced the production of endothelial C3 and C5. Moreover, C5a was shown in vitro to up-regulate the expression of IL-8, MCP-1, E-selectin, and ICAM-1 by HMVEC-d with a dose-dependent manner.In HSP, the activation of the complement system in part through the alternative pathway may have resulted in increased plasma levels of C3a and C5a, which, especially C5a, may play a role in the disease pathogenesis by activating endothelium of cutaneous small vessels.

  11. Plasmodium chabaudi-Infected Erythrocytes Adhere to CD36 and Bind to Microvascular Endothelial Cells in an Organ-Specific Way

    Science.gov (United States)

    Mota, Maria M.; Jarra, William; Hirst, Elizabeth; Patnaik, Pradeep K.; Holder, Anthony A.

    2000-01-01

    Adherence of erythrocytes infected with Plasmodium falciparum to microvascular endothelial cells (sequestration) is considered to play an important role in parasite virulence and pathogenesis. However, the real importance of sequestration for infection and disease has never been fully assessed. The absence of an appropriate in vivo model for sequestration has been a major barrier. We have examined the rodent malaria parasite Plasmodium chabaudi chabaudi AS in mice as a potential model. Erythrocytes infected with this parasite adhere in vitro to purified CD36, a critical endothelium receptor for binding P. falciparum-infected erythrocytes. P. c. chabaudi-infected erythrocytes adhere in vitro to endothelial cells in a gamma interferon-dependent manner, suggesting the involvement of additional adhesion molecules in the binding process, as is also the case with P. falciparum-infected cells. Furthermore, plasma or sera from infected and hyperimmune mice, respectively, have the ability to block binding of infected erythrocytes to endothelial cells. In vivo, erythrocytes containing mature P. c. chabaudi parasites are sequestered from the peripheral circulation. Sequestration is organ specific, occurring primarily in the liver, although intimate contact between infected erythrocytes and endothelial cells is also observed in the spleen and brain. The results are discussed in the context of the use of this model to study (i) the relationship between endothelial cell activation and the level of sequestration and (ii) the primary function of sequestration in malaria infection. PMID:10858230

  12. Uncaria tomentosa alkaloidal fraction reduces paracellular permeability, IL-8 and NS1 production on human microvascular endothelial cells infected with dengue virus.

    Science.gov (United States)

    Lima-Junior, Raimundo Sousa; Mello, Cintia da Silva; Siani, Antonio Carlos; Valente, Ligia M Marino; Kubelka, Claire Fernandes

    2013-11-01

    Dengue is the major Arbovirus in the world, annually causing morbidity and death. Severe dengue is associated with changes in the endothelial barrier function due to the production of inflammatory mediators by immune cells and by the endothelium. Dengue virus (DENV) replicates efficiently in human endothelial cells in vitro and elicits immune responses resulting in endothelial permeability. Uncaria tomentosa (Willd.) DC.(Rubiaceae), known as cat's claw, has been used in folk medicine for the treatment of a wide-array of symptoms, and several scientific studies reported its antiviral, immunomodulatory, anti-inflammatory and antioxidant properties. Here we infected a human lineage of dermal microvascular endothelial cells (HMEC-1) with DENV-2 and treated it with an alkaloidal fraction from U. tomentosa bark (AFUT). We showed antiviral and immunomodulatory activities of U. tomentosa by determining the NS1 antigen and IL-8 in supernatant of DENV-2 infected HMEC-1. Furthermore, by measurement of transendothelial electrical resistance (TEER) we demonstrated, for the first time, that a plant derivative contributed to the reduction of paracellular permeability in DENV-2 infected HMEC-1. We also showed that IL-8 contributed significantly to the induction of permeability. Although further investigations should be conducted before a new drug can be suggested, our in vitro data support evidence that AFUT could be potentially useful in developing a treatment for severe dengue.

  13. Advanced glycation end products accelerate ischemia/reperfusion injury through receptor of advanced end product/nitrative thioredoxin inactivation in cardiac microvascular endothelial cells.

    Science.gov (United States)

    Liu, Yi; Ma, Yanzhuo; Wang, Rutao; Xia, Chenhai; Zhang, Rongqing; Lian, Kun; Luan, Ronghua; Sun, Lu; Yang, Lu; Lau, Wayne B; Wang, Haichang; Tao, Ling

    2011-10-01

    The advanced glycation end products (AGEs) are associated with increased cardiac endothelial injury. However, no causative link has been established between increased AGEs and enhanced endothelial injury after ischemia/reperfusion. More importantly, the molecular mechanisms by which AGEs may increase endothelial injury remain unknown. Adult rat cardiac microvascular endothelial cells (CMECs) were isolated and incubated with AGE-modified bovine serum albumin (BSA) or BSA. After AGE-BSA or BSA preculture, CMECs were subjected to simulated ischemia (SI)/reperfusion (R). AGE-BSA increased SI/R injury as evidenced by enhanced lactate dehydrogenase release and caspase-3 activity. Moreover, AGE-BSA significantly increased SI/R-induced oxidative/nitrative stress in CMECs (as measured by increased inducible nitric oxide synthase expression, total nitric oxide production, superoxide generation, and peroxynitrite formation) and increased SI/R-induced nitrative inactivation of thioredoxin-1 (Trx-1), an essential cytoprotective molecule. Supplementation of EUK134 (peroxynitrite decomposition catalyst), human Trx-1, or soluble receptor of advanced end product (sRAGE) (a RAGE decoy) in AGE-BSA precultured cells attenuated SI/R-induced oxidative/nitrative stress, reduced SI/R-induced Trx-1 nitration, preserved Trx-1 activity, and reduced SI/R injury. Our results demonstrated that AGEs may increase SI/R-induced endothelial injury by increasing oxidative/nitrative injury and subsequent nitrative inactivation of Trx-1. Interventions blocking RAGE signaling or restoring Trx activity may be novel therapies to mitigate endothelial ischemia/reperfusion injury in the diabetic population.

  14. Generation of Brain Microvascular Endothelial-Like Cells from Human Induced Pluripotent Stem Cells by Co-Culture with C6 Glioma Cells.

    Science.gov (United States)

    Minami, Haruka; Tashiro, Katsuhisa; Okada, Atsumasa; Hirata, Nobue; Yamaguchi, Tomoko; Takayama, Kazuo; Mizuguchi, Hiroyuki; Kawabata, Kenji

    2015-01-01

    The blood brain barrier (BBB) is formed by brain microvascular endothelial cells (BMECs) and tightly regulates the transport of molecules from blood to neural tissues. In vitro BBB models from human pluripotent stem cell (PSCs)-derived BMECs would be useful not only for the research on the BBB development and function but also for drug-screening for neurological diseases. However, little is known about the differentiation of human PSCs to BMECs. In the present study, human induced PSCs (iPSCs) were differentiated into endothelial cells (ECs), and further maturated to BMECs. Interestingly, C6 rat glioma cell-conditioned medium (C6CM), in addition to C6 co-culture, induced the differentiation of human iPSC-derived ECs (iPS-ECs) to BMEC-like cells, increase in the trans-endothelial electrical resistance, decreased in the dextran transport and up-regulation of gene expression of tight junction molecules in human iPS-ECs. Moreover, Wnt inhibitors attenuated the effects of C6CM. In summary, we have established a simple protocol of the generation of BMEC-like cells from human iPSCs, and have demonstrated that differentiation of iPS-ECs to BMEC-like cells is induced by C6CM-derived signals, including canonical Wnt signals.

  15. Generation of Brain Microvascular Endothelial-Like Cells from Human Induced Pluripotent Stem Cells by Co-Culture with C6 Glioma Cells.

    Directory of Open Access Journals (Sweden)

    Haruka Minami

    Full Text Available The blood brain barrier (BBB is formed by brain microvascular endothelial cells (BMECs and tightly regulates the transport of molecules from blood to neural tissues. In vitro BBB models from human pluripotent stem cell (PSCs-derived BMECs would be useful not only for the research on the BBB development and function but also for drug-screening for neurological diseases. However, little is known about the differentiation of human PSCs to BMECs. In the present study, human induced PSCs (iPSCs were differentiated into endothelial cells (ECs, and further maturated to BMECs. Interestingly, C6 rat glioma cell-conditioned medium (C6CM, in addition to C6 co-culture, induced the differentiation of human iPSC-derived ECs (iPS-ECs to BMEC-like cells, increase in the trans-endothelial electrical resistance, decreased in the dextran transport and up-regulation of gene expression of tight junction molecules in human iPS-ECs. Moreover, Wnt inhibitors attenuated the effects of C6CM. In summary, we have established a simple protocol of the generation of BMEC-like cells from human iPSCs, and have demonstrated that differentiation of iPS-ECs to BMEC-like cells is induced by C6CM-derived signals, including canonical Wnt signals.

  16. Opiates Upregulate Adhesion Molecule Expression in Brain MicroVascular Endothelial Cells (BMVEC: Implications for Altered Blood Brain Barrier (BBB Permeability

    Directory of Open Access Journals (Sweden)

    Madhavan P.N. Nair

    2006-01-01

    Full Text Available The blood-brain barrier (BBB is an intricate cellular system composed of vascular endothelial cells and perivascular astrocytes that restrict the passage of immunocompetent cells into the central nervous system (CNS. Expression of the adhesion molecules, intercellular adhesion molecule 1 (ICAM-1 and vascular cell adhesion molecule-1 (VCAM-1 on brain microvascular endothelial cells (BMVEC and their interaction with human immunodeficiency virus (HIV-1 viral proteins may help enhance viral adhesion and virus-cell fusion resulting in increased infectivity. Additionally, transmigration through the BBB is facilitated by both endothelial and monocyte/macrophage-derived nitric oxide (NO. Dysregulated production of NO by BMVEC due to opiates and HIV-1 viral protein interactions play a pivotal role in brain endothelial injury, resulting in the irreversible loss of BBB integrity, which may lead to enhanced infiltration of virus-carrying cells across the BBB. Opioids act as co-factors in the neuropathogenesis of HIV-1 by facilitating BBB dysfunction however, no studies have been done to investigate the role of opiates alone or in combination with HIV-1 viral proteins on adhesion molecule expression in BMVEC. We hypothesize that opiates such as heroin and morphine in conjunction with the HIV-1 viral protein gp120 increase the expression of adhesion molecules ICAM-1 and VCAM-1 and these effects are mediated via the modulation of NO. Results show that opiates alone and in synergy with gp120 increase both the genotypic and phenotypic expression of ICAM-1 and VCAM-1 by BMVEC, additionally, these opiate induced effects may be the result of increased NO production. These studies will provide a better understanding of how opiate abuse in conjunction with HIV-1 infection facilitates the breakdown of the BBB and exacerbates the neuropathogenesis of HIV-1. Elucidation of the mechanisms of BBB modulation will provide new therapeutic approaches to maintain BBB integrity

  17. The Src family tyrosine kinases src and yes have differential effects on inflammation-induced apoptosis in human pulmonary microvascular endothelial cells.

    Science.gov (United States)

    Nelin, Leif D; White, Hilary A; Jin, Yi; Trittmann, Jennifer K; Chen, Bernadette; Liu, Yusen

    2016-05-01

    Endothelial cells are essential for normal lung function: they sense and respond to circulating factors and hemodynamic alterations. In inflammatory lung diseases such as acute respiratory distress syndrome, endothelial cell apoptosis is an inciting event in pathogenesis and a prominent pathological feature. Endothelial cell apoptosis is mediated by circulating inflammatory factors, which bind to receptors on the cell surface, activating signal transduction pathways, leading to caspase-3-mediated apoptosis. We hypothesized that yes and src have differential effects on caspase-3 activation in human pulmonary microvascular endothelial cells (hPMVEC) due to differential downstream signaling effects. To test this hypothesis, hPMVEC were treated with siRNA against src (siRNAsrc), siRNA against yes (siRNAyes), or their respective scramble controls. After recovery, the hPMVEC were treated with cytomix (LPS, IL-1β, TNF-α, and IFN-γ). Treatment with cytomix induced activation of the extracellular signal-regulated kinase (ERK) pathway and caspase-3-mediated apoptosis. Treatment with siRNAsrc blunted cytomix-induced ERK activation and enhanced cleaved caspase-3 levels, while treatment with siRNAyes enhanced cytomix-induced ERK activation and attenuated levels of cleaved caspase-3. Inhibition of the ERK pathway using U0126 enhanced cytomix-induced caspase-3 activity. Treatment of hPMVEC with cytomix induced Akt activation, which was inhibited by siRNAsrc. Inhibition of the phosphatidylinositol 3-kinase/Akt pathway using LY294002 prevented cytomix-induced ERK activation and augmented cytomix-induced caspase-3 cleavage. Together, our data demonstrate that, in hPMVEC, yes activation blunts the ERK cascade in response to cytomix, resulting in greater apoptosis, while cytomix-induced src activation induces the phosphatidylinositol 3-kinase pathway, which leads to activation of Akt and ERK and attenuation of apoptosis.

  18. Microvascular filtration is increased in the forearms of patients with breast cancer-related lymphedema

    DEFF Research Database (Denmark)

    Jensen, Mads Radmer; Simonsen, Lene; Karlsmark, Tonny

    2013-01-01

    Breast cancer-related lymphedema (BCRL) is a frequent and debilitating complication of breast cancer treatment. The pathophysiology is complex and remains poorly understood; however, data suggest that changes in the peripheral circulation may contribute to edema formation. In 13 volunteers...

  19. N-n-butyl haloperidol iodide ameliorates hypoxia/reoxygenation injury through modulating the LKB1/AMPK/ROS pathway in cardiac microvascular endothelial cells

    Science.gov (United States)

    Lu, Binger; Wang, Bin; Zhong, Shuping; Zhang, Yanmei; Gao, Fenfei; Chen, Yicun; Zheng, Fuchun; Shi, Ganggang

    2016-01-01

    Endothelial cells are highly sensitive to hypoxia and contribute to myocardial ischemia/reperfusion injury. We have reported that N-n-butyl haloperidol iodide (F2) can attenuate hypoxia/reoxygenation (H/R) injury in cardiac microvascular endothelial cells (CMECs). However, the molecular mechanisms remain unclear. Neonatal rat CMECs were isolated and subjected to H/R. Pretreatment of F2 leads to a reduction in H/R injury, as evidenced by increased cell viability, decreased lactate dehydrogenase (LDH) leakage and apoptosis, together with enhanced AMP-activated protein kinase (AMPK) and liver kinase B1 (LKB1) phosphorylation in H/R ECs. Blockade of AMPK with compound C reversed F2-induced inhibition of H/R injury, as evidenced by decreased cell viability, increased LDH release and apoptosis. Moreover, compound C also blocked the ability of F2 to reduce H/R-induced reactive oxygen species (ROS) generation. Supplementation with the ROS scavenger N-acetyl-L-cysteine (NAC) reduced ROS levels, increased cell survival rate, and decreased both LDH release and apoptosis after H/R. In conclusion, our data indicate that F2 may mitigate H/R injury by stimulating LKB1/AMPK signaling pathway and subsequent suppression of ROS production in CMECs. PMID:27166184

  20. Apigenin Isolated from the Medicinal Plant Elsholtzia rugulosa Prevents β-Amyloid 25–35-Induces Toxicity in Rat Cerebral Microvascular Endothelial Cells

    Directory of Open Access Journals (Sweden)

    Qingshan Liu

    2011-05-01

    Full Text Available Endothelial cells of cerebral capillaries forming the blood-brain barrier play an important role in the pathogenesis and therapy of Alzheimer’s disease. Amyloid-β peptides are key pathological elements in the development of this disease. Apigenin (4’,5,7-tetrahydroxyflavone is a plant flavonoid and pharmacologically active agent that can be isolated from several plant species. In the present study, effects of apigenin obtained from the medicinal plant Elsholtzia rugulosa (Labiatae on primary cultured rat cerebral microvascular endothelial cells (CMECs mediated by amyloid-β peptide 25–35 (Aβ25–35 were examined. Aβ25–35 showed toxic effects on CMECs, involving reduction of cell viability, release of lactate dehydrogenase (LDH, increase of nuclear condensation, over-production of intracellular reactive oxygen species (ROS, decrease of superoxide dismutase (SOD activity, and breakage of the barrier integrity and function. Based on this model, we demonstrated that apigenin from the medicinal plant Elsholtzia rugulosa protected cultured rat CMECs by increasing cell viability, reducing LDH release, relieving nuclear condensation, alleviating intracellular ROS generation, increasing SOD activity, and strengthening the barrier integrity through the preservation of transendothelial electrical resistance, permeability property and characteristic enzymatic activity after being exposed to Aβ25–35. In conclusion, apigenin isolated from Elsholtzia rugulosa has the ability to protect rat CMECs against Aβ25–35-induced toxicity.

  1. Effect of oxygen and glucose deprivation on VEGF and its receptors in microvascular endothelial cells co-cultured with mast cells.

    Science.gov (United States)

    Wang, Zhihua; Tao, Jianping; Zhang, Qingyong; Wei, Meng

    2015-09-01

    The aim of this study was to determine the correlation between angiogenesis and the differential expression of vascular endothelial growth factor (VEGF) and its receptors in myocardial microvascular endothelial cells (MMVECs) co-cultured with mast cells (MCs) or mast cell granules (MCGs) under oxygen and glucose deprivation (OGD). MMVECs and MCs were isolated from Wistar rats. MCs spontaneously degranulated in OGD. The expression of VEGF peaked at 8 h and decreased from 16 h in OGD. However, the expression of its receptor, fms-like tyrosine kinase-1 (Flt-1), and fetal liver kinase-1 (Flk-1), decreased significantly, and angiogenic potential of MMVECs decreased in OGD. Expression of VEGF, Flt-1, and Flk-1 increased significantly when MMVECs were co-cultured with MCGs or active MCs, but MCs had only a limited ability to induce angiogenesis in OGD. The angiogenic potential of MMVECs cultured in OGD (even with MCGs) was inferior to that of MMVECs cultured under normoxic conditions. OGD have a profound effect on angiogenesis, which is more pronounced than the effect of MCs on angiogenesis.

  2. The protective role of isorhamnetin on human brain microvascular endothelial cells from cytotoxicity induced by methylglyoxal and oxygen-glucose deprivation.

    Science.gov (United States)

    Li, Wenlu; Chen, Zhigang; Yan, Min; He, Ping; Chen, Zhong; Dai, Haibin

    2016-02-01

    As the first target of stroke, cerebral endothelial cells play a key role in brain vascular repair and maintenance, and their function is impeded in diabetes. Methylglyoxal (MGO), a reactive dicarbonyl produced during glucose metabolism, accumulates in diabetic patients. MGO and MGO-induced advanced glycation end-products (AGEs) could ameliorate stroke-induced brain vascular damage, closely related with ECs dysfunction. Using MGO plus oxygen-glucose deprivation (OGD) to mimic diabetic stroke, we reported the protective effect of isorhamnetin on OGD-induced cytotoxicity after MGO treatment on primary human brain microvascular endothelial cells (HBMEC) and explored the underlying mechanisms. Treatment of MGO for 24 h significantly enhanced 3-h OGD-induced HBMEC toxic effect, which was inhibited by pretreatment of isorhamnetin (100 μmol/L). Moreover, the protective effect of isorhamnetin is multiple function dependent, which includes anti-inflammation, anti-oxidative stress and anti-apoptosis effects. Besides its well-known inhibition on the mitochondria-dependent or intrinsic apoptotic pathway, isorhamnetin also reduced activation of the extrinsic apoptotic pathway, as characterized by the decreased expression and activity of caspase 3 and caspase 8. Furthermore, pretreatment with isorhamnetin specifically inhibited FAS/FASL expression and suppressed nuclear factor-kappa B nuclear translocation. Taken together, our results indicated that isorhamnetin protected against OGD-induced cytotoxicity after MGO treatment in cultured HBMEC due to its multiple protective effects and could inhibit Fas-mediated extrinsic apoptosis. Therefore, isorhamnetin is a promising reagent for the treatment of hyperglycemia and ischemia-induced cerebral vascular degeneration. A proposed model of the potential protective mechanism of isorhamnetin, a metabolite of quercetin, on methylglyoxal (MGO) treatment plus oxygen-glucose deprivation (OGD) exposure-induced cytotoxicity in cultured human

  3. Human breast microvascular endothelial cells retain phenotypic traits in long-term finite life span culture

    DEFF Research Database (Denmark)

    Sigurdsson, Valgardur; Fridriksdottir, Agla J R; Kjartansson, Jens

    2007-01-01

    uptake of low-density lipoprotein, and had E-selectin induced upon treatment with tumor necrosis factor-alpha. The first signs of senescence in passage 14 were accompanied by gain of trisomy 11. At passage 18 cells showed chromosomal aberrations and growth arrest as revealed by beta...

  4. The novel mitochondria-targeted hydrogen sulfide (H2S) donors AP123 and AP39 protect against hyperglycemic injury in microvascular endothelial cells in vitro.

    Science.gov (United States)

    Gerő, Domokos; Torregrossa, Roberta; Perry, Alexis; Waters, Alicia; Le-Trionnaire, Sophie; Whatmore, Jacqueline L; Wood, Mark; Whiteman, Matthew

    2016-11-01

    The development of diabetic vascular complications is initiated, at least in part, by mitochondrial reactive oxygen species (ROS) production in endothelial cells. Hyperglycemia induces superoxide production in the mitochondria and initiates changes in the mitochondrial membrane potential that leads to mitochondrial dysfunction. Hydrogen sulfide (H2S) supplementation has been shown to reduce the mitochondrial oxidant production and shows efficacy against diabetic vascular damage in vivo. However, the half-life of H2S is very short and it is not specific for the mitochondria. We have therefore evaluated two novel mitochondria-targeted anethole dithiolethione and hydroxythiobenzamide H2S donors (AP39 and AP123 respectively) at preventing hyperglycemia-induced oxidative stress and metabolic changes in microvascular endothelial cells in vitro. Hyperglycemia (HG) induced significant increase in the activity of the citric acid cycle and led to elevated mitochondrial membrane potential. Mitochondrial oxidant production was increased and the mitochondrial electron transport decreased in hyperglycemic cells. AP39 and AP123 (30-300nM) decreased HG-induced hyperpolarisation of the mitochondrial membrane and inhibited the mitochondrial oxidant production. Both H2S donors (30-300nM) increased the electron transport at respiratory complex III and improved the cellular metabolism. Targeting H2S to mitochondria retained the cytoprotective effect of H2S against glucose-induced damage in endothelial cells suggesting that the molecular target of H2S action is within the mitochondria. Mitochondrial targeting of H2S also induced >1000-fold increase in the potency of H2S against hyperglycemia-induced injury. The high potency and long-lasting effect elicited by these H2S donors strongly suggests that these compounds could be useful against diabetic vascular complications.

  5. Effects ofPlasmodium falciparum-infected erythrocytes on matrix metalloproteinase-9 regulation in human microvascular endothelial cells

    Institute of Scientific and Technical Information of China (English)

    Sarah D Alessandro; Nicoletta Basilico; Mauro Prato

    2013-01-01

    Objective:To investigate the regulation of matrix metalloproteinases(MMPs) and tissue inhibitors of metalloproteinases(TIMPs) in human microvascular endothelium(HMEC-1) exposed to erythrocytes infected by different strains ofPlasmodium falciparum (P. falciparum).Methods:HMEC-1 cells were co-incubated for72 h with erythrocytes infected by late stage trophozoite of D10(chloroquine-sensitive) orW2(chloroquine-resistant)P. falciparum strains.Cell supernatants were then collected and the levels of pro- or active gelatinasesMMP-9 andMMP-2 were evaluated by gelatin zymography and densitometry.The release of pro-MMP-9,MMP-3,MMP-1 andTIMP-1 proteins was analyzed by western blotting and densitometry.Results:Infected erythrocytes inducedde novo proMMP-9 andMMP-9 release.Neither basal levels of proMMP-2 were altered, nor activeMMP-2 was found.MMP-3 andMMP-1 secretion was significantly enhanced, whereas basalTIMP-1 was unaffected.All effects were similar for both strains. Conclusions:P. falciparum parasites, either chloroquine-sensitive or -resistant, induce the release of activeMMP-9 protein from human microvascular endothelium, by impairing balances between proMMP-9 and its inhibitor, and by enhancing the levels of its activators.This work provides new evidence onMMP involvement in malaria, pointing atMMP-9 as a possible target in adjuvant therapy.

  6. Orally administered S-1 suppresses circulating endothelial cell counts in metastatic breast cancer patients.

    OpenAIRE

    2013-01-01

    [Background]S-1 is an oral cytotoxic preparation that contains tegafur. Gamma-butyrolactone (GBL) is a metabolite of tegafur that is known to suppress vascular endothelial growth factor (VEGF)-mediated angiogenic activity. The aim of this study was to determine the change in circulating endothelial cell (CEC) counts, GBL levels, and angiogenesis-related factors during S-1 administration in metastatic breast cancer (MBC) patients. [Methods]Patients with HER2-negative MBC were eligible. S-1 was...

  7. Expression and Clinical Significance of Vascular Endothelial Growth Factor in Benign and Malignant Tissues of Breast

    Institute of Scientific and Technical Information of China (English)

    XU Wei-gu; LIU Yang; WANG Gang; SONG Ji-ning; YANG Xiao-qing; WANG Wen-ya

    2008-01-01

    Objective:To detect the expression of vascular endothelial growth factor(VEGF)and microvessel density(MVD)count in breast benign affection,breast atypical hyperplasia and breast carcinoma in situ,and to clarify the relationship between VEGF expression,MVD and the clinicopathological features of these diseases. Methods:The expression of VEGF and MVD count in 115 cases breast benign diseases(including 40 breast fibroid tumor,40 breast cystic hyperplasia and 35 intraductal papilloma,19 breast atypical hyperplasias and 32 breast carcinomas in situ were examined by immunohistochemistry staining(SP-method). Results:The positive rate of VEGF in breast benign diseases,breast atypical hyperplasia and breast carcinoma in situ were 21.74%(25/115)、31.58.%(6/19)and 53.13%(17/32)respectively.It was the lowest in breast benign affection group,and was the highest breast carcinoma in situ group.The expression of VEGF increased gradually in the three groups(P<0.05).The MVD count of the three groups were 14.41±2.59,18.89±4.47 and 21.13±4.12 respectively,It was the lowest in breast benign affection group,and was the highest breast carcinoma in situ group.The MVD count of the three groups increased gradually(P<0.05).In VEGF positive group,MVD count was 19.41±4.78;In VEGF negative group,MVD count was 14.91±3.15.The MVD count was higher in VEGF positive group than that in VEGF negative group(P<0.05). Conclusion:The results of this study suggested that VEGF could promote microvessel growth in breast tumors.The occurrence and progression of breast cancer might be related with the expression of VEGF.

  8. Cytotoxicity and Proliferation Studies with Arsenic in Established Human Cell Lines: Keratinocytes, Melanocytes, Dendritic Cells, Dermal Fibroblasts, Microvascular Endothelial Cells, Monocytes and T-Cells

    Directory of Open Access Journals (Sweden)

    Hari H. P. Cohly

    2003-01-01

    Full Text Available Abstract: Based on the hypothesis that arsenic exposure results in toxicity and mitogenecity, this study examined the dose-response of arsenic in established human cell lines of keratinocytes (HaCaT, melanocytes (1675, dendritic cells (THP-1/A23187, dermal fibroblasts (CRL1904, microvascular endothelial cells (HMEC, monocytes (THP-1, and T cells (Jurkat. Cytotoxicity was determined by incubating THP-1, THP-1+ A23187 and JKT cells in RPMI 1640, 1675 in Vitacell, HMEC in EBM, and dermal fibroblasts and HaCaT in DMEM with 10% fetal bovine serum, 1% streptomycin and penicillin for 72 hrs in 96-well microtiter plates, at 37oC in a 5% CO2 incubator with different concentrations of arsenic using fluorescein diacetate (FDA. Cell proliferation in 96-well plates was determined in cultured cells starved by prior incubation for 24 hrs in 1% FBS and exposed for 72 hours, using the 96 cell titer proliferation solution (Promega assay. Cytotoxicity assays yielded LD50s of 9 μg/mL for HaCaT, 1.5 μg/mL for CRL 1675, 1.5 μg/mL for dendritic cells, 37 μg/mL for dermal fibroblasts, 0.48 μg/mL for HMEC, 50 μg/mL for THP-1 cells and 50 μg/mL for JKT-T cells. The peak proliferation was observed at 6 μg/mL for HaCaT and THP-1 cells, 0.19 μg/mL for CRL 1675, dendritic cells, and HMEC, and 1.5 μg/mL for dermal fibroblasts and Jurkat T cells. These results show that arsenic is toxic at high doses to keratinocytes, fibroblasts, monocytes and T cells, and toxic at lower doses to melanocytes, microvascular endothelial cells and dendritic cells. Proliferation studies showed sub-lethal doses of arsenic to be mitogenic.

  9. Atorvastatin prevents hypoxia-induced inhibition of endothelial nitric oxide synthase expression but does not affect heme oxygenase-1 in human microvascular endothelial cells

    NARCIS (Netherlands)

    Loboda, Agnieszka; Jazwa, Agnieszka; Jozkowicz, Alicj A.; Dorosz, Jerzy; Balla, Jozsef; Molema, Grietje; Dulak, Jozef

    2006-01-01

    Beneficial cardiovascular effects of statins, the inhibitors of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase, are particularly assigned to the modulation of inflammation. Endothelial nitric oxide synthase (eNOS) and heme oxygenase-1 (HO-1) are listed among the crucial protective, anti-i

  10. Breast cancer cells stimulate osteoprotegerin (OPG production by endothelial cells through direct cell contact

    Directory of Open Access Journals (Sweden)

    Holen Ingunn

    2009-07-01

    Full Text Available Abstract Background Angiogenesis, the sprouting of capillaries from existing blood vessels, is central to tumour growth and progression, however the molecular regulation of this process remains to be fully elucidated. The secreted glycoprotein osteoprotegerin (OPG is one potential pro-angiogenic factor, and clinical studies have demonstrated endothelial cells within a number of tumour types to express high levels of OPG compared to those in normal tissue. Additionally, OPG can increase endothelial cell survival, proliferation and migration, as well as induce endothelial cell tube formation in vitro. This study aims to elucidate the processes involved in the pro-angiogenic effects of OPG in vitro, and also how OPG levels may be regulated within the tumour microenvironment. Results It has previously been demonstrated that OPG can induce tube formation on growth factor reduced matrigel. In this study, we demonstrate that OPG enhances the pro-angiogenic effects of VEGF and that OPG does not stimulate endothelial cell tube formation through activation of the VEGFR2 receptor. We also show that cell contact between HuDMECs and the T47D breast cancer cell line increases endothelial cell OPG mRNA and protein secretion levels in in vitro co-cultures. These increases in endothelial cell OPG secretion were dependent on ανβ3 ligation and NFκB activation. In contrast, the pro-angiogenic factors VEGF, bFGF and TGFβ had no effect on HuDMEC OPG levels. Conclusion These findings suggest that the VEGF signalling pathway is not involved in mediating the pro-angiogenic effects of OPG on endothelial cells in vitro. Additionally, we show that breast cancer cells cause increased levels of OPG expression by endothelial cells, and that direct contact between endothelial cells and tumour cells is required in order to increase endothelial OPG expression and secretion. Stimulation of OPG secretion was shown to involve ανβ3 ligation and NFκB activation.

  11. Human Brain Microvascular Endothelial Cells Derived from the BC1 iPS Cell Line Exhibit a Blood-Brain Barrier Phenotype.

    Science.gov (United States)

    Katt, Moriah E; Xu, Zinnia S; Gerecht, Sharon; Searson, Peter C

    2016-01-01

    The endothelial cells that form capillaries in the brain are highly specialized, with tight junctions that minimize paracellular transport and an array of broad-spectrum efflux pumps that make drug delivery to the brain extremely challenging. One of the major limitations in blood-brain barrier research and the development of drugs to treat central nervous system diseases is the lack of appropriate cell lines. Recent reports indicate that the derivation of human brain microvascular endothelial cells (hBMECs) from human induced pluripotent stem cells (iPSCs) may provide a solution to this problem. Here we demonstrate the derivation of hBMECs extended to two new human iPSC lines: BC1 and GFP-labeled BC1. These hBMECs highly express adherens and tight junction proteins VE-cadherin, ZO-1, occludin, and claudin-5. The addition of retinoic acid upregulates VE-cadherin expression, and results in a significant increase in transendothelial electrical resistance to physiological values. The permeabilities of tacrine, rhodamine 123, and Lucifer yellow are similar to values obtained for MDCK cells. The efflux ratio for rhodamine 123 across hBMECs is in the range 2-4 indicating polarization of efflux transporters. Using the rod assay to assess cell organization in small vessels and capillaries, we show that hBMECs resist elongation with decreasing diameter but show progressive axial alignment. The derivation of hBMECs with a blood-brain barrier phenotype from the BC1 cell line highlights that the protocol is robust. The expression of GFP in hBMECs derived from the BC1-GFP cell line provides an important new resource for BBB research.

  12. Activation of melatonin receptor (MT1/2) promotes P-gp transporter in methamphetamine-induced toxicity on primary rat brain microvascular endothelial cells.

    Science.gov (United States)

    Jumnongprakhon, Pichaya; Sivasinprasasn, Sivanan; Govitrapong, Piyarat; Tocharus, Chainarong; Tocharus, Jiraporn

    2017-02-20

    Melatonin has been known as a neuroprotective agent for the central nervous system (CNS) and the blood-brain barrier (BBB), which is the primary structure that comes into contact with several neurotoxins including methamphetamine (METH). Previous studies have reported that the activation of melatonin receptors (MT1/2) by melatonin could protect against METH-induced toxicity in brain endothelial cells via several mechanisms. However, its effects on the P-glycoprotein (P-gp) transporter, the active efflux pump involved in cell homeostasis, are still unclear. Thus, this study investigated the role of melatonin and its receptors on the METH-impaired P-gp transporter in primary rat brain microvascular endothelial cells (BMVECs). The results showed that METH impaired the function of the P-gp transporter, significantly decreasing the efflux of Rho123 and P-gp expression, which caused a significant increase in the intracellular accumulation of Rho123, and these responses were reversed by the interaction of melatonin with its receptors. Blockade of the P-gp transporter by verapamil caused oxidative stress, apoptosis, and cell integrity impairment after METH treatment, and these effects could be reversed by melatonin. Our results, together with previous findings, suggest that the interaction of melatonin with its receptors protects against the effects of the METH-impaired P-gp transporter and that the protective role in METH-induced toxicity was at least partially mediated by the regulation of the P-gp transporter. Thus, melatonin and its receptors (MT1/2) are essential for protecting against BBB impairment caused by METH.

  13. Oxidative Stress Induced by Cigarette Smoke Extracts in Human Brain Cells (T98G) and Human Brain Microvascular Endothelial Cells (HBMEC) in Mono- and Co-Culture.

    Science.gov (United States)

    Kim, Ju-Hyeong; Cho, Myung-Haing; Choi, Kyung-Chul; Lee, Kyuhong; Kim, Kwang-Sik; Shim, Soon-Mi

    2015-01-01

    The objective of the current study was to examine oxidative stress induced by cigarette smoke extract (CSE) or cigarette smoke condensate (CSC) in human brain cells (T98G) and human brain microvascular endothelial cells (HBMEC) in mono- and co-culture systems. Cell viability of T98G cells exposed to CSC (0.05-4 mg/ml) was significantly decreased compared to CSE (0.025-20%). There were no marked differences between quantities of reactive oxygen species (ROS) generation by either CSE (2, 4, and 10%) or CSC (0.2, 0.4, and 0.8 mg/ml) treatment compared to control. However, a significant effect was noted in ROS generation following CSC incubation at 4mg/ml. Cellular integrity of HBMEC decreased to 74 and 64% within 120 h of exposure at the IC50 value of CSE and CSC, respectively. This study suggests that chronic exposure to cigarette smoking might initiate damage to the blood-brain barrier.

  14. In vitro model of cerebral ischemia by using brain microvascular endothelial cells derived from human induced pluripotent stem cells.

    Science.gov (United States)

    Kokubu, Yasuhiro; Yamaguchi, Tomoko; Kawabata, Kenji

    2017-04-29

    Brain-derived microvascular endothelial cells (BMECs), which play a central role in blood brain barrier (BBB), can be used for the evaluation of drug transport into the brain. Although human BMEC cell lines have already been reported, they lack original properties such as barrier integrity. Pluripotent stem cells (PSCs) can be used for various applications such as regenerative therapy, drug screening, and pathological study. In the recent study, an induction method of BMECs from PSCs has been established, making it possible to more precisely study the in vitro human BBB function. Here, using induced pluripotent stem (iPS) cell-derived BMECs, we examined the effects of oxygen-glucose deprivation (OGD) and OGD/reoxygenation (OGD/R) on BBB permeability. OGD disrupted the barrier function, and the dysfunction was rapidly restored by re-supply of the oxygen and glucose. Interestingly, TNF-α, which is known to be secreted from astrocytes and microglia in the cerebral ischemia, prevented the restoration of OGD-induced barrier dysfunction in an apoptosis-independent manner. Thus, we could establish the in vitro BBB disease model that mimics the cerebral ischemia by using iPS cell-derived BMECs.

  15. Statins suppress glucose-induced plasminogen activator inhibitor-1 expression by regulating RhoA and nuclear factor-κB activities in cardiac microvascular endothelial cells.

    Science.gov (United States)

    Ni, Xiao-Qing; Zhu, Jian-Hua; Yao, Ning-Hua; Qian, Juan; Yang, Xiang-Jun

    2013-01-01

    The aim of this study was to investigate the possible proinflammatory signaling pathways involved in statin inhibition of glucose-induced plasminogen activator inhibitor-1 (PAI-1) expression in cardiac microvascular endothelial cells (CMECs). Primary rat CMECs were grown in the presence of 5.7 or 23 mmol/L glucose. PAI-1 mRNA and protein expression levels were measured by realtime polymerase chain reaction, Western blotting and enzyme-linked immunosorbent assay, respectively. A pull-down assay was performed to determine RhoA activity. IκBα protein expression was measured by Western blotting, nuclear factor (NF)-κB activation was detected by electrophoretic mobility shift assay and its transcription activity was determined by a dual luciferase reporter gene assay. PAI-1 mRNA and protein expression levels were both increased with high glucose concentrations, but they were significantly suppressed by simvastatin and atorvastatin treatment (P statins (P statins may occur partly by regulating the RhoA/ROCK-NF-κB pathway. The multifunctional roles of statins may be particularly beneficial for patients with metabolic syndrome.

  16. Culture and Identification of microvascular endothelial cells from human ovarian carcinoma%人卵巢癌微血管内皮细胞的培养与鉴定

    Institute of Scientific and Technical Information of China (English)

    徐燕; 梁志清; 王丹; 谢尧

    2009-01-01

    目的 建立人卵巢癌微血管内皮细胞(ovarian carcinoma-derived microvascular endothelial cells,ODMECs)体外培养体系.方法 采用胶原酶、胰酶联合消化法分离微血管内皮,经percoll梯度密度离心纯化.光镜、电镜、流式细胞术、免疫细胞化学对所获得的ODMECs进行鉴定.结果 所获ODMECs流式分析有内皮标志物CD34/VEGF-R2表达;免疫荧光FⅦ-RAg阳性;电镜下见细胞多核、有丰富的微丝、Weibel-Palade小体等;培养中的内皮细胞生长状态良好、呈现典型的铺路石征,可传代培养.结论 本研究建立了人ODMECs体外培养体系,对了解卵巢癌血管内皮的异质性有重要价值,为抗卵巢癌血管生成的研究奠定了基础.

  17. Differentially expressed genes of human microvascular endothelial cells in response to anti-dengue virus NS1 antibodies by suppression subtractive hybridization.

    Science.gov (United States)

    Yin, Yue; Jiang, Lan; Fang, Danyun; Jiang, Lifang; Zhou, Junmei

    2013-06-01

    It has been previously shown that anti-dengue virus (DENV) nonstructural protein NS1 antibodies could act as autoantibodies that direct against one or more of the host's own proteins, which has potential implications for dengue hemorrhagic fever pathogenesis. In the present study, we have employed suppression subtractive hybridization (SSH) to identify the differentially expressed genes from human microvascular endothelial cells (HMEC-1) in response to anti-dengue virus type 2 NS1 antibodies (anti-DENV2 NS1 Abs). A total of 35 clones from the SSH cDNA library were randomly selected for further analysis using bioinformatics tools after vector screening. After searching for sequence homology in NCBI GenBank database with BLASTN and BLASTX programs, 23 obtained sequences with significant matches (E-values <1×10(-4)) in the SSH library. The predicted genes in the subtracted library include immune response molecules (CD59 antigen preproprotein preproprotein, MURR1), signal transduction molecules (Nuclear casein kinase and cyclin-dependent kinase substrate 1), calcium-binding proteins (S100A6, Annexin A2 isoform 1/2), and cell-membrane component (Yip1 domain family). From these clones, 5 upregulated genes were selected for differential expression profiling by real-time RT-PCR to confirm their upregulated status. The results confirmed their differential upregulation, and thus verified the success of SSHs and the likely involvement of these genes in dengue pathogenesis.

  18. Tanshinone IIA inhibits endothelin-1 production in TNF-α-induced brain microvascular endothelial cells through suppression of endothelin-converting enzyme-1 synthesis

    Institute of Scientific and Technical Information of China (English)

    Chao TANG; An-hua WU; Hong-li XUE; Yun-jie WANG

    2007-01-01

    Aim: To investigate the effects of tanshinone ⅡA (Tan ⅡA) on the regulation of the production of endothelin (ET)-1 (including large ET-1), mRNA levels of ET-1,endothelin-converting enzyme- 1 (ECE- 1), endothelin-A receptor (ETA) and endothelin-B receptor (ETB) induced by TNF-α in rat brain microvascular endo-thelial cells (BMVEC). Methods: The ET-1 release (including large ET-1) into the culture medium was determined by enzyme immunoassay. The levels of ET-1,ECE- 1, ETA, and ETB mRNA were measured by RT-PCR. Endothelin receptor bind-ing was also tested. Results: The induction of ET- 1 release by TNF-α from cul-tured BMVEC was dose-dependently reduced by Tan IIA, but large ET-1 levels progressively increased in response to Tan IIA; the mRNA expression of ET-1 was unaffected. Tan ⅡA also caused a decrease in ETA receptor mRNA and ECE-1expression in a dose-dependent manner. Endothelin receptor binding was unal-tered in BMVEC stimulated with TNF-α alone or a combination of TNF-α and Tan ⅡA. Conclusion: These findings suggest that Tan ⅡA may inhibit ET-1 produc-tion in TNF-α-induced BMVEC through the suppression of ECE-1 synthesis.

  19. Modulation of human dermal microvascular endothelial cell and human gingival fibroblast behavior by micropatterned silica coating surfaces for zirconia dental implant applications

    Science.gov (United States)

    Laranjeira, Marta S; Carvalho, Ângela; Pelaez-Vargas, Alejandro; Hansford, Derek; Ferraz, Maria Pia; Coimbra, Susana; Costa, Elísio; Santos-Silva, Alice; Fernandes, Maria Helena; Monteiro, Fernando Jorge

    2014-01-01

    Dental ceramic implants have shown superior esthetic behavior and the absence of induced allergic disorders when compared to titanium implants. Zirconia may become a potential candidate to be used as an alternative to titanium dental implants if surface modifications are introduced. In this work, bioactive micropatterned silica coatings were produced on zirconia substrates, using a combined methodology of sol–gel processing and soft lithography. The aim of the work was to compare the in vitro behavior of human gingival fibroblasts (HGFs) and human dermal microvascular endothelial cells (HDMECs) on three types of silica-coated zirconia surfaces: flat and micropatterned (with pillars and with parallel grooves). Our results showed that cells had a higher metabolic activity (HGF, HDMEC) and increased gene expression levels of fibroblast-specific protein-1 (FSP-1) and collagen type I (COL I) on surfaces with pillars. Nevertheless, parallel grooved surfaces were able to guide cell growth. Even capillary tube-like networks of HDMEC were oriented according to the surface geometry. Zirconia and silica with different topographies have shown to be blood compatible and silica coating reduced bacteria adhesion. All together, the results indicated that microstructured bioactive coating seems to be an efficient strategy to improve soft tissue integration on zirconia implants, protecting implants from peri-implant inflammation and improving long-term implant stabilization. This new approach of micropatterned silica coating on zirconia substrates can generate promising novel dental implants, with surfaces that provide physical cues to guide cells and enhance their behavior. PMID:27877662

  20. Modulation of human dermal microvascular endothelial cell and human gingival fibroblast behavior by micropatterned silica coating surfaces for zirconia dental implant applications

    Directory of Open Access Journals (Sweden)

    Marta S Laranjeira

    2014-03-01

    Full Text Available Dental ceramic implants have shown superior esthetic behavior and the absence of induced allergic disorders when compared to titanium implants. Zirconia may become a potential candidate to be used as an alternative to titanium dental implants if surface modifications are introduced. In this work, bioactive micropatterned silica coatings were produced on zirconia substrates, using a combined methodology of sol–gel processing and soft lithography. The aim of the work was to compare the in vitro behavior of human gingival fibroblasts (HGFs and human dermal microvascular endothelial cells (HDMECs on three types of silica-coated zirconia surfaces: flat and micropatterned (with pillars and with parallel grooves. Our results showed that cells had a higher metabolic activity (HGF, HDMEC and increased gene expression levels of fibroblast-specific protein-1 (FSP-1 and collagen type I (COL I on surfaces with pillars. Nevertheless, parallel grooved surfaces were able to guide cell growth. Even capillary tube-like networks of HDMEC were oriented according to the surface geometry. Zirconia and silica with different topographies have shown to be blood compatible and silica coating reduced bacteria adhesion. All together, the results indicated that microstructured bioactive coating seems to be an efficient strategy to improve soft tissue integration on zirconia implants, protecting implants from peri-implant inflammation and improving long-term implant stabilization. This new approach of micropatterned silica coating on zirconia substrates can generate promising novel dental implants, with surfaces that provide physical cues to guide cells and enhance their behavior.

  1. Modulation of human dermal microvascular endothelial cell and human gingival fibroblast behavior by micropatterned silica coating surfaces for zirconia dental implant applications

    Science.gov (United States)

    Laranjeira, Marta S.; Carvalho, Ângela; Pelaez-Vargas, Alejandro; Hansford, Derek; Ferraz, Maria Pia; Coimbra, Susana; Costa, Elísio; Santos-Silva, Alice; Fernandes, Maria Helena; Monteiro, Fernando Jorge

    2014-04-01

    Dental ceramic implants have shown superior esthetic behavior and the absence of induced allergic disorders when compared to titanium implants. Zirconia may become a potential candidate to be used as an alternative to titanium dental implants if surface modifications are introduced. In this work, bioactive micropatterned silica coatings were produced on zirconia substrates, using a combined methodology of sol-gel processing and soft lithography. The aim of the work was to compare the in vitro behavior of human gingival fibroblasts (HGFs) and human dermal microvascular endothelial cells (HDMECs) on three types of silica-coated zirconia surfaces: flat and micropatterned (with pillars and with parallel grooves). Our results showed that cells had a higher metabolic activity (HGF, HDMEC) and increased gene expression levels of fibroblast-specific protein-1 (FSP-1) and collagen type I (COL I) on surfaces with pillars. Nevertheless, parallel grooved surfaces were able to guide cell growth. Even capillary tube-like networks of HDMEC were oriented according to the surface geometry. Zirconia and silica with different topographies have shown to be blood compatible and silica coating reduced bacteria adhesion. All together, the results indicated that microstructured bioactive coating seems to be an efficient strategy to improve soft tissue integration on zirconia implants, protecting implants from peri-implant inflammation and improving long-term implant stabilization. This new approach of micropatterned silica coating on zirconia substrates can generate promising novel dental implants, with surfaces that provide physical cues to guide cells and enhance their behavior.

  2. Photodynamic efficacy of liposome-delivered hypocrellin B in microvascular endothelial cells in vitro and chicken combs in vivo: a potential photosensitizer for port wine stain

    Science.gov (United States)

    Chen, H. X.; Yang, Z. F.; Zou, X. B.; Zhu, J. G.; Deng, H.; Zhao, J. Q.; Gu, Y.

    2013-02-01

    Photodynamic therapy (PDT) has been proved a successful method for port wine stain (PWS), but the prolonged skin photosensitivity induced by the photosensitizers used currently seriously limits the clinical application of PDT. In this study, we investigate the feasibility of hypocrellin B (HB), a promising second-generation photosensitizer for the treatment of PWS. The photodynamic effect of liposome-delivered HB was evaluated in vitro with microvascular endothelial cells (MEC) and in vivo with chicken combs. The dark cytotoxicity and photocytotoxicity of liposomal HB in MEC were evaluated using the MTT assay. Gross and histological examinations were performed to investigate the selective occlusion of the superficial dermal microvasculature in the chicken comb. The result showed that photocytotoxicity of liposomal HB was dependent on both light dose and drug concentration. PDT with HB (0.5-1 mg kg-1) and a light dose of 120 J cm-2 showed selective destruction of the superficial dermal microvasculature of the chicken comb, leaving the overlying epidermis intact. This is the first study to investigate the potential efficacy of HB-PDT as a novel modality for the treatment of PWS. These findings suggest that liposomal HB is a safe and effective photosensitizer for PWS.

  3. Effects of mir-21 on Cardiac Microvascular Endothelial Cells After Acute Myocardial Infarction in Rats: Role of Phosphatase and Tensin Homolog (PTEN)/Vascular Endothelial Growth Factor (VEGF) Signal Pathway

    Science.gov (United States)

    Yang, Feng; Liu, Wenwei; Yan, Xiaojuan; Zhou, Hanyun; Zhang, Hongshen; Liu, Jianfei; Yu, Ming; Zhu, Xiaoshan; Ma, Kezhong

    2016-01-01

    Background This study investigated how miR-21 expression is reflected in acute myocardial infarction and explored the role of miR-21 and the PTEN/VEGF signaling pathway in cardiac microvascular endothelial cells. Material/Methods We used an in vivo LAD rat model to simulate acute myocardial infarction. MiR-21 mimics and miR-21 inhibitors were injected and transfected into model rats in order to alter miR-21 expression. Cardiac functions were evaluated using echocardiographic measurement, ELISA, and Masson staining. In addition, lenti-PTEN and VEGF siRNA were transfected into CMEC cells using standard procedures for assessing the effect of PTEN and VEGE on cell proliferation, apoptosis, and angiogenesis. MiR-21, PTEN, and VEGF expressions were examined by RT-PCR and Western blot. The relationship between miR-21 and PTEN was determined by the luciferase activity assay. Results We demonstrated that miR-21 bonded with the 3′-UTR of PTEN and suppressed PTEN expressions. Established models significantly induced cardiac infarct volume and endothelial injury marker expressions as well as miR-21 and PTEN expressions (PMiR-21 mimics exhibited significantly protective effects since they down-regulated both infarction size and injury marker expressions by increasing VEGF expression and inhibiting PTEN expression (PmiR-21 on cell proliferation, apoptosis, and angiogenesis (PMiR-21 exerts protective effects on endothelial injury through the PTEN/VEGF pathway after acute myocardial infarction. PMID:27708252

  4. Destruction of human microvascular endothelial cell capillary-like microtubules by Brazilian purpuric fever-associated Haemophilus influenzae biogroup aegyptius.

    Science.gov (United States)

    Quinn, F D; Weyant, R S; Candal, F J; Ades, E W

    1994-01-01

    When grown in the presence of Matrigel, monolayers of an immortalized human microvascular cell line (HMEC-1) form capillary-like microtubule networks. Previous work, using HMEC-1 monolayers, demonstrated a significant difference in in vitro cytotoxicity between Brazilian purpuric fever (BPF)-associated Haemophilus influenzae biogroup aegyptius (HAE) strains and non-BPF-associated HAE strains. The present study demonstrates that BPF-related cytotoxic differences can also be observed in HMEC-1 microtubule networks. At a multiplicity of infection (MOI) of 2 x 10(-2) bacteria/tissue culture cell, BPF-associated strain F3031 disrupted the microtubule network, producing random clumps of rounded cells at 48 h of incubation. Infection with non-BPF-associated strain F1947 at the same MOI produced no observable microtubule disruption. The ability of HMEC-1 microtubule model to differentiate virulent and avirulent HAE in vitro will further aid in the study of BPF pathogenesis. In addition, the fact that the HMEC-1 cells can be induced to form microtubules make it an excellent model system for the general study of many of the agents of vascular purpura.

  5. Repeatability of the evaluation of systemic microvascular endothelial function using laser doppler perfusion monitoring: clinical and statistical implications

    Directory of Open Access Journals (Sweden)

    Eduardo Tibiriçá

    2011-01-01

    Full Text Available OBJECTIVE: An awareness of the repeatability of biological measures is required to properly design and calculate sample sizes for longitudinal interventional studies. We investigated the day-to-day repeatability of measures of systemic microvascular reactivity using laser Doppler perfusion monitoring. METHODS: We performed laser Doppler perfusion monitoring in combination with skin iontophoresis using acetylcholine and sodium nitroprusside as well as post-occlusive reactive and thermal hyperemia twice within two weeks. The repeatability was assessed by calculating the within-subject standard deviations, limits of agreement, typical errors and intra-class correlation coefficients between days 1 and 2. The ratio of the within-subject standard deviation to the mean values obtained on days 1 and 2 (within-subject standard deviation/GM was used to determine the condition with the best repeatability. RESULTS: Twenty-four healthy subjects, aged 24.6 + 3.8 years, were recruited. The area under the curve of the vasodilatory response to post-occlusive reactivity showed marked variability (within-subject standard deviation/GM = 0.83, while the area under the curve for acetylcholine exhibited less variability (within-subject standard deviation/ GM = 0.52 and was comparable to the responses to sodium nitroprusside and thermal treatment (within-subject standard deviations/GM of 0.67 and 0.56, respectively. The area under the blood flow/time curve for vasodilation during acetylcholine administration required the smallest sample sizes, the area under the blood flow/time curve during post-occlusive reactivity required the largest sample sizes, and the area under the blood flow/time curves of vasodilation induced by sodium nitroprusside and thermal treatment required intermediate sizes. CONCLUSIONS: In view of the importance of random error related to the day-to-day repeatability of laser Doppler perfusion monitoring, we propose an original and robust statistical

  6. Signaling mechanisms in tumor necrosis factor alpha-induced death of microvascular endothelial cells of the corpus luteum

    Directory of Open Access Journals (Sweden)

    Rueda Bo R

    2003-02-01

    Full Text Available Abstract The microvasculature of the corpus luteum (CL, which comprises greater than 50% of the total number of cells in the CL, is thought to be the first structure to undergo degeneration via apoptosis during luteolysis. These studies compared the apoptotic potential of various cytokines (tumor necrosis factor α, TNFα; interferon gamma, IFNγ; soluble Fas ligand, sFasL, a FAS activating antibody (FasAb, and the luteolytic hormone prostaglandin F2α (PGF2α on CL-derived endothelial (CLENDO cells. Neither sFasL, FasAb nor PGF2α had any effect on CLENDO cell viability. Utilizing morphological and biochemical parameters it was evident that TNFα and IFNγ initiated apoptosis in long-term cultures. However, TNFα was the most potent stimulus for CLENDO cell apoptosis at early time points. Unlike many other studies described in non-reproductive cell types, TNFα induced apoptosis of CLENDO cells occurs in the absence of inhibitors of protein synthesis. TNFα-induced death is typically associated with acute activation of distinct intracellular signaling pathways (e.g. MAPK and sphingomyelin pathways. Treatment with TNFα for 5–30 min activated MAPKs (ERK, p38, and JNK, and increased ceramide accumulation. Ceramide, a product of sphingomyelin hydrolysis, can serve as an upstream activator of members of the MAPK family independently in numerous cell types, and is a well-established pro-apoptotic second messenger. Like TNFα, treatment of CLENDO cells with exogenous ceramide significantly induced endothelial apoptosis. Ceramide also activated the JNK pathway, but had no effect on ERK and p38 MAPKs. Pretreatment of CLENDO cells with glutathione (GSH, an intracellular reducing agent and known inhibitor of reactive oxygen species (ROS or TNFα-induced apoptosis, significantly attenuated TNFα-induced apoptosis. It is hypothesized that TNFα kills CLENDO cells through elevation of reactive oxygen species, and intracellular signals that promote

  7. Endothelial Cell Migration and Vascular Endothelial Growth Factor Expression Are the Result of Loss of Breast Tissue Polarity

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Amy; Cuevas, Ileana; Kenny, Paraic A; Miyake, Hiroshi; Mace, Kimberley; Ghajar, Cyrus; Boudreau, Aaron; Bissell, Mina; Boudreau, Nancy

    2009-05-26

    Recruiting a new blood supply is a rate-limiting step in tumor progression. In a three-dimensional model of breast carcinogenesis, disorganized, proliferative transformed breast epithelial cells express significantly higher expression of angiogenic genes compared with their polarized, growth-arrested nonmalignant counterparts. Elevated vascular endothelial growth factor (VEGF) secretion by malignant cells enhanced recruitment of endothelial cells (EC) in heterotypic cocultures. Significantly, phenotypic reversion of malignant cells via reexpression of HoxD10, which is lost in malignant progression, significantly attenuated VEGF expression in a hypoxia-inducible factor 1{alpha}-independent fashion and reduced EC migration. This was due primarily to restoring polarity: forced proliferation of polarized, nonmalignant cells did not induce VEGF expression and EC recruitment, whereas disrupting the architecture of growth-arrested, reverted cells did. These data show that disrupting cytostructure activates the angiogenic switch even in the absence of proliferation and/or hypoxia and restoring organization of malignant clusters reduces VEGF expression and EC activation to levels found in quiescent nonmalignant epithelium. These data confirm the importance of tissue architecture and polarity in malignant progression.

  8. Vascular endothelial growth factor and microvessel density for detection and prognostic evaluation of invasive breast cancer

    Institute of Scientific and Technical Information of China (English)

    Lukui Yang; Long Li; Xiangyu Cui; Dalei Yang

    2015-01-01

    Objective The purpose of this study was to evaluate the distribution of vascular endothelial growth factor (VEGF) and CD105-microvessel density (MVD) in invasive breast carcinomas. We also aimed to analyze the relationship between VEGF and MVD expression with other standard prognostic parameters associated with invasive breast cancer, such as size, grade, stage of the cancer, metastases, and tumor recurrence. Methods Immunohistochemistry via the Ultra SensitiveTM S-P method was used to detect VEGF and MVD expression in 128 cases of invasive breast carcinoma. Specimens were evaluated for CD105 expres-sion. Positively stained microvessels were counted in dense vascular foci under 400× magnification. MVD in the peripheral area adjacent to the lesion and in the central area within the lesion in invasive breast carcinomas and benign leisions groups were also assessed. Fifty cases of benign breast disease tissue were selected as the control group. Results Results showed that 64.1% of invasive breast cancer samples were VEGF-positive, higher than in benign breast disease tissue (22.0%, P 0.05). MVD of the peripheral area adja-cent to the lesion was significantly higher than those central area within the lesion in both invasive breast cancer and benign breast disease groups (P 50 years) or the two tumor diameter groups (≤2 cm vs.>2 cm), P > 0.05. Conclusion Overexpression of VEGF and MVD may be important biological markers for invasion and lymph node and distant metastases of invasive breast cancer. Combined detection of the two tumor mark-ers could provide better prognostic monitoring for disease recurrence and metastasis, as wel as aid with clinical staging of breast tumors. Prediction of the risk for metastasis and recurrence, as wel as recurrence patterns based on VEGF and MVD post-surgery, could aid design of better fol ow-up regimens and appro-priate treatment strategies for breast cancer patients.

  9. WSS25 inhibits Dicer, downregulating microRNA-210, which targets Ephrin-A3, to suppress human microvascular endothelial cell (HMEC-1) tube formation.

    Science.gov (United States)

    Xiao, Fei; Qiu, Hong; Zhou, Ling; Shen, Xiaokun; Yang, Liping; Ding, Kan

    2013-05-01

    WSS25 is a sulfated polysaccharide that inhibits angiogenesis. However, the mechanism underlying the regulation of angiogenesis by WSS25 is not well understood. Using microRNA (miRNA) microarray analysis, a total of 25 miRNAs were found to be upregulated and 12 (including miR-210) downregulated by WSS25 in human microvascular endothelial cells (HMEC-1). Interestingly, Dicer, a key enzyme for miRNA biosynthesis, was downregulated by WSS25 in HMEC-1 cells. Further studies indicated that HMEC-1 cell tube formation and miR-210 expression were suppressed while Ephrin-A3 expression was enhanced by the silencing of Dicer. In contrast, HMEC-1 cell tube formation and miR-210 expression were induced while Ephrin-A3 expression was suppressed by Dicer overexpression. Moreover, miR-210 was downregulated while Ephrin-A3 was upregulated by WSS25 in HMEC-1 cells. HMEC-1 cell migration and tube formation were arrested, while Ephrin-A3 expression was augmented by anti-miR-210. In addition, HMEC-1 cell tube formation was significantly attenuated or augmented when Ephrin-A3 was overexpressed or silenced, respectively. Nevertheless, the tube formation blocked by WSS25 could be partially rescued by manipulation of Dicer, miR-210 and Ephrin-A3. These results suggest a new pathway whereby WSS25 inhibits angiogenesis via suppression of Dicer, leading to downregulation of miR-210 and upregulation of Ephrin-A3.

  10. Aryl hydrocarbon Receptor is Necessary to Protect Fetal Human Pulmonary Microvascular Endothelial Cells against Hyperoxic Injury: Mechanistic Roles of Antioxidant Enzymes and RelB

    Science.gov (United States)

    Zhang, Shaojie; Patel, Ananddeep; Chu, Chun; Jiang, Weiwu; Wang, Lihua; Welty, Stephen E.; Moorthy, Bhagavatula; Shivanna, Binoy

    2015-01-01

    Hyperoxia contributes to the development of bronchopulmonary dysplasia (BPD) in premature infants. Activation of the aryl hydrocarbon receptor (AhR) protects adult and newborn mice against hyperoxic lung injury by mediating increases in the expression of phase I (cytochrome P450 (CYP) 1A) and phase II (NADP(H) quinone oxidoreductase (NQO1)) antioxidant enzymes (AOE). AhR positively regulates the expression of RelB, a component of the nuclear factor-kappaB (NF-κB) protein that contributes to anti-inflammatory processes in adult animals. Whether AhR regulates the expression of AOE and RelB, and protects fetal primary human lung cells against hyperoxic injury is unknown. Therefore, we tested the hypothesis that AhR-deficient fetal human pulmonary microvascular endothelial cells (HPMEC) will have decreased RelB activation and AOE, which will in turn predispose them to increased oxidative stress, inflammation, and cell death compared to AhR-sufficient HPMEC upon exposure to hyperoxia. AhR-deficient HPMEC showed increased hyperoxia-induced reactive oxygen species (ROS) generation, cleavage of poly (ADP-ribose) polymerase (PARP), and cell death compared to AhR-sufficient HPMEC. Additionally, AhR-deficient cell culture supernatants displayed increased macrophage inflammatory protein 1α and 1β, indicating a heightened inflammatory state. Interestingly, loss of AhR was associated with a significantly attenuated CYP1A1, NQO1, superoxide dismutase 1(SOD1), and nuclear RelB protein expression. These findings support the hypothesis that decreased RelB activation and AOE in AhR-deficient cells is associated with increased hyperoxic injury compared to AhR-sufficient cells. PMID:25831079

  11. Development and Validation of an In-Cell Western for Quantifying P-Glycoprotein Expression in Human Brain Microvascular Endothelial (hCMEC/D3) Cells.

    Science.gov (United States)

    McInerney, Mitchell P; Pan, Yijun; Short, Jennifer L; Nicolazzo, Joseph A

    2017-01-05

    An in-cell western (ICW) protocol detecting the relative expression of P-glycoprotein (P-gp) in human cerebro-microvascular endothelial cells (hCMEC/D3) was developed and optimized, with the intention of improving throughput relative to western blotting (WB). For validation of the ICW protocol, hCMEC/D3 cells were incubated with known P-gp upregulators (10 μM rifampicin and 5 μM SR12813) and treated with siRNA targeted against MDR1, before measuring changes in P-gp expression, using both ICW and WB in parallel. To confirm a relationship between the detected P-gp expression and function, the uptake of the P-gp substrate rhodamine-123 was assessed following SR12813 treatment. Rifampicin and SR12813 significantly upregulated P-gp expression (1.5-fold and 1.9-fold, respectively) compared to control, as assessed by the ICW protocol. WB analysis of the same treatments revealed 1.4-fold and 1.5-fold upregulations. MDR1 siRNA reduced P-gp abundance by 20% and 35% when assessed by ICW and WB, respectively. SR12813 treatment reduced rhodamine-123 uptake by 18%, indicating that the observed changes in P-gp expression by ICW were associated with comparable functional changes. The correlation of P-gp upregulation by WB, rhodamine-123 uptake, and the ICW protocol provide validation of a new ICW method as an alternative method for quantification of P-gp in hCMEC/D3 cells.

  12. Endothelial progenitor cells, microvascular obstruction, and left ventricular remodeling in patients with ST elevation myocardial infarction undergoing primary percutaneous coronary intervention.

    Science.gov (United States)

    Porto, Italo; De Maria, Giovanni Luigi; Leone, Antonio Maria; Dato, Ilaria; D'Amario, Domenico; Burzotta, Francesco; Niccoli, Giampaolo; Trani, Carlo; Biasucci, Luigi Marzio; Bolognese, Leonardo; Crea, Filippo

    2013-09-15

    Endothelial progenitor cells (EPCs) are released from the bone marrow during cardiac ischemic events, potentially influencing vascular and myocardial repair. We assessed the clinical and angiographic correlates of EPC mobilization at the time of primary percutaneous coronary intervention in 78 patients with ST elevation myocardial infarction and the impact of both baseline and follow-up EPC levels on left ventricular (LV) remodeling. Blood samples were drawn from the aorta and the culprit coronary artery for cytofluorimetric EPC detection (CD34+CD45dimKDR+ cells, in percentage of cytofluorimetric counts). Area at risk was assessed by Bypass Angioplasty Revascularization Investigation myocardial jeopardy index, thrombotic burden as thrombus score and microvascular obstruction (MVO) as a combination of ST segment resolution and myocardial blush grade. Echocardiographic evaluation of LV remodeling was performed at 1-year follow-up in 54 patients, whereas peripheral EPC levels were reassessed in 40 patients. EPC levels during primary percutaneous coronary intervention were significantly higher in intracoronary than in aortic blood (0.043% vs 0.0006%, p <0.001). Both intracoronary and aortic EPC were related to area at risk extent, to intracoronary thrombus score (p <0.001), and inversely to MVO (p = 0.001). Peripheral EPC levels at 1-year follow-up were lower in patients with LV remodeling than in those without (0.001% [0.001 to 0.002] vs 0.003% [0.002 to 0.010]; p = 0.01) and independently predicted absence of remodeling at multivariate analysis. In conclusion, a rapid intracoronary EPC recruitment takes place in the early phases of ST elevation myocardial infarction, possibly reflecting an attempted reparative response. The extent of this mobilization seems to be correlated to the area at risk and to the amount of MVO. Persistently low levels of EPC are associated to LV remodeling.

  13. Effects of antioxidant and NF-κB on the induction of iNOS gene in rat pulmonary microvascular endothelial cells in vitro

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    AIM:To investigate the role of NF-κB in the activation of inducible nitric oxide synthase (iNOS) gene by tumor necrosis factor alpha (TNF α) and lipopolysaccharide (LPS) in endothelial cells and effect of antioxidant on the induction of iNOS. METHODS:Rat pulmonary microvascular endothelial cell (RPMEC) was cultured and the cells were identified with antiendothelial cell antibody CD31 using immunohistochemistry(ABC). The concentration of nitrite in the culture media was determined based on Griess reaction. iNOS mRNA was analyzed using RT-PCR and Northern blot. NF-κB in cell nuclei was detected with electrophoresis mobility shift assay (EMSA). RESULTS:A marked production of nitrite in RPMECs was found after 24 hours treatment with TNF α(105 U/L) and LPS (1 mg/L) (P<0.01). The level of iNOS mRNA increased significantly after adding TNF α(105 U/L) and LPS (1 mg/L) to the cell media for 2 hours (P<0.05). Pretreatment with cycloheximide (CHX, 10 mg/L) or antioxidant, PDTC (0.1 mmol/L) or NAC (20 mmol/L) significantly decreased nitrite production and iNOS mRNA expression induced by TNF α(105 U/L) and LPS (1 mg/L) (P<0.05). Furthermore, there was a dose-effect relationship between PDTC/NAC and inhibitory effect. TNF α (105 U/L) and LPS (1 mg/L) triggered the activation and translocation of NF-κB. This effect was blocked by adding PDTC (0.1 mmol/L) or NAC (20 mmol/L) to the cell media for 1.5 hours.CONCLUSION:1.TNFα and LPS may induce iNOS gene expression at transcriptional or posttranscriptional level. The upregulation of iNOS depends on new protein synthesis. 2. The induction of iNOS gene expression by TNFα and LPS is dependent on the activation of NF-κB. 3. Antioxidants may inhibit the induction of iNOS gene through the inhibition of NF-κB activation.

  14. Regulation of human cerebro-microvascular endothelial baso-lateral adhesion and barrier function by S1P through dual involvement of S1P1 and S1P2 receptors.

    Science.gov (United States)

    Wiltshire, Rachael; Nelson, Vicky; Kho, Dan Ting; Angel, Catherine E; O'Carroll, Simon J; Graham, E Scott

    2016-01-27

    Herein we show that S1P rapidly and acutely reduces the focal adhesion strength and barrier tightness of brain endothelial cells. xCELLigence biosensor technology was used to measure focal adhesion, which was reduced by S1P acutely and this response was mediated through both S1P1 and S1P2 receptors. S1P increased secretion of several pro-inflammatory mediators from brain endothelial cells. However, the magnitude of this response was small in comparison to that mediated by TNFα or IL-1β. Furthermore, S1P did not significantly increase cell-surface expression of any key cell adhesion molecules involved in leukocyte recruitment, included ICAM-1 and VCAM-1. Finally, we reveal that S1P acutely and dynamically regulates microvascular endothelial barrier tightness in a manner consistent with regulated rapid opening followed by closing and strengthening of the barrier. We hypothesise that the role of the S1P receptors in this process is not to cause barrier dysfunction, but is related to controlled opening of the endothelial junctions. This was revealed using real-time measurement of barrier integrity using ECIS ZΘ TEER technology and endothelial viability using xCELLigence technology. Finally, we show that these responses do not occur simply though the pharmacology of a single S1P receptor but involves coordinated action of S1P1 and S1P2 receptors.

  15. Aryl hydrocarbon receptor is necessary to protect fetal human pulmonary microvascular endothelial cells against hyperoxic injury: Mechanistic roles of antioxidant enzymes and RelB

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Shaojie; Patel, Ananddeep; Chu, Chun; Jiang, Weiwu; Wang, Lihua; Welty, Stephen E.; Moorthy, Bhagavatula; Shivanna, Binoy, E-mail: shivanna@bcm.edu

    2015-07-15

    Hyperoxia contributes to the development of bronchopulmonary dysplasia (BPD) in premature infants. Activation of the aryl hydrocarbon receptor (AhR) protects adult and newborn mice against hyperoxic lung injury by mediating increases in the expression of phase I (cytochrome P450 (CYP) 1A) and phase II (NADP(H) quinone oxidoreductase (NQO1)) antioxidant enzymes (AOE). AhR positively regulates the expression of RelB, a component of the nuclear factor-kappaB (NF-κB) protein that contributes to anti-inflammatory processes in adult animals. Whether AhR regulates the expression of AOE and RelB, and protects fetal primary human lung cells against hyperoxic injury is unknown. Therefore, we tested the hypothesis that AhR-deficient fetal human pulmonary microvascular endothelial cells (HPMEC) will have decreased RelB activation and AOE, which will in turn predispose them to increased oxidative stress, inflammation, and cell death compared to AhR-sufficient HPMEC upon exposure to hyperoxia. AhR-deficient HPMEC showed increased hyperoxia-induced reactive oxygen species (ROS) generation, cleavage of poly(ADP-ribose) polymerase (PARP), and cell death compared to AhR-sufficient HPMEC. Additionally, AhR-deficient cell culture supernatants displayed increased macrophage inflammatory protein 1α and 1β, indicating a heightened inflammatory state. Interestingly, loss of AhR was associated with a significantly attenuated CYP1A1, NQO1, superoxide dismutase 1(SOD1), and nuclear RelB protein expression. These findings support the hypothesis that decreased RelB activation and AOE in AhR-deficient cells is associated with increased hyperoxic injury compared to AhR-sufficient cells. - Highlights: • AhR deficiency potentiates oxygen toxicity in human fetal lung cells. • Deficient AhR signaling increases hyperoxia-induced cell death. • AhR deficiency increases hyperoxia-induced ROS generation and inflammation. • Anti-oxidant enzyme levels are attenuated in AhR-deficient lung cells

  16. Angiotensin II induces apoptosis of human pulmonary microvascular endothelial cells in acute aortic dissection complicated with lung injury patients through modulating the expression of monocyte chemoattractant protein-1.

    Science.gov (United States)

    Wu, Zhiyong; Dai, Feifeng; Ren, Wei; Liu, Huagang; Li, Bowen; Chang, Jinxing

    2016-01-01

    Patients with acute aortic dissection (AAD) usually showed acute lung injury (ALI). However, its pathogenesis is still not well defined. Apoptosis of pulmonary microvascular endothelial cells (PMVECs) is closely related to the alveolus-capillary barrier injury and the increased vascular permeability. In this study, we aim to investigate the human PMVECs (hPMVECs) apoptosis induced by angiotensin II (AngII) and monocyte chemoattractant protein-1 (MCP-1) and their potential interaction in the pathogenesis of AAD complicated with ALI. Fifty-eight newly diagnosed AAD, 12 matched healthy individuals were included. Pulmonary tissues of AAD complicated with lung injury were obtained from 2 cadavers to determine the levels of AngII type 1 receptor (AT1-R) and MCP-1. Serum AngII was measured using commercial ELISA kit. H&E staining and immunohistostaining were performed to determine the expression of AT1-R and MCP-1. For the in vitro experiment, hPMVECs were divided into control, AngII group, AngII+Bindarit group and Bindarit group, respectively. Flow cytometry was performed to analyze the apoptosis in each group. Reverse transcription-polymerase chain reaction was performed to determine the mRNA expression of MCP-1. Western blot analysis was performed to evaluate the expression of MCP-1 and apoptosis related protein. Apoptosis of hPMVECs was observed in the lung tissues in the cadavers with AAD complicated with ALI. Besides, the expression of AT1-R and MCP-1 was remarkably elevated. Compared with normal individuals and the non-lung injury AAD patients, the expression of serum AngII was remarkably elevated in AAD patients complicated with ALI. In vitro experiments showed AngII contributed to the apoptosis and elevation of MCP1 in hPMVECs. Besides, it involved in the down-regulation of Bcl-2 protein, and up-regulation of Bax and Caspase-3. Such phenomenon was completely reversed after administration of MCP-1 inhibitor (Bindarit). The production of MCP-1 and cellular

  17. The Diagnostic Value of Superb Microvascular Imaging (SMI) in Detecting Blood Flow Signals of Breast Lesions: A Preliminary Study Comparing SMI to Color Doppler Flow Imaging.

    Science.gov (United States)

    Ma, Yan; Li, Gang; Li, Jing; Ren, Wei-dong

    2015-09-01

    The correlation between color Doppler flow imaging (CDFI) and Superb Microvascular Imaging (SMI) for detecting blood flow in breast lesions was investigated, as was the diagnostic value of SMI in differentiating benign from malignant breast lesions.These lesions were evaluated using both CDFI and SMI according to Adler's method. Pathologic examination showed 57 malignant lesions and 66 benign lesions. The number of blood vessels in a single mass was detected by 2 techniques (SMI and CDFI), and the difference between the 2 values (SMI-CDFI) was calculated. The optimal threshold for the diagnosis of malignant neoplasms and the diagnostic performances of SMI, CDFI, and SMI-CDFI were calculated.For the total lesions and malignant lesions alone, the difference between SMI and CDFI for detecting blood flow was significant (P < 0.01), but the difference was not significant for benign lesions (P = 0.15). The area under the receiver operating characteristic curve was 0.73 (95% confidence interval [CI]: 0.64-0.82) for CDFI; 0.81 (95% CI: 0.74-0.89) for SMI; and 0.89 (95% CI: 0.82-0.95) for SMI-CDFI. Furthermore, the modality of "SMI-CDFI" showed the best diagnostic performance.SMI provides further microvessel information in breast lesions. The diagnostic modality of "SMI-CDFI" can improve the diagnostic performance of ultrasound in the differentiation between benign and malignant masses.

  18. Role of vascular endothelial growth factor in the stimulation of cellular invasion and signaling of breast cancer cells.

    Science.gov (United States)

    Price, D J; Miralem, T; Jiang, S; Steinberg, R; Avraham, H

    2001-03-01

    The expression of vascular endothelial growth factor (VEGF) by breast tumors has been previously correlated with a poor prognosis in the pathogenesis of breast cancer. Furthermore, VEGF secretion is a prerequisite for tumor development. Although most of the effects of VEGF have been shown to be attributable to the stimulation of endothelial cells, we present evidence here that breast tumor cells are capable of responding to VEGF. We show that VEGF stimulation of T-47D breast cancer cells leads to changes in cellular signaling and invasion. VEGF increases the cellular invasion of T-47D breast cancer cells on Matrigel/ fibronectin-coated transwell membranes by a factor of two. Northern analysis for the expression of the known VEGF receptors shows the presence of moderate levels of Flt-1 and low levels of Flk-1/KDR mRNAs in a variety of breast cancer cell lines. T-47D breast cancer cells bind 125I-labeled VEGF with a Kd of 13 x 10(-9) M. VEGF induces the activation of the extracellular regulated kinases 1,2 as well as activation of phosphatidylinositol 3'-kinase, Akt, and Forkhead receptor L1. These findings in T-47D breast cancer cells strongly suggest an autocrine role for VEGF contributing to the tumorigenic phenotype.

  19. 改良的人视网膜血管内皮细胞的培养与鉴定方法%A novel method for culture and identification of primary human retinal microvascular endothelial cells

    Institute of Scientific and Technical Information of China (English)

    毛羽翔; 林少芬; 曾美珍; 田景毅; 唐仕波

    2013-01-01

    Background To optimize the culture method of human retinal microvascular endothelial cells is very important for the study of retinal angiogenesis disease.Human retinal microvascular endothelial cells have been successfully cultured in previous studies,but further improvement of the culture method to harvest higher yields and purity cells is still needed.Objective This study was to design a modified method to isolate and purify human retinal microvascular endothelial cells much easily and quickly,and to compare the expression of specific markers of vascular endothelial cells,factor Ⅷ and CD31/CD34 in the cells.Methods The use of human donor eyeballs was approved by the Ethic Commission of Zhongshan Ophthalmic Center of Sun Yat-sen University.The retina tissue from healthy donor was isolated and digested by the two-step digestion method with 2% trypsin and 0.133% collagenase Ⅳ.Human retinal microvascular endothelial cells were collected and plated in 60 mm dishes coated by 0.1% fibronectin and cultured in endothelial cell-specialized medium supplemented with 10% fetal bovine serum,0.3 mg/L β-endothelial cell growth factor (ECGF) and 100 ng/L sodium heparin.During the culturing,the growth situation of the cells was monitored by morphological observation,and immunohistochemical staining was performed to probe vascular endothelial cell-specific membrane protein CD31,CD34 and factor Ⅷ for identification of the cell purity.Results Human retinal microvascular endothelial cells were isolated successfully from the retina by the twostep digestion method.The primary cultured cells adhered to well 72 hours later and achieved confluence with the typical cobblestone appearance 9 to 10 days after cultured.The cells exhibited the blue nuclei and reddish cytoplasm by regular haematoxylin and eosin stain and showed a strong positive response for CD31,CD34 and factor Ⅷ by immunohistochemistry.The positive dye of CD31 and CD34 was lower than Ⅷ factor in both

  20. Endothelial protein C receptor function in murine and human breast cancer development.

    Directory of Open Access Journals (Sweden)

    Florence Schaffner

    Full Text Available Several markers identify cancer stem cell-like populations, but little is known about the functional roles of stem cell surface receptors in tumor progression. Here, we show that the endothelial protein C receptor (EPCR, a stem cell marker in hematopoietic, neuronal and epithelial cells, is crucial for breast cancer growth in the orthotopic microenvironment of the mammary gland. Mice with a hypomorphic allele of EPCR show reduced tumor growth in the PyMT-model of spontaneous breast cancer development and deletion of EPCR in established PyMT tumor cells significantly attenuates transplanted tumor take and growth. We find expansion of EPCR(+ cancer stem cell-like populations in aggressive, mammary fat pad-enhanced human triple negative breast cancer cells. In this model, EPCR-expressing cells have markedly increased mammosphere- and tumor-cell initiating activity compared to another stable progenitor-like subpopulation present at comparable frequency. We show that receptor blocking antibodies to EPCR specifically attenuate in vivo tumor growth initiated by either EPCR(+ cells or the heterogenous mixture of EPCR(+ and EPCR(- cells. Furthermore, we have identified tumor associated macrophages as a major source for recognized ligands of EPCR, suggesting a novel mechanism by which cancer stem cell-like populations are regulated by innate immune cells in the tumor microenvironment.

  1. Fumaric Acid Esters Do Not Reduce Inflammatory NF-κB/p65 Nuclear Translocation, ICAM-1 Expression and T-Cell Adhesiveness of Human Brain Microvascular Endothelial Cells.

    Science.gov (United States)

    Haarmann, Axel; Nehen, Mathias; Deiß, Annika; Buttmann, Mathias

    2015-08-13

    Dimethyl fumarate (DMF) is approved for disease-modifying treatment of patients with relapsing-remitting multiple sclerosis. Animal experiments suggested that part of its therapeutic effect is due to a reduction of T-cell infiltration of the central nervous system (CNS) by uncertain mechanisms. Here we evaluated whether DMF and its primary metabolite monomethyl fumarate (MMF) modulate pro-inflammatory intracellular signaling and T-cell adhesiveness of nonimmortalized single donor human brain microvascular endothelial cells at low passages. Neither DMF nor MMF at concentrations of 10 or 50 µM blocked the IL-1β-induced nuclear translocation of NF-κB/p65, whereas the higher concentration of DMF inhibited the nuclear entry of p65 in human umbilical vein endothelium cultured in parallel. DMF and MMF also did not alter the IL-1β-stimulated activation of p38 MAPK in brain endothelium. Furthermore, neither DMF nor MMF reduced the basal or IL-1β-inducible expression of ICAM-1. In accordance, both fumaric acid esters did not reduce the adhesion of activated Jurkat T cells to brain endothelium under basal or inflammatory conditions. Therefore, brain endothelial cells probably do not directly mediate a potential blocking effect of fumaric acid esters on the inflammatory infiltration of the CNS by T cells.

  2. Salvianolic acid B improves the disruption of high glucose-mediated brain microvascular endothelial cells via the ROS/HIF-1α/VEGF and miR-200b/VEGF signaling pathways.

    Science.gov (United States)

    Yang, Ming-Chao; You, Fu-Li; Wang, Zhe; Liu, Xiang-Nan; Wang, Yan-Feng

    2016-09-06

    The study investigated the roles and mechanisms of Salvianolic acid B (Sal B) on permeability of rat brain microvascular endothelial cells (RBMECs) exposed to high glucose. The results demonstrated that Sal B greatly up-regulated the expression of tight junction (TJ) proteins and decreased the permeability of RBMECs compared with the control group. And the increase of reactive oxidative species (ROS) production, the upregulation of hypoxia-inducible factor-1 alpha (HIF-1α) and vascular endothelial growth factor (VEGF) protein induced by high glucose were antagonized by Sal B. In addition, a great decrease of microRNA-200b (miR-200b) was observed in the RBMECs under high-glucose condition, which was significantly increased by Sal B pretreatment. And overexpression of miR-200b markedly attenuated the RBMECs permeability and inhibited the expression of VEGF protein by targeting with 3'-UTR of its mRNA. This led to the conclusion that Sal B-mediated improvement of blood-brain barrier dysfunction induced by high-glucose is related to the ROS/HIF-1α/VEGF and miR-200b/VEGF signaling pathways.

  3. Fumaric Acid Esters Do Not Reduce Inflammatory NF-κB/p65 Nuclear Translocation, ICAM-1 Expression and T-Cell Adhesiveness of Human Brain Microvascular Endothelial Cells

    Directory of Open Access Journals (Sweden)

    Axel Haarmann

    2015-08-01

    Full Text Available Dimethyl fumarate (DMF is approved for disease-modifying treatment of patients with relapsing-remitting multiple sclerosis. Animal experiments suggested that part of its therapeutic effect is due to a reduction of T-cell infiltration of the central nervous system (CNS by uncertain mechanisms. Here we evaluated whether DMF and its primary metabolite monomethyl fumarate (MMF modulate pro-inflammatory intracellular signaling and T-cell adhesiveness of nonimmortalized single donor human brain microvascular endothelial cells at low passages. Neither DMF nor MMF at concentrations of 10 or 50 µM blocked the IL-1β-induced nuclear translocation of NF-κB/p65, whereas the higher concentration of DMF inhibited the nuclear entry of p65 in human umbilical vein endothelium cultured in parallel. DMF and MMF also did not alter the IL-1β-stimulated activation of p38 MAPK in brain endothelium. Furthermore, neither DMF nor MMF reduced the basal or IL-1β-inducible expression of ICAM-1. In accordance, both fumaric acid esters did not reduce the adhesion of activated Jurkat T cells to brain endothelium under basal or inflammatory conditions. Therefore, brain endothelial cells probably do not directly mediate a potential blocking effect of fumaric acid esters on the inflammatory infiltration of the CNS by T cells.

  4. Endothelial cells provide a notch-dependent pro-tumoral niche for enhancing breast cancer survival, stemness and pro-metastatic properties.

    Directory of Open Access Journals (Sweden)

    Pegah Ghiabi

    Full Text Available Treating metastasis has been challenging due to tumors complexity and heterogeneity. This complexity is partly related to the crosstalk between tumor and its microenvironment. Endothelial cells -the building blocks of tumor vasculature- have been shown to have additional roles in cancer progression than angiogenesis and supplying oxygen and nutrients. Here, we show an alternative role for endothelial cells in supporting breast cancer growth and spreading independent of their vascular functions. Using endothelial cells and breast cancer cell lines MDA-MB231 and MCF-7, we developed co-culture systems to study the influence of tumor endothelium on breast tumor development by both in vitro and in vivo approaches. Our results demonstrated that endothelial cells conferred survival advantage to tumor cells under complete starvation and enriched the CD44HighCD24Low/- stem cell population in tumor cells. Moreover, endothelial cells enhanced the pro-metastatic potential of breast cancer cells. The in vitro and in vivo results concordantly confirmed a role for endothelial Jagged1 to promote breast tumor through notch activation. Here, we propose a role for endothelial cells in enhancing breast cancer progression, stemness, and pro-metastatic traits through a perfusion-independent manner. Our findings may be beneficial in developing novel therapeutic approaches.

  5. Lymphatic Reprogramming of Adult Endothelial Stem Cells for a Cell-Based Therapy for Lymphedema in Breast Cancer Patients

    Science.gov (United States)

    2008-09-01

    Therapy for Lymphedema inBreast Cancer Patients PRINCIPAL INVESTIGATOR: Young Kwon Hong, Ph.D. CONTRACTING ORGANIZATION...5a. CONTRACT NUMBER 4. TITLE AND SUBTITLE Lymphatic Reprogramming of Adult Endothelial Stem Cells for a Cell-Based Therapy for Lymphedema in... lymphedema patients. The key significance of our proposal is to utilize the elusive circulating adult stem cells to avoid the ethical and immunological

  6. Reprint of "The role of cytoskeleton in the regulation of vascular endothelial barrier function" [Microvascular Research 76 (2008) 202-207].

    Science.gov (United States)

    Bogatcheva, Natalia V; Verin, Alexander D

    2009-01-01

    The cytoskeleton is vital to the function of virtually all cell types in the organism as it is required for cell division, cell motility, endo- or exocytosis and the maintenance of cell shape. Endothelial cells, lining the inner surface of the blood vessels, exploit cytoskeletal elements to ensure the integrity of cell monolayer in quiescent endothelium, and to enable the disintegration of the formed barrier in response to various agonists. Vascular permeability is defined by the combination of transcellular and paracellular pathways, with the latter being a major contributor to the inflammation-induced barrier dysfunction. This review will analyze the cytoskeletal elements, which reorganization affects endothelial permeability, and emphasize signaling mechanisms with barrier-protective or barrier-disruptive potential.

  7. Distribution of a 69-kD laminin-binding protein in aortic and microvascular endothelial cells: modulation during cell attachment, spreading, and migration

    DEFF Research Database (Denmark)

    Yannariello-Brown, J; Wewer, U; Liotta, L;

    1988-01-01

    Affinity chromatography and immunolocalization techniques were used to investigate the mechanism(s) by which endothelial cells interact with the basement membrane component laminin. Bovine aortic endothelial cells (BAEC) membranes were solubilized and incubated with a laminin-Sepharose affinity...... column. SDS-PAGE analysis of the eluted proteins identified a 69-kD band as the major binding protein, along with minor components migrating at 125, 110, 92, 85, 75, 55, and 30 kD. Polyclonal antibodies directed against a peptide sequence of the 69-kD laminin-binding protein isolated from human tumor......, with a granular perinuclear distribution and in linear arrays throughout the cell. During migration a redistribution from diffuse to predominanately linear arrays that co-distributed with actin microfilaments was noted in double-label experiments. The 69-kD laminin-binding protein colocalized with actin filaments...

  8. Impaired systemic tetrahydrobiopterin bioavailability and increased dihydrobiopterin in adult falciparum malaria: association with disease severity, impaired microvascular function and increased endothelial activation.

    Directory of Open Access Journals (Sweden)

    Tsin W Yeo

    2015-03-01

    Full Text Available Tetrahydrobiopterin (BH₄ is a co-factor required for catalytic activity of nitric oxide synthase (NOS and amino acid-monooxygenases, including phenylalanine hydroxylase. BH4 is unstable: during oxidative stress it is non-enzymatically oxidized to dihydrobiopterin (BH₂, which inhibits NOS. Depending on BH₄ availability, NOS oscillates between NO synthase and NADPH oxidase: as the BH₄/BH₂ ratio decreases, NO production falls and is replaced by superoxide. In African children and Asian adults with severe malaria, NO bioavailability decreases and plasma phenylalanine increases, together suggesting possible BH₄ deficiency. The primary three biopterin metabolites (BH₄, BH₂ and B₀ [biopterin] and their association with disease severity have not been assessed in falciparum malaria. We measured pterin metabolites in urine of adults with severe falciparum malaria (SM; n=12, moderately-severe malaria (MSM, n=17, severe sepsis (SS; n=5 and healthy subjects (HC; n=20 as controls. In SM, urinary BH₄ was decreased (median 0.16 ¼mol/mmol creatinine compared to MSM (median 0.27, SS (median 0.54, and HC (median 0.34]; p<0.001. Conversely, BH₂ was increased in SM (median 0.91 ¼mol/mmol creatinine, compared to MSM (median 0.67, SS (median 0.39, and HC (median 0.52; p<0.001, suggesting increased oxidative stress and insufficient recycling of BH2 back to BH4 in severe malaria. Overall, the median BH₄/BH₂ ratio was lowest in SM [0.18 (IQR: 0.04-0.32] compared to MSM (0.45, IQR 0.27-61, SS (1.03; IQR 0.54-2.38 and controls (0.66; IQR 0.43-1.07; p<0.001. In malaria, a lower BH₄/BH₂ ratio correlated with decreased microvascular reactivity (r=0.41; p=0.03 and increased ICAM-1 (r=-0.52; p=0.005. Decreased BH4 and increased BH₂ in severe malaria (but not in severe sepsis uncouples NOS, leading to impaired NO bioavailability and potentially increased oxidative stress. Adjunctive therapy to regenerate BH4 may have a role in improving NO

  9. Xiang-Qi-Tang and its active components exhibit anti-inflammatory and anticoagulant properties by inhibiting MAPK and NF-κB signaling pathways in LPS-treated rat cardiac microvascular endothelial cells.

    Science.gov (United States)

    He, Chang-Liang; Yi, Peng-Fei; Fan, Qiao-Jia; Shen, Hai-Qing; Jiang, Xiao-Lin; Qin, Qian-Qian; Song, Zhou; Zhang, Cui; Wu, Shuai-Cheng; Wei, Xu-Bin; Li, Ying-Lun; Fu, Ben-Dong

    2013-04-01

    Xiang-Qi-Tang (XQT) is a Chinese herbal formula containing Cyperus rotundus, Astragalus membranaceus and Andrographis paniculata. Alpha-Cyperone (CYP), astragaloside IV (AS-IV) and andrographolide (AND) are the three major active components in this formula. XQT may modulate the inflammatory or coagulant responses. We therefore assessed the effects of XQT on lipopolysaccharide (LPS)-induced inflammatory model of rat cardiac microvascular endothelial cells (RCMECs). XQT, CYP, AS-IV and AND inhibited the production of tumor necrosis factor alpha (TNF-α), intercellular cell adhesion molecule-1 (ICAM-1) and plasminogen activator inhibitor-1 (PAI-1), and up-regulated the mRNA expression of Kruppel-like factor 2 (KLF2). XQT and CYP inhibited the secretion of tissue factor (TF). To further explore the mechanism, we found that XQT, or its active components CYP, AS-IV and AND significantly inhibited extracellular signal-regulated kinase (ERK), c-jun NH2-terminal kinase (JNK) and p38 phosphorylation protein expression as well as decreased the phosphorylation levels of nuclear factor κB (NF-κB) p65 proteins in LPS-stimulated RCMECs. These results suggested that XQT and its active components inhibited the expression of inflammatory and coagulant mediators via mitogen-activated protein kinase (MAPKs) and NF-κB signaling pathways. These findings may contribute to future research on the action mechanisms of this formula, as well as therapy for inflammation- or coagulation-related diseases.

  10. Aryl hydrocarbon receptor regulates CYP1B1 but not ABCB1 and ABCG2 in hCMEC/D3 human cerebral microvascular endothelial cells after TCDD exposure.

    Science.gov (United States)

    Jacob, Aude; Potin, Sophie; Chapy, Hélène; Crete, Dominique; Glacial, Fabienne; Ganeshamoorthy, Kayathiri; Couraud, Pierre-Olivier; Scherrmann, Jean-Michel; Declèves, Xavier

    2015-07-10

    The aryl hydrocarbon receptor (AhR) is a ligand-dependent transcription factor activated by a variety of widespread persistent environmental pollutants such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). It can transactivate the expression of several target genes. Recently AhR transcripts were detected in isolated human brain microvessels and in the hCMEC/D3 human cerebral microvascular endothelial cell line, an in vitro model of the human cerebral endothelium. To date AhR implication in the co-regulation of ABCB1, ABCG2 and CYP1B1 at human cerebral endothelium has not been addressed. Here we investigated whether AhR could co-regulate ABCB1, ABCG2 and CYP1B1 expressions in the hCMEC/D3 cell line. Exposure to TCDD induced a concentration-dependent increase in CYP1B1 expression. We demonstrated AhR involvement in the TCDD-mediated increase in CYP1B1 expression by using small interfering RNA against AhR. Western blotting analysis also revealed an increase in CYP1B1 protein expression following TCDD exposure in hCMEC/D3. Regarding ABCB1 and ABCG2, exposure to TCDD had no effect on their protein expressions and functional activities. In conclusion our data indicated a differential modulation of CYP1B1 and ABCB1/ABCG2 expressions in hCMEC/D3 cells following TCDD exposure.

  11. Protein kinase C-α signals P115RhoGEF phosphorylation and RhoA activation in TNF-α-induced mouse brain microvascular endothelial cell barrier dysfunction

    Directory of Open Access Journals (Sweden)

    Deng Xiaolu

    2011-04-01

    Full Text Available Abstract Background Tumor necrosis factor-α (TNF-α, a proinflammatory cytokine, is capable of activating the small GTPase RhoA, which in turn contributes to endothelial barrier dysfunction. However, the underlying signaling mechanisms remained undefined. Therefore, we aimed to determine the role of protein kinase C (PKC isozymes in the mechanism of RhoA activation and in signaling TNF-α-induced mouse brain microvascular endothelial cell (BMEC barrier dysfunction. Methods Bend.3 cells, an immortalized mouse brain endothelial cell line, were exposed to TNF-α (10 ng/mL. RhoA activity was assessed by pull down assay. PKC-α activity was measured using enzyme assasy. BMEC barrier function was measured by transendothelial electrical resistance (TER. p115RhoGEF phosphorylation was detected by autoradiography followed by western blotting. F-actin organization was observed by rhodamine-phalloidin staining. Both pharmacological inhibitors and knockdown approaches were employed to investigate the role of PKC and p115RhoGEF in TNF-α-induced RhoA activation and BMEC permeability. Results We observed that TNF-α induces a rapid phosphorylation of p115RhoGEF, activation of PKC and RhoA in BMECs. Inhibition of conventional PKC by Gö6976 mitigated the TNF-α-induced p115RhoGEF phosphorylation and RhoA activation. Subsequently, we found that these events are regulated by PKC-α rather than PKC-β by using shRNA. In addition, P115-shRNA and n19RhoA (dominant negative mutant of RhoA transfections had no effect on mediating TNF-α-induced PKC-α activation. These data suggest that PKC-α but not PKC-β acts as an upstream regulator of p115RhoGEF phosphorylation and RhoA activation in response to TNF-α. Moreover, depletion of PKC-α, of p115RhoGEF, and inhibition of RhoA activation also prevented TNF-α-induced stress fiber formation and a decrease in TER. Conclusions Taken together, our results show that PKC-α phosphorylation of p115RhoGEF mediates TNF

  12. Neisseria meningitidis causes cell cycle arrest of human brain microvascular endothelial cells at S phase via p21 and cyclin G2.

    Science.gov (United States)

    Oosthuysen, Wilhelm F; Mueller, Tobias; Dittrich, Marcus T; Schubert-Unkmeir, Alexandra

    2016-01-01

    Microbial pathogens have developed several mechanisms to modulate and interfere with host cell cycle progression. In this study, we analysed the effect of the human pathogen Neisseria meningitidis on cell cycle in a brain endothelial cell line as well as in primary brain endothelial cells. We found that N.  Meningitidis causes an accumulation of cells in the S phase early at 3 and at 24 h post-infection that was paralleled by a decrease of cells in G2/M phase. Importantly, the outer membrane proteins of the colony opacity-associated (Opa) protein family as well as the Opc protein proved to trigger the accumulation of cells in the S phase. A focused cell cycle reverse transcription quantitative polymerase chain reaction-based array and integrated network analysis revealed changes in the abundance of several cell cycle regulatory mRNAs, including the cell cycle inhibitors p21(WAF1/CIP1) and cyclin G2. These alterations were reflected in changes in protein expression levels and/or relocalization in N. meningitidis-infected cells. Moreover, an increase in p21(WAF1/CIP1) expression was found to be p53 independent. Genetic ablation of p21(WAF1/CIP1) and cyclin G2 abrogated N. meningitidis-induced S phase accumulation. Finally, by measuring the levels of the biomarker 8-hydroxydeoxyguanosine and phosphorylation of the histone variant H2AX, we provide evidence that N. meningitidis induces oxidative DNA damage in infected cells.

  13. Matrix metalloproteinase 2 (MMP-2) degrades soluble vasculotropic amyloid-beta E22Q and L34V mutants, delaying their toxicity for human brain microvascular endothelial cells.

    Science.gov (United States)

    Hernandez-Guillamon, Mar; Mawhirt, Stephanie; Fossati, Silvia; Blais, Steven; Pares, Mireia; Penalba, Anna; Boada, Merce; Couraud, Pierre-Olivier; Neubert, Thomas A; Montaner, Joan; Ghiso, Jorge; Rostagno, Agueda

    2010-08-27

    Patients carrying mutations within the amyloid-beta (Abeta) sequence develop severe early-onset cerebral amyloid angiopathy with some of the related variants manifesting primarily with hemorrhagic phenotypes. Matrix metalloproteases (MMPs) are typically associated with blood brain barrier disruption and hemorrhagic transformations after ischemic stroke. However, their contribution to cerebral amyloid angiopathy-related hemorrhage remains unclear. Human brain endothelial cells challenged with Abeta synthetic homologues containing mutations known to be associated in vivo with hemorrhagic manifestations (AbetaE22Q and AbetaL34V) showed enhanced production and activation of MMP-2, evaluated via Multiplex MMP antibody arrays, gel zymography, and Western blot, which in turn proteolytically cleaved in situ the Abeta peptides. Immunoprecipitation followed by mass spectrometry analysis highlighted the generation of specific C-terminal proteolytic fragments, in particular the accumulation of Abeta-(1-16), a result validated in vitro with recombinant MMP-2 and quantitatively evaluated using deuterium-labeled internal standards. Silencing MMP-2 gene expression resulted in reduced Abeta degradation and enhanced apoptosis. Secretion and activation of MMP-2 as well as susceptibility of the Abeta peptides to MMP-2 degradation were dependent on the peptide conformation, with fibrillar elements of AbetaE22Q exhibiting negligible effects. Our results indicate that MMP-2 release and activation differentially degrades Abeta species, delaying their toxicity for endothelial cells. However, taking into consideration MMP ability to degrade basement membrane components, these protective effects might also undesirably compromise blood brain barrier integrity and precipitate a hemorrhagic phenotype.

  14. Microvascular pericytes in healthy and diseased kidneys

    Directory of Open Access Journals (Sweden)

    Pan SY

    2014-01-01

    Full Text Available Szu-Yu Pan,1,2 Yu-Ting Chang,3 Shuei-Liong Lin1,31Renal Division, Department of Internal Medicine, National Taiwan University Hospital, Taipei, Taiwan; 2Department of Internal Medicine, National Taiwan University Hospital, Yun-Lin Branch, Yun-Lin, Taiwan; 3Graduate Institute of Physiology, College of Medicine, National Taiwan University, Taipei, TaiwanAbstract: Pericytes are interstitial mesenchymal cells found in many major organs. In the kidney, microvascular pericytes are defined anatomically as extensively branched, collagen-producing cells in close contact with endothelial cells. Although many molecular markers have been proposed, none of them can identify the pericytes with satisfactory specificity or sensitivity. The roles of microvascular pericytes in kidneys were poorly understood in the past. Recently, by using genetic lineage tracing to label collagen-producing cells or mesenchymal cells, the elusive characteristics of the pericytes have been illuminated. The purpose of this article is to review recent advances in the understanding of microvascular pericytes in the kidneys. In healthy kidney, the pericytes are found to take part in the maintenance of microvascular stability. Detachment of the pericytes from the microvasculature and loss of the close contact with endothelial cells have been observed during renal insult. Renal microvascular pericytes have been shown to be the major source of scar-forming myofibroblasts in fibrogenic kidney disease. Targeting the crosstalk between pericytes and neighboring endothelial cells or tubular epithelial cells may inhibit the pericyte-myofibroblast transition, prevent peritubular capillary rarefaction, and attenuate renal fibrosis. In addition, renal pericytes deserve attention for their potential to produce erythropoietin in healthy kidneys as pericytes stand in the front line, sensing the change of oxygenation and hemoglobin concentration. Further delineation of the mechanisms underlying the

  15. GENISTEIN INHIBITS EXPRESSION OF VASCULAR ENDOTHELIAL GROWTH FACTOR IN HER-2/NEU TRANSFECTED HUMAN BREAST CANCER MCF-7 CELLS

    Institute of Scientific and Technical Information of China (English)

    ZHU Jun-dong; YU Xiao-ping; MI Man-tian

    2006-01-01

    Objective: our previous studies have demonstrated that HER-2/neu gene expression in human breast cancer MCF-7 cells promotes angiogenesis in MCF-7 cells xenograft tumors, and genistein inhibits angiogenesis in MCF-7 cells with HER-2/neu expression xenograft tumors. Here, the effects of genistein on the expression of vascular endothelial growth factor (VEGF) inMCR-7 cells with HER-2/neu expression were further studied for exploring the molecular mechanism of anti-angiogenesis in HER-2/neu-overexpressing breast cancer by genistein. Methods: HER-2/neu-overexpressing MCF-7 cells (MCF-7/HER-2)were established by transfecting HER-2/neu gene into HER-2/neu negative expression breast cancer MCF-7 cells.Immunocytochemical staining, western blot and reverse transcription-polymerase chain reaction (RT-PCR) were adopted to measure the expression of VEGF in MCF-7/HER-2 cells treated by genistein for 24, 48 and 72h. Results: HER-2/neu expression up-regulated VEGF mRNA and protein in MCF-7 cells, genistein decreased VEGF mRNA and protein level in MCF-7/HER-2 cells in a time-dependent manner. Conclusion: These results suggest that VEGF plays an important role in HER-2/neu gene expression promoted antiogenesis in breast cancer and genistein induced down-regulation of the expression of VEGF may be one of the molecular mechanisms of its anti-angiogenesis in HER-2/neu-overexpressing breast cancer.

  16. Anti-angiogenesis effect of generation 4 polyamidoamine/vascular endothelial growth factor antisense oligodeoxynucleotide on breast cancer in vitro

    Institute of Scientific and Technical Information of China (English)

    Shan-zhi GU; Xin-han ZHAO; Ling-xiao ZHANG; Li LI; Zhi-yu WANG; Min MENG; Gai-li AN

    2009-01-01

    Objective: To study the effects of the generation 4 polyamidoamine/vascular endothelial growth factor antisense oligodeoxynucleotide (G4PAMAMNEGFASODN) compound on the expressions of vascular endothelial growth factor (VEGF) and its mRNA of breast cancer cells and on the inhibition of vascular endothelial cells. Methods: We examined the morphology of G4PAMAM/VEGFASODN compound and its pH stability, in vitro transfection efficiency and toxicity, and the expressions of VEGF and its mRNA. Methyl thiazolyl tetrazolium assay was used to detect the inhibitory function of the compound on vascular endothelial cells. Results: The compound was about 10 nm in diameter and was homogeneously netlike. From pH 5 to 10, it showed quite a buffered ability. The 48-h transfection rate in the charge ratio of 1:40 was 98.76%, significantly higher than that of the liposome group (P<0.05). None of the transfection products showed obvious toxicity on the cells. The expressions of both VEGF protein and its mRNA after G4PAMAM/VEGFASODN transfection decreased markedly. Conclusion: With a low toxicity, high safety, and high transfection rate, G4PAMAMNEGFASODN could be a promising gene vector. Specifically, it inhibits VEGF gene expression efficiently, laying a basis for further in vivo animal studies.

  17. Culture and identification of microvascular endothelial cells from human endometriosis%卵巢子宫内膜异位症病灶微血管内皮细胞的培养与鉴定

    Institute of Scientific and Technical Information of China (English)

    欧阳卓; 卢丹; 张为远

    2011-01-01

    目的 建立卵巢子宫内膜异位症(内异症)病灶微血管内皮细胞( OEMEC)的体外培养体系并进行鉴定.方法 采用Ⅰ型和Ⅱ型胶原酶、胰蛋白酶一乙二胺四乙酸(EDTA)联合消化法,分离卵巢内异症患者术后病理检查确诊的囊肿组织中的OEMEC,经差速贴壁法和杂细胞擦刮法纯化;采用免疫细胞化学染色法染色,通过光镜、透射电镜观察微血管内皮细胞特异性标志物——CD34、第八因子多克隆抗体(FⅧ-Rag)和微血管内皮细胞特异性结构——Weibel-Palade小体的表达,对所获得的OEMEC进行鉴定.结果 光镜下观察OEMEC呈现典型的铺路石样生长;免疫细胞化学染色显示有CD34和FⅧ-Rag的强表达,其阳性细胞的百分率分别为91.4%和92.5%;透射电镜下观察OEMEC内可见Weibel-Palade小体的存在;培养的子代微血管内皮细胞生长良好,细胞群体倍增时间为4.5d.结论 OEMEC体外培养体系的成功建立,对了解人OEMEC的异质性具有较好的实用价值,将为内异症病灶中新生微血管特点的研究奠定细胞学实验基础.%Objective To establish the methods of isolating and culturing human ovarian endometriosis-derived microvascular endothelial cells (OEMEC).Methods The tissues of human endometriotic cyst of ovary were finely minced with scissors,then digested by collagenase Ⅰ,Ⅱ and trypsinethylene diamine tetraacetic acid (EDTA).The cells were purified by using centrifugation of 2000 r/min speed.OEMEC were identified by light microscope and transmission electron microscope observing CD34,FⅧ-Rag and Weibel-Palade in microvascular endothelial cells.Results The OEMEC grew as confluent monolayer like cobblestones under light microscope.CD34 and FⅧ-Rag were expressed strongly,and the percentages of CD34 and FⅧ-Rag positive cells were 91.4% and 92.5%.Weibel-Palade bodies could be observed under transmission electron microscope.The time of cell doubling proliferation was

  18. Human embryonic stem cell-derived endothelial cells as cellular delivery vehicles for treatment of metastatic breast cancer.

    Science.gov (United States)

    Su, Weijun; Wang, Lina; Zhou, Manqian; Liu, Ze; Hu, Shijun; Tong, Lingling; Liu, Yanhua; Fan, Yan; Kong, Deling; Zheng, Yizhou; Han, Zhongchao; Wu, Joseph C; Xiang, Rong; Li, Zongjin

    2013-01-01

    Endothelial progenitor cells (EPCs) have shown tropism towards primary tumors or metastases and are thus potential vehicles for targeting tumor therapy. However, the source of adult EPCs is limited, which highlights the need for a consistent and renewable source of endothelial cells for clinical applications. Here, we investigated the potential of human embryonic stem cell-derived endothelial cells (hESC-ECs) as cellular delivery vehicles for therapy of metastatic breast cancer. In order to provide an initial assessment of the therapeutic potency of hESC-ECs, we treated human breast cancer MDA-MB-231 cells with hESC-EC conditioned medium (EC-CM) in vitro. The results showed that hESC-ECs could suppress the Wnt/β-catenin signaling pathway and thereby inhibit the proliferation and migration of MDA-MB-231 cells. To track and evaluate the possibility of hESC-EC-employed therapy, we employed the bioluminescence imaging (BLI) technology. To study the therapeutic potential of hESC-ECs, we established lung metastasis models by intravenous injection of MDA-MB-231 cells labeled with firefly luciferase (Fluc) and green fluorescent protein (GFP) to NOD/SCID mice. In mice with lung metastases, we injected hESC-ECs armed with herpes simplex virus truncated thymidine kinase (HSV-ttk) intravenously on days 11, 16, 21, and 26 after MDA-MB-231 cell injection. The NOD/SCID mice were subsequently treated with ganciclovir (GCV), and the growth status of tumor was monitored by Fluc imaging. We found that MDA-MB-231 tumors were significantly inhibited by intravenously injected hESC-ECs. The tumor-suppressive effects of the hESC-ECs, by inhibiting Wnt/β-catenin signaling pathway and inducing tumor cell death through bystander effect in human metastatic breast cancer model, provide previously unexplored therapeutic modalities for cancer treatment.

  19. Comprehensive characterization of rat pulmonary microvascular endothelial cells cultured by tissue-sticking method%组织块法培养大鼠肺微血管内皮细胞的综合鉴定

    Institute of Scientific and Technical Information of China (English)

    徐顺贵; 吴国明; 徐智; 冯起甲; 王关嵩; 张健鹏

    2007-01-01

    目的 为组织块法培养的大鼠肺微血管内皮细胞(rat pulmonary microvascular endothelial cells,RPMVECs)建立合理可靠的多指标综合鉴定方案.方法 采用外周肺组织贴块法培养RPMVECs,抽取组织块进行切片观察,以大鼠肺动脉平滑肌细胞和人脐静脉内皮细胞为对照,对培养细胞进行CD34、植物凝集素BSI、Ⅷ因子相关抗原免疫细胞化学鉴定,通过光镜和透射电镜观察细胞形态和超微结构.结果 组织切片显示组织块源于外周肺组织,CD34免疫细胞化学染色阳性,植物凝集素BSI结合试验阳性,而Ⅷ因子相关抗原染色阴性,透射电镜未见Weibel-Palade小体.结论 Ⅷ因子相关抗原和Weibel-Palade小体并非RPMVECs鉴定的理想指标,联合应用外周肺组织切片、CD34和植物凝集素BSI三指标为组织块法培养的RPMVECs提供了一个简单易行、合理、可靠的综合鉴定方案.

  20. Unfractionated heparin suppresses lipopolysaccharide-induced monocyte chemoattractant protein-1 expression in human microvascular endothelial cells by blocking Krüppel-like factor 5 and nuclear factor-κB pathway.

    Science.gov (United States)

    Li, Xu; Li, Xin; Zheng, Zhen; Liu, Yina; Ma, Xiaochun

    2014-10-01

    Unfractionated heparin (UFH) and low-molecular-weight heparins (LMWH), apart from anticoagulant activities, contain a variety of biological properties such as anti-inflammatory actions possibly affecting sepsis. Chemokines are vital for promoting the movement of circulating leukocytes to the site of infection and are involved in the pathogenesis of sepsis. The purpose of this study was to investigate the effects and potential mechanisms of UFH on lipopolysaccharide (LPS)-induced chemokine production in human pulmonary microvascular endothelial cells (HPMECs). HPMECs were pretreated with UFH (0.1 U/ml and 1 U/ml), 15 min prior to stimulation with LPS (10 μg/ml). Cells were cultured under various experimental conditions for 2 h and 6 h for analysis. UFH markedly decreased LPS-induced interleukin (IL)-8 and monocyte chemoattractant protein-1 (MCP-1) mRNA and protein expression in HPMECs. UFH also attenuated the secretion of these chemokines in culture supernatants. In addition, UFH blocked the chemotactic activities of LPS-stimulated HPMECs supernatants on monocytes migration as expected. UFH inhibited LPS-induced Krüppel-like factor 5 (KLF-5) mRNA and protein levels. Concurrently, UFH reduced nuclear factor (NF)-κB nuclear translocation. Importantly, transfection with siRNA targeting KLF-5 reduced NF-κB activation and chemokines expression. These results demonstrate that interfering with KLF-5 mediated NF-κB activation might contribute to the inhibitory effects of chemokines and monocytes migration by UFH in LPS-stimulated HPMECs.

  1. Effects of polysaccharides from pulsatilla decoction on glycocalyx sugar chains of microvascular endothelial cells%白头翁汤中总多糖对微血管内皮细胞糖萼糖链表达的影响

    Institute of Scientific and Technical Information of China (English)

    杨重锦; 孙雄; 穆祥; 张涛

    2016-01-01

    [目的]研究白头翁汤中总多糖对微血管内皮细胞(microvascular endothelial cells,MVECs)糖萼糖链的影响.[方法]体外培养猪小肠黏膜MVECs,以50μg/mL白头翁汤中总多糖刺激48 h后,利用凝集素荧光技术检测刀豆凝集素(concanavalin A,Con A)、麦胚凝集素(wheat germ agglutinin,WGA)、双花扁豆凝集素(dolichos bifows agglutinin,DBA)、荆豆凝集素(ulex europaeus agglutinin Ⅰ,UEA Ⅰ)、花生凝集素(peanut agglutinin,PNA)、雪花莲凝集素(galanthus nivalis lectin,GNL)、番茄凝集素(lycopersicon esculentum lectin,LEL)受体糖链的表达.[结果]正常情况下猪小肠黏膜MVECs WGA、LEL、Con A和GNL四种凝集素荧光染色呈强阳性,DBA和PNA荧光染色呈弱阳性,UEA Ⅰ荧光染色呈阴性;白头翁汤中总多糖能显著提高WGA和LEL荧光染色的强度.[结论]白头翁汤中总多糖能上调猪小肠黏膜MVECs表达N-乙酰氨基葡萄糖.

  2. Perfusion parameters of dynamic contrast-enhanced magnetic resonance imaging in patients with rectal cancer: Correlation with microvascular density and vascular endothelial growth factor expression

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Yeo Eun [Dept. of Radiology, Seoul Medical Center, Seoul (Korea, Republic of); Lim, Joon Seok; Kim, Myeong Jin; Kim, Ki Whang; Choi, Jun Jeong [Yonsei University Health System, Seoul (Korea, Republic of); Kim, Dae Hong [Molecular Imaging and Therapy Branch, National Cancer Center, Goyang (Korea, Republic of); Myoung, Sung Min [Dept. of Medical Information, Jungwon University, Goesan (Korea, Republic of)

    2013-12-15

    To determine whether quantitative perfusion parameters of dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) correlate with immunohistochemical markers of angiogenesis in rectal cancer. Preoperative DCE-MRI was performed in 63 patients with rectal adenocarcinoma. Transendothelial volume transfer (K{sup trans}) and fractional volume of the extravascular-extracellular space (Ve) were measured by Interactive Data Language software in rectal cancer. After surgery, microvessel density (MVD) and vascular endothelial growth factor (VEGF) expression scores were determined using immunohistochemical staining of rectal cancer specimens. Perfusion parameters (K{sup trans}, Ve) of DCE-MRI in rectal cancer were found to be correlated with MVD and VEGF expression scores by Spearman's rank coefficient analysis. T stage and N stage (negative or positive) were correlated with perfusion parameters and MVD. Significant correlation was not found between any DCE-MRI perfusion parameters and MVD (rs = -0.056 and p 0.662 for K{sup trans}; rs = -0.103 and p = 0.416 for Ve), or between any DCE-MRI perfusion parameters and the VEGF expression score (rs = -0.042, p 0.741 for K{sup trans}; r = 0.086, p = 0.497 for Ve) in rectal cancer. TN stage showed no significant correlation with perfusion parameters or MVD (p > 0.05 for all). DCE-MRI perfusion parameters, K{sup trans} and Ve, correlated poorly with MVD and VEGF expression scores in rectal cancer, suggesting that these parameters do not simply denote static histological vascular properties.

  3. Vascular endothelial growth inhibitor affects the invasion, apoptosis and vascularisation in breast cancer cell line MDA-MB-231

    Institute of Scientific and Technical Information of China (English)

    Gao Yinguang; Ge Zhicheng; Zhang Zhongtao; Bai Zhigang; Ma Xuemei; Wang Yu

    2014-01-01

    Background Breast cancer is one of the most common malignant female diseases worldwide.It is a significant threat to every woman's health.Vascular endothelial growth inhibitor (VEGI) is known to be abundant in endothelial cells.According to previous literature,overexpression of VEGI has been shown to inhibit tumor neovascularisation and progression in cellular and animal models,but there has been limited research on the significance of VEGI in the breast cancer.Methods In our study,cell lines MDA-MB-231 were first constructed in which VEGI mediated by lentivirus over-expressed.The effects of VEGI over-expression on MDA-MB-231 cells were investigated both in vitro and in vivo.The expression of VEGI in the MDA-MB-231 cells after infection of lentivirus was analyzed using real-time PCR and Western blotting.The effect of the biological characteristics of MDA-MB-231 cells was assessed by growth,invasion,adhesion,and migration assay with subcutaneous tumor-bearing nude mice models.Then the growth curves of the subcutaneous tumors were studied.Expressions of VEGI,CD31 and CD34 in the tumors were analyzed by immunohistochemistry and apoptosis was detected by flow cytometry and immunohistochemistry.Results Infection of MDA-MB-231 cells within the lentivirus resulted in approximately a 1 000-fold increase in the expression of VEGI.As can be seen in the invasion,adhesion and migration assay,the over-expression of VEGI can inhibit the ability of MDA-MB-231 cells during migration,adhesion and invasion.The volume of the subcutaneous tumor in the over-expression group was distinctly and significantly less than that of the control groups.Immunohistochemistry analysis of the tumor biopsies cleady showed the expression of VEGI in the over-expression group increased while CD31 and CD34 decreased significantly.In vitro and in vivo,the early apoptosis rate and the apoptosis index were increased within the VEGI over-expression group as compared with the control group.Conclusions Taken

  4. Myocardial Slit2/Robo4 expression and impact of exogenous Slit2 on proliferation and migration of cardiac microvascular endothelial cells%Slit2/Robo4信号通路对小鼠心肌微血管内皮细胞增殖和迁移的影响

    Institute of Scientific and Technical Information of China (English)

    陈桂秀; 王浩宇; 刘涛; 杨明涛; 周振宇; 冯刚

    2013-01-01

    and explore the impact of exogenous Slit2 on proliferation and migrate of mouse cardiac microvascular endothelial cells.Methods Slit2 and Robo4 expression in mouse ventricular muscle blood vessel was detected by immunohistochemistry.Slit2 and Robo4 expression in cardiac microvascular endothelial cells isolated from mouse ventricular muscle were detected by euzymelinked immunosorbent assay and immunofluorescence,respectively.The effects of various concentrations exogenous Slit2 on proliferation of mouse cardiac microvascular endothelial cells was examined by CCK-8 cell proliferation kit.Transwell chamber was used to detect migration of mouse cardiac microvascular endothelial cells treated with 800 μl M199 culture medium containing 20% FBS (negative control),10 ng/ml VEGF (positive control),100 ng/ml Slit2(Slit2) and 100 ng/ml Slit2 + 10 ng/ml VEGF (Slit2 + VEGF) and incubated for 18 h at 37 ℃ and 5% CO2.Results Both Slit2 and Robo4 protein expressions were detected in ventricular muscle blood vessel.Slit2 protein expression was detected in mouse microvascular endothelial cells.Protein and mRNA Robo4 expressions were also evidenced in mouse microvascular endothelial cells.Proliferation of mouse cardiac microvascular endothelial cells was not affected by exogenous Slit2.Migration of mouse cardiac microvascular endothelial cells was not affected by exogenous Slit2 (22.1 ± 2.8 vs.23.2 ± 3.8 in negative control,P > 0.05) and significantly enhanced by VEGF (65.3 ± 3.8,P < O.05 vs.Slip2 and negative control),this effect could be blocked by cotreatment with Slip2 (29.2 ± 3.4 in Slip2 + VEGF,P <0.05 vs.VEGF).Conclusion Slit2 and Robo4 are expressed in mouse ventricular muscle blood vessels and cardiac microvascular endothelial cells.Exogenous Slit2 has no impact on the proliferation of mouse cardiac microvascular endothelial cells but could inhibit VEGF-induced mouse cardiac microvascular endothelial cell migration.

  5. Culture supernatants of breast cancer cell line MDA-MB-231 treated with parthenolide inhibit the proliferation, migration, and lumen formation capacity of human umbilical vein endothelial cells

    Institute of Scientific and Technical Information of China (English)

    LI Cai-juan; GUO Su-fen; SHI Tie-mei

    2012-01-01

    Background Parthenolide has been tested for anti-tumor activities,such as anti-proliferation and pro-apoptosis in recent studies.However,little is known about its role in the process of tumor angiogenesis.This study aims to investigate the effects and potential mechanisms of parthenolide on the proliferation,migration and lumen formation capacity of human umbilical vein endothelial cells.Methods Different concentrations of parthenolide were applied to the human breast cancer cell line MDA-MB-231 cells.After 24-hour incubation,the culture supematants were harvested and used to treat human umbilical vein endothelial cells for 24 hours.Then an inverted fluorescence phase contrast microscope was used to evaluate the human umbilical vein endothelial cells.The secretion of vascular endothelial growth factor (VEGF),interleukin (IL)-8 and matrix metalloproteinases (MMP)-9 in the culture supernatant of the MDA-MB-231 cells was then measured with enzyme-linked immunosorbent assay (ELISA) assays.Results Suppression of proliferation,migration,and the lumen formation capacity of human umbilical vein endothelial cells was observed in the presence of the culture supernatants from the breast cancer cell line treated with different concentrations of parthenolide.Parthenolide decreased the levels of the angiogenic factors MMP-9,VEGF,and IL-8secreted by the MDA-MB-231 cells.Conclusions Parthenolide may suppress angiogenesis through decreasing angiogenic factors secreted by breast cancer cells to interfere with the proliferation,migration and lumen-like structure formation of endothelial cells,thereby inhibiting tumor growth.It is a promising potential anti-angiogenic drug.

  6. Renal and cardiac microvascular endothelium: injury and repair

    NARCIS (Netherlands)

    Oosterhuis, N.R.

    2016-01-01

    Injury to the capillary endothelium can be devastating for renal and cardiac function. To halt the progression of chronic kidney disease (CKD) and heart failure (HF) preservation of the microvascular endothelial cell (EC) function and structure is of great importance.1 Increasing knowledge about mic

  7. A Simple and Steady Method for Primary Culture of Newborn Mouse Cerebral Microvascular Endothelial Cells%一种简易稳定的乳小鼠脑微血管内皮细胞原代培养方法

    Institute of Scientific and Technical Information of China (English)

    熊永洁; 尹波; 甘莉; 王倩; 张苏明

    2012-01-01

    Objective To find a simple and steady method for primary culture of cerebral microvascular endothelial cells (CMECs). Methods The cerebral cortices of 1 to 3 day newborn C57 mice were dissected and the meninges were removed carefully,followed by digestion with two kinds of enzymes and centrifugation at 4'C. The CMECs were harvested and observed under the inverted, pna.se contrast microscopy and the transmission electron microscopy(TEM). The growth curve was drawn through MTT assay and factor Ⅷ| associated antigen was detected by using immunotluorescence analysis to characterize CMECs. Results After primary culture for 24 h,the cells appeared "polygon shaped" or "short spindle shaped" and clustered like islands. As the cells grew, they formed confluent monolayers demonstrating "swirl" or "cobblestone" morphology. The structure of tight junctions was observed under the TEM and the immunofluorescence staining revealed that the cells were posi tive for anti factorⅧ. Conclusion Using the above method, a higher purity of CMECs was obtained. This economical, simple and repetitive primary culture could offer material to the further study of the properties in the aspects of physiology, biochemis try and pharmacology,as well as the function study of the in vitro model of blood brain barrier or co culture system.%目的 寻找一种简单并稳定的原代小鼠脑微血管内皮细胞的培养方法.方法 采用两次酶消化法及4℃梯度离心法获得细胞,光镜和电镜下观察细胞形态,用检测Ⅷ因子相关抗原等方法鉴定细胞.结果 光镜下细胞呈多角形或短梭形,岛屿样聚集,随细胞生长及不断融合而出现"漩涡状"、"铺路石"样排列.电镜下观察可见紧密连接结构.Ⅷ因子相关抗原免疫荧光染色阳性.结论 上述方法可获得纯度较高的小鼠脑微血管内皮细胞,为其原代培养提供了一种经济、简易、重复性好的操作方法,也为进一步的生理、生化、药理及体

  8. A comparative study of the cell cycle status and primitive cell adhesion molecule profile of human CD34+ cells cultured in stroma-free versus porcine microvascular endothelial cell cultures.

    Science.gov (United States)

    Chute, J P; Saini, A A; Kampen, R L; Wells, M R; Davis, T A

    1999-02-01

    Porcine microvascular endothelial cells (PMVECs) plus cytokines support a rapid proliferation and expansion of human CD34+CD38- cells that are capable of multilineage engraftment within the bone marrow of a secondary host. CD34+CD38- cells contain the self-renewing, long-term culture-initiating cells (LTC-IC) that are ideal targets for retroviral gene transfer experiments. Previous experiments attempting retroviral infection of CD34+CD38- cells have failed partly because these cells do not enter cell cycle in response to cytokine combinations. In this study, we determined the cell cycle status and the cell adhesion molecule profile on purified CD34+ cells and the CD34+CD38- subset before and after ex vivo expansion on PMVECs. Purified human CD34+ cells were cocultured with PMVECs for 7 days in the presence of optimal concentrations of granulocyte/macrophage-colony-stimulating factor (GM-CSF) + interleukin (IL)-3 + IL-6 + stem cell factor (SCF) + Flt-3 ligand. The total CD34+ population and the CD34+CD38- subset increased 8.4- and 67-fold, respectively, with absolute increases in the number of colony-forming unit-granulocyte macrophage (CFU-GM) (28.2-fold), CFU-Mix (8.7 fold), and burst-forming unit-erythroid (BFU-E) (4.0-fold) progenitor cells. After 7 days of coculture with PMVECs, 44% of the CD34+CD38+ subset were found to be in G1, and 51% were in G2/S/M phase of the cell cycle. More remarkably, 53% of the CD34+CD38- subset were in G1, and 17% were in G2/S/M phase after 7 days of PMVEC coculture. In contrast, only 22% of the CD34+CD38- subset remaining after 7 days of stroma-free culture were in G1, and 6% were in G2/S/M phase. Despite the high level of cellular activation and proliferation induced by PMVEC coculture, the surface expression of adhesion molecules CD11a (LFA-1), CD11b, CD15s (sialyl-Lewis x), CD43, and CD44 (HCAM) on the total CD34+ population was maintained, and the surface expression of CD49d (VLA-4), CD54 (ICAM), CD58, and CD62L (L selectin

  9. Effect of VIM in EV71 infection in human microvascular endothelial cells%VIM在肠道病毒71型侵染人脑微血管内皮细胞中的作用

    Institute of Scientific and Technical Information of China (English)

    钟艳云; 张宝; 何明亮; 曹宇娟; 吴娴波

    2016-01-01

    Objectives To study the effect of VIM in Enterovirus 71 (EV71) infection of (human brain microvascular endothelial cells (HBMEC) and elaborating the mechanism of EV71 infection in the nervous system. Methods Knocked out the VIM by CRISPR technology , the differences in EV71 absorption , replication , release between wild VIM and VIM knocked-out (VIM-KO) HBMEC were detected by fluorescence quantitative PCR. Results 4 ℃ absorption experiment conformed that EV71 adsorption in VIM- KO is 40% less than in the normal HBMEC. After EV71 infect HBMEC for 48 h (48 h p. i.), the quantitative PCR result showed intracellular viral RNA in VIM-KO was only 1/12 of that in the normal HBMEC. Also the extracellular viral RNA was quantified, and the number of cells in VIM-KO had been reduced 1.4 times compared with the normal HBMEC. Conclusions Once VIM knocking out, EV71 attachment has been obviously reduced. Meanwhile, the level of viral RNA replication and release are decreased compared with the normal HBMEC. VIM may be an attachment receptor of EV71 in HBMEC , when the virus invades HBMEC with the binding of VIM. Moreover , VIM plays an important role in the replication and release of EV71.%目的:研究VIM在肠道病毒71型(EV71)侵染人脑微血管内皮细胞(HBMEC)中的作用,为阐明EV71感染神经系统机制提供方向。方法:采用CRISPR技术敲除VIM,通过荧光定量PCR检测EV71在对照和敲除的细胞中吸附、复制和释放的差异。结果:4℃吸附实验证实VIM敲除细胞EV71病毒的吸附量比对照细胞少40%;EV71感染HBMEC 48 h后,VIM敲除细胞EV71病毒核酸量是对照细胞的1/12。细胞培养液上清 EV71病毒核酸量检测结果表明,VIM 敲除的细胞比对照细胞少1.4倍。结论:敲除 VIM 后, EV71吸附细胞能力降低,在细胞内的复制和释放也减少。VIM可能作为HBMEC表面的EV71受体,并影响EV71在细胞中的复制和释放。

  10. Angiogenesis in male breast cancer

    Directory of Open Access Journals (Sweden)

    Kanthan Rani

    2005-03-01

    Full Text Available Abstract Background Male breast cancer is a rare but aggressive and devastating disease. This disease presents at a later stage and in a more advanced fashion than its female counterpart. The immunophenotype also appears to be distinct when compared to female breast cancer. Angiogenesis plays a permissive role in the development of a solid tumor and provides an avenue for nutrient exchange and waste removal. Recent scrutiny of angiogenesis in female breast cancer has shown it to be of significant prognostic value. It was hypothesized that this holds true in invasive ductal carcinoma of the male breast. In the context of male breast cancer, we investigated the relationship of survival and other clinico-pathological variables to the microvascular density of the tumor tissue. Methods Seventy-five cases of primary male breast cancer were identified using the records of the Saskatchewan Cancer Agency over a period of 26 years. Forty-seven cases of invasive ductal carcinoma of the male breast had formalin-fixed paraffin-embedded tissue blocks that were suitable for this study. All cases were reviewed. Immunohistochemical staining was performed for the angiogenic markers (cluster designations 31 (CD31, 34 (CD34 and 105 (CD105, von Willebrand factor (VWF, and vascular endothelial growth factor (VEGF. Microvascular density (MVD was determined using average, centre, and highest microvessel counts (AMC, CMC, and HMC, respectively. Statistical analyses compared differences in the distribution of survival times and times to relapse between levels of MVD, tumor size, node status and age at diagnosis. In addition, MVD values were compared within each marker, between each marker, and were also compared to clinico-pathological data. Results Advanced age and tumor size were related to shorter survival times. There were no statistically significant differences in distributions of survival times and times to relapse between levels of MVD variables. There was no

  11. Soluble vascular endothelial growth factor receptors 2 (sVEGFR-2) and 3 (sVEGFR-3) and breast cancer risk in the Swedish Mammography Cohort.

    Science.gov (United States)

    Harris, Holly; Wolk, Alicja; Larsson, Anders; Vasson, Marie-Paule; Basu, Samar

    2016-01-01

    Vascular endothelial growth factor (VEGF) is a signalling protein that has been established as a contributor to tumor angiogenesis, and expression of VEGF and its soluble receptors (sVEGFR2 and sVEGFR3) have been demonstrated in breast cancer cells. However, no prospective studies have examined the association between prediagnostic sVEGFR levels and breast cancer risk. We conducted a prospective case-control study nested within the Swedish Mammography Cohort examining the association between sVEGFR2 and 3 levels and breast cancer risk. The analysis included 69 incident breast cancer cases diagnosed after blood collection and 719 controls. Logistic regression models were used to calculate odds ratios and 95% confidence intervals. After adjustment for breast cancer risk factors, sVEGFR2 levels were associated with breast cancer risk (OR=1.28; 95% CI=1.06-1.56 per 1000 ng/L increase in concentration) while sVEGFR3 levels were not related to such risk (OR=1.00; 95% CI=0.93-1.07). Our results suggest that sVEGFR2 levels may be positively associated with breast cancer risk, however future studies with larger case groups are necessary to confirm this association.

  12. 超微血管成像与乳腺肿瘤微血管密度的相关性研究%Correlation analysis between superb microvascular imaging and pathological microvessel density in breast tumors

    Institute of Scientific and Technical Information of China (English)

    马燕; 李晶; 任卫东; 邓力军

    2016-01-01

    Objective: To compare the difference in the ability of blood lfow detection between superb microvascular imaging (SMI) and color Doppler flow imaging (CDFI) among benign and malignant breast tumors. The correlation between the two imaging techniques and microvessel density (MVD) were also evaluated.Methods:A total of 79 women patients with 88 lesions (48 benign, 40 malignant) were included. CDFI and SMI were used to detect blood lfows of breast masses according to Adler’s grading criteria, respectively. hTe Kappa analysis was used to assess the consistency. hTe anti-CD34 immunohistochemical staining was performed to detect the expression of microvessels in the tumor, and the correlation coeffcient of Spearman was used to analyze the correlation between the expression of MVD and the Adler classiifcation of the SMI and CDFI, respectively.Results:hTe consistency of SMI and CDFI for all tumors is unfavorable (κ=0.67), poor for malignant tumor (κ=0.39), favorable for benign tumor (κ=0.79). Compared with CDFI, SMI and MVD have a signiifcant positive correlation (r=0.82,P<0.01). Conclusion:The detection ability of SMI on microvessels of the breast malignant tumor is better than CDFI, which is signiifcantly related to MVD. SMI can be used as a non-invasive method to evaluate the microvessel of breast malignant tumor before surgery.%目的:探讨超微血管成像(superb microvascular imaging,SMI)与彩色多普勒血流显像(color Doppler flow imaging,CDFI)检出乳腺良恶性肿物的血流情况的差异,并分析二者与肿物病理微血管密度(microvessel density,MVD)的相关性。方法:对79名女性乳腺肿物患者共88个病灶(良性48个、恶性40个)进行SMI及CDFI检测,参照Adler分级标准,应用上述两种成像技术分别观察同一病灶的血流情况,Kappa分析以评估其一致性。对术后病理标本进行CD34免疫组织化学染色,检测肿瘤MVD表达,Spearman相关系数,分别分析MVD表达与两

  13. Hypoxia Affects the Structure of Breast Cancer Cell-Derived Matrix to Support Angiogenic Responses of Endothelial Cells.

    Science.gov (United States)

    Hielscher, Abigail; Qiu, Connie; Porterfield, Josh; Smith, Quinton; Gerecht, Sharon

    2013-01-01

    Hypoxia, a common feature of the tumor environment and participant in tumor progression, is known to alter gene and protein expression of several Extracellular Matrix (ECM) proteins, many of which have roles in angiogenesis. Previously, we reported that ECM deposited from co-cultures of Neonatal Fibroblasts (NuFF) with breast cancer cells, supported 3-dimensional vascular morphogenesis. Here, we sought to characterize the hypoxic ECM and to identify whether the deposited ECM induce angiogenic responses in Endothelial Cells (ECs). NuFF and MDA-MB-231 breast cancer cells were co-cultured, subjected to alternating cycles of 24 hours of 1% (hypoxia) and 21% (atmospheric) oxygen and de-cellularized for analyses of deposited ECM. We report differences in mRNA expression profiles of matrix proteins and crosslinking enzymes relevant to angiogenesis in hypoxia-exposed co-cultures. Interestingly, overt differences in the expression of ECM proteins were not detected in the de-cellularized ECM; however, up-regulation of the cell-binding fragment of fibronecin was observed in the conditioned media of hypoxic co-cultures. Ultrastructure analyses of the de-cellularized ECM revealed differences in fiber morphology with hypoxic fibers more compact and aligned, occupying a greater percent area and having larger diameter fibers than atmospheric ECM. Examining the effect of hypoxic ECM on angiogenic responses of ECs, morphological differences in Capillary-Like Structures (CLS) formed atop de-cellularized hypoxic and atmospheric ECM were not evident. Interestingly, we found that hypoxic ECM regulated the expression of angiogenic factors and matrix metalloproteinases in CLS. Overall, we report that in vitro, hypoxia does not alter the composition of the ECM deposited by co-cultures of NuFF/MDA-MB-231, but rather alters fiber morphology, and induces vascular expression of angiogenic growth factors and metalloproteinases. Taken together, these results have important implications for

  14. Investigation of the therapeutic efficacy of codelivery of psiRNA-vascular endothelial growth factor and pIL-4 into chitosan nanoparticles in the breast tumor model.

    Science.gov (United States)

    Şalva, Emine; Turan, Suna O; Kabasakal, Levent; Alan, Saadet; Özkan, Naziye; Eren, Fatih; Akbuğa, Jülide

    2014-03-01

    Angiogenesis has been known to increase tumor growth and for its metastatic potential in human tumors. Vascular endothelial growth factor (VEGF) plays an important role in tumor angiogenesis and is a promising therapeutic target for breast cancer. VEGF is an essential target for RNAi-based gene therapy of breast cancer. Interleukin-4 (IL-4) may act as an anti-angiogenic molecule that inhibits tumor growth and migration in rats. The purpose of the present study was to improve therapeutic efficacy in breast cancer with the codelivery of siRNA-expressing plasmid targeting VEGF and IL-4-expressing plasmid encapsulating into chitosan nanoparticles (NPs). The codelivery of psiVEGF and pIL-4 plasmids greatly enhanced in vitro and in vivo gene-silencing efficiency. For the in vitro study, when psiVEGF and pIL-4 into chitosan NPs were combined (81%), the gene-silencing effect was higher than psiVEGF and pIL-4 NPs alone. The in vivo study breast tumor model demonstrated that the administration of coencapsulation of psiVEGF and pIL-4 into chitosan NPs caused an additive effect on breast tumor growth inhibition (97%), compared with containing NPs psiVEGF or pIL-4 alone. These results indicate that chitosan NPs can be effectively used for the codelivery of pIL-4 and siVEGF-expressing plasmid in a combination therapy against breast cancer.

  15. Molecular characterization of glutathione S-transferase, endothelial nitric oxide synthase and Vitamin D receptor genes in breast cancer cases

    Directory of Open Access Journals (Sweden)

    Rizk El-Baz(1; Azza Ismail(2 ; Maher Amer(2; Mai Elshahat(3; Amira Kazamel(2; Ahmad Settin

    2012-10-01

    Full Text Available Background: Enzymes of the Glutathione S-transferase system (GST modulate the effects of exposure to several cytotoxic and genotoxic agents. Nitric oxide (NO is constitutively synthesized in the endothelium by endothelial nitric oxide synthase (eNOS and acts as a pleiotropic regulator involved in carcinogenesis. Vitamin D levels may influence breast cancer development. The vitamin D receptor (VDR is a crucial mediator for the cellular effects of vitamin D and additionally interacts with other cell-signaling pathways that influence cancer development. Objectives: To check for the association of polymorphisms of GST, eNOS3 and VDR genes with the susceptibility and severity of breast cancer in Egyptian cases. Subjects: This work included 100 cases with breast cancer and 100 healthy individuals. The mean age of cases was 48.31±11.40 years. They included 100 females.Methods: DNA was amplified using PCR-RFLP for detection of polymorphisms related to eNOS3 and VDR , also DNA was amplified using PCR-SSP for detection of polymorphisms related to GST and calculating the odds ratios and their 95% confidence intervals.Results: Total cases showed high significant frequency of eNOS3-786 CC (P<0.05, OR=18.58 genotypes, GSTT1(null (OR = 2.68; CI 95%=1.51-4.75; p=0.001. These were considered risk genotypes for disease susceptibility. On the other hand, total cases showed low significant frequency with homozygosity for eNOS3-786 TT (P=0.01 and the GSTT1 gene was present in 42.0% of the cancers and in 66.0% of controls (OR = 0.37; CI 95%= 0.21-0.66; p=0.001. These may be considered low risk genotypes. No significant difference in frequencies of null and present genotypes of GSTM1 and VDR FOKI in total cases compared to controls. Conclusions: Polymorphisms related to eNOS3-786, GSTT1 and VDR FOKI genes may be considered genetic markers for BC among Egyptian cases. This may have potential impact on family counselling as well as future management plans.

  16. 低频脉冲磁场对大鼠心肌微血管内皮细胞的影响%Effects of low frequency pulsed magnetic fields on cardiac microvascular endothelial cells

    Institute of Scientific and Technical Information of China (English)

    郭影; 李飞; 郭文怡; 孙冬冬; 路晓艳; 张荣庆

    2010-01-01

    Objective To investigate the effects of low frequency pulsed magnetic fields (LF-PMFs) on the proliferation of cardiac microvascular endothelial cells (CMECs) and their ultrastructure,migration and angiogenic potential. Methods CMECs from rats were exposed in vitro to low frequency square wave pulsed magnetic fields (15 Hz) 2 h/d for 5 d.The cells were randomly divided into 4 groups (control,1.0 mT,1.4 mT and 1.8 mT).After 5 days of exposure,proliferation was detected in terms of the cells' growth curves,their cycle was detected with flow cytometry,and their ultrastructure was observed using transmission electric microscopy. A scratch assay was used to evaluate the CMECs migration,and their angiogenic potential was measured using a tube formation assay.Results There was no significant effect of a 1.0 mT magnetic field on the ceils' growth curve,cell cycle or ultrastructure.The 1.4 mT magnetic field did,however,accelerate the CMECs' proliferation.The peak of the cells'growth curve was higher and moved forward,and the percentage of cells in the S phase increased significantly compared with the control group.The effects of a 1.8 mT magnetic field on S phase development were similar to those of the 1.4 mT field,but the peak of the cells' growth curve was not moved forward.After exposure to a 1.4 mT or 1.8 mT magnetic field,the CMECs' ultrastructure changed and they appeared more viable and powerful.Their nucleoil became bigger and clearer than those of the control group.There were cavernous nucleoli or two nucleoli.The number of mitochondria increased.The endoplasmic reticulum was richer and full of protein secretions inside with many microvilli on the surface.The magnetic fields facilitated migration and tube formation in the CMECs significantly,and these effects were correlated with the magnetic field intensity (1.4 mT> 1.8 mT > 1.0 mT).The cell migratory percentage in the 1.4 mT group was 86.1% ,while in the control group it was only 45.3%.When the CMECs were

  17. 大鼠原代脑微血管内皮细胞体外分离与培养的实验研究%In Vitro Isolation and Culture of Primary Rat Brain Microvascular Endothelial Cells

    Institute of Scientific and Technical Information of China (English)

    李振; 刘云会; 薛一雪; 刘丽波; 王萍

    2012-01-01

    目的 探讨获取大鼠脑微血管内皮细胞(BMEC)的简单、有效方法,为构建体外血肿瘤屏障(BTB)模型提供材料.方法 采集出生3~5 d的Wistar胎鼠大脑皮质,应用酶消化法及葡聚糖离心法获得脑微血管段后,接种于培养皿中进行原代培养,采用倒置显微镜对所培养的细胞进行形态学观察;以Ⅷ因子相关抗原免疫组化染色法鉴定细胞;将BMEC与C6脑胶质瘤细胞共培养,构建体外BTB模型,并采用免疫组化法和免疫荧光法检测BMEC间紧密连接相关蛋白occludin的表达.结果 体外培养2h时脑微血管段贴壁,12~48 h见圆形生发中心形成,2~3d单层内皮细胞自生发中心长出,4~5 d见较大单层内皮细胞团,5~7 d可见融合成片的内皮细胞单层,外观呈“铺路石”样;第Ⅷ因子免疫组化结果显示“铺路石”样细胞胞质呈棕黄色染色;紧密连接相关蛋白occludin的免疫组化和荧光结果证明共培养的BMEC间表达BTB的特性.结论 本方法能成功地进行大鼠原代BMEC培养,构建大鼠体外BTB模型,进而应用于BTB的生理、生化及药理学研究.%Objective To explore a simple and reprodueable method lor the in vitro isolation and eulture of rat brain microvascular endothelial cells (BMEC), so as to provide materials for the construction of in vilro blood tumor barrier ( BTB) models. Methods Relative!* pure cerebral microvessel fragments were obtained from the cortex of 3-5 days old wistar rats through dissection,enzyme digestion,and dex-tran centrifugation. Then, these fragments were seeded on dishes for primary culture. The morphology of BMEC was observed under an invert microscope. BMEC were identified by immunohistoehemistry with factor VIII-associated antigen. In vitro BTB models were constructed by co-cultivation of BMEC with C6 glioma cells. The expression of tight junction-related protein oeeludin in BMEC of BTB was measured by immunohistoehemistry and

  18. Effects of amelogenins on angiogenesis-associated processes of endothelial cells

    DEFF Research Database (Denmark)

    Almqvist, S; Kleinman, H K; Werthén, M;

    2011-01-01

    To study the effects of an amelogenin mixture on integrin-dependent adhesion, DNA synthesis and apoptosis of cultured human dermal microvascular endothelial cells and angiogenesis in an organotypic assay.......To study the effects of an amelogenin mixture on integrin-dependent adhesion, DNA synthesis and apoptosis of cultured human dermal microvascular endothelial cells and angiogenesis in an organotypic assay....

  19. Source of the seed cell in bone tissue engineering revascularization:renal microvascular endothelial cell cultured in vitro and its cellular phenotype in rabbits%骨组织工程再血管化种子细胞来源:兔肾微血管内皮细胞体外培养及其细胞表型的研究

    Institute of Scientific and Technical Information of China (English)

    王明海; 吴明暖; 张锡庆; 王晓东; 张亚; 王科文; 尹航; 冯林; 刘柏龄

    2004-01-01

    vitro in a short period with relatively high purity can be used as ideal seed cells in revascularization for tissue-engineered bone.OBJECTIVE: To explore the culture in vitro method for renal microvascular endothelial cells for its verification and phenotype study, which is prepared as ideal seed cell for revascularization in bone tissue engineering.DESIGN: Single sample study.SETTING: Department of Orthopaedics of Children' s Hospital affiliated to Soochow University, Institute of Biotechnology, Academy for Life Science,Soochow University.MATERIALS: Study was accomplished in the Laboratory of the Department of Orthopaedics of Children' s Hospital affiliated to Soochow University during May 2003 to April 2004. Experimental materials include CO2 incubator,M199, fetal calf serum, Ⅳ collagenase, 40 g/L of paraform, 20 g/L of cidex,ECGF, mouse-anti-human FⅧ monoclonal antibody(anti-FⅧ-RAg), multi-marked immune analyser(1 420, WALLAC BLCTOR2), centrifuge, inverted microscope, and immunofluorescence microscope, etc.METHODS: Three-step gradient sieve method was used. Glomerulus of relatively higher purity was obtained at first, and then rental microvascular endothelial cells were cultured in vitro with glomerulus external implantation method. The rental microvascular endothelial cells were further verified by immunohistochemical Ⅷ factor-related antigen. Cytoplasmic Weibel-Palade body was observed under tranmision microscope. The expression of CD31,CD34 and CD44 was detected with indirect immunofluorescence method.MAIN OUTCOME MEASURES: The morphology, activity and phenotype of renal microvascular endothelial cell.RESULTS: Vascular endothelial cells in very high purity were cultured in vitro with 11 passages. Besides, immunohistochemistry revealed positive reaction to Ⅷ-related antigen, Weibel-Palade body was observed under electron microscope, and the expressions of CD31 and CD34 were positive but CD44 was negative under immunofluorescence analysis.CONCLUSION: This

  20. Skin microvascular reactivity in patients with hypothyroidism.

    Science.gov (United States)

    Mihor, Ana; Gergar, Maša; Gaberšček, Simona; Lenasi, Helena

    2016-11-04

    Hypothyroidism is associated with impaired vascular function; however, little is known about its impact on microcirculation. We aimed to determine skin microvascular reactivity in hypothyroidism focusing on endothelial function and the sympathetic response. We measured skin laser Doppler (LD) flux (LDF) on the volar forearm and the finger pulp using LD flowmetry in hypothyroid patients (N = 13) and healthy controls (N = 15). Skin microvascular reactivity was assessed by a three-minute occlusion of the brachial artery, inducing postocclusive reactive hyperaemia (PRH), and by a four-minute local cooling of the hand. An electrocardiogram (ECG), digital artery blood pressure and skin temperature at the measuring sites were recorded. Baseline LDF, the digital artery blood pressure and the heart rate were comparable between patients and controls. On the other hand, patients exhibited significantly longer PRH duration, significantly higher blood pressure during cooling (unpaired t-test, p skin microcirculation and an apparent increase in sympathetic reactivity after local cooling in hypothyroid patients. Hypothyroidism induces subtle changes of some haemodynamic parameters in skin microcirculation implying altered endothelial function and altered sympathetic reactivity.

  1. LGALS3BP, lectin galactoside-binding soluble 3 binding protein, induces vascular endothelial growth factor in human breast cancer cells and promotes angiogenesis.

    Science.gov (United States)

    Piccolo, Enza; Tinari, Nicola; Semeraro, Daniela; Traini, Sara; Fichera, Imma; Cumashi, Albana; La Sorda, Rossana; Spinella, Francesca; Bagnato, Anna; Lattanzio, Rossano; D'Egidio, Maurizia; Di Risio, Annalisa; Stampolidis, Pavlos; Piantelli, Mauro; Natoli, Clara; Ullrich, Axel; Iacobelli, Stefano

    2013-01-01

    Elevated serum or tissue levels of lectin galactoside-binding soluble 3 binding protein (LGALS3BP) have been associated with short survival and development of metastasis in a variety of human cancers. However, the role of LGALS3BP, particularly in the context of tumor-host relationships, is still missing. Here, we show that LGALS3BP knockdown in MDA-MB-231 human breast cancer cells leads to a decreased adhesion to fibronectin, a reduced transendothelial migration and, more importantly, a reduced expression of vascular endothelial growth factor (VEGF). Production of VEGF, that was restored by exposure of silenced cells to recombinant LGALS3BP, required an intact PI3k/Akt signaling. Furthermore, we show that LGALS3BP was able to directly stimulate HUVEC tubulogenesis in a VEGF-independent, galectin-3-dependent manner. Immunohistochemical analysis of human breast cancer tissues revealed a correlation among LGALS3BP expression, VEGF expression, and blood vessel density. We propose that in addition to its prometastatic role, LGALS3BP secreted by breast cancer cells functions critically as a pro-angiogenic factor through a dual mechanism, i.e by induction of tumor VEGF and stimulation of endothelial cell tubulogenesis.

  2. Immunohistochemical analysis of cancer stem cell markers in invasive breast carcinoma and associated ductal carcinoma in situ: relationships with markers of tumor hypoxia and microvascularity.

    Science.gov (United States)

    Currie, Margaret J; Beardsley, Brooke E; Harris, Gavin C; Gunningham, Sarah P; Dachs, Gabi U; Dijkstra, Birgit; Morrin, Helen R; Wells, J Elisabeth; Robinson, Bridget A

    2013-03-01

    We performed immunohistochemical analysis of 3 cancer stem cell-related markers (CD44(+)/CD24(-/low), aldehyde dehydrogenase [ALDH]-1, CD133) in 94 invasive ductal carcinomas and assessed relationships with markers of hypoxia (carbonic anhydrase IX [CAIX]), tumor microvessel density (CD31), and clinicopathologic variables. Overall, 10% of tumors were CD44(+)/CD24(-/low), 13% were ALDH-1(+), 25% were CD133(+), 35% were immunonegative, and 1 tumor was immunopositive for all 3 markers. Associated ductal carcinoma in situ (DCIS) was present in 48% of tumors. Marker immunopositivity was detected in DCIS in 13% (CD44(+)/CD24(-/low)), 7% (ALDH-1(+)), and 32% (CD133(+)) of these tumors and was more likely present in DCIS when also detected in the invasive compartment (P = .03, P = .001, and P = .009, respectively). CD44(+)/CD24(-/low) cells were more common in progesterone receptor-negative tumors (P breast cancers (P breast cancer and showed that CD44(+)/CD24(-/low) and CD133(+) cells were more frequently observed in hypoxic regions of tumor, whereas ALDH-1(+) cells more commonly colocalized to tumors with high microvessel density.

  3. Monitoring in microvascular surgery.

    Science.gov (United States)

    Furnas, H; Rosen, J M

    1991-03-01

    The importance of monitoring in microvascular surgery is underscored by the high reported salvage rates of failing free flaps and replants. In this overview, we begin by defining the physiology of ischemic tissue with emphasis given to the no-reflow phenomenon and the secondary critical ischemia times. Based on the physiological changes accompanying ischemia, several variables are defined that can be monitored to reflect the vascular state of a free flap or replant. Multifarious monitoring systems are then reviewed, including clinical observation, temperature, isotope clearance, ultrasonic Doppler, laser Doppler, transcutaneous oxygen tension, reflection plethysmography, dermofluorometry, pH, electromagnetic flowmetry, serial hematocrits, interstitial fluid pressure, and magnetic resonance imaging.

  4. 细胞饥饿及 TNF-α干预后内皮细胞膜微粒对人脑微血管内皮细胞的影响%Effects of cell starvation and TNF-αderived endothelial microvesicles on human brain microvascular endothelial cells

    Institute of Scientific and Technical Information of China (English)

    潘群文; 何彩霞; 刘雅静; 张惠婷; 王艳; 戴炳琰; 马晓瑭

    2015-01-01

    目的:探讨饥饿及TNF-α刺激条件下,内皮细胞膜微粒( EMVs )对人脑微血管内皮细胞( HBMECs )增殖、迁移及血管生成功能的影响。方法体外培养HBMECs,分为PBS组、饥饿组、TNF-α组。于饥饿24 h后提取饥饿后产生的EMVs( sHB-MVs),于TNF-α刺激24 h后提取TNF-α刺激下产生的EMVs(αHB-MVs)。将sHB-MVs与αHB-MVs按照1×108个/mL、每孔10μL分别与饥饿组及TNF-α组HBMECs共培养,PBS组给予10μL PBS处理。采用MTT法测定各组HBMECs增殖能力,倒置显微镜下测定HBMECs迁移距离,观察血管形成数。结果饥饿组增殖能力高于PBS组(P<0.01),TNF-α组HBMECs增殖能力低于PBS组(P<0.01)。饥饿组迁移距离长于PBS组(P<0.05),TNF-α组迁移距离短于PBS组(P<0.01)。饥饿组血管生成数多于PBS组(P<0.01),TNF-α组血管生成数少于PBS组(P<0.01)。结论饥饿刺激下产生的EMVs可促进HBMECs增殖、迁移及血管生成功能,TNF-α刺激下产生的EMVs可抑制HBMECs增殖、迁移及血管生成功能。%Objective To investigate the effects of cell starvation and TNF-αderived endothelial microvesicles (EMVs) on the proliferation, migration and angiogenesis of human brain microvascular endothelial cells (HBMECs) in vitro.Methods HBMECs were cultured and were divided into the PBS group, starvation group and TNF-αgroup.EMVs were extracted from HB-MECs cultured in a serum deprivation (SD) medium (starving stress, sHB-MVs) or SD medium containing tumor necrosis fac-tor-(TNF-α) (apoptotic stress,αHB-MVs).The HBMECs of the starvation group and TNF-αgroup were cultured with sHB-MVs andαHB-MVs (1 ×108/mL, 10μL each hole), respectively;and the PBS group was treated with 10μL PBS.The prolif-eration of HBMECs in each group was determined by MTT, the migration distance of HBMECs was measured by inverted micro-scope and the number of angiogenesis was observed under the

  5. In Vitro Endothelialization Test of Biomaterials Using Immortalized Endothelial Cells.

    Directory of Open Access Journals (Sweden)

    Ken Kono

    Full Text Available Functionalizing biomaterials with peptides or polymers that enhance recruitment of endothelial cells (ECs can reduce blood coagulation and thrombosis. To assess endothelialization of materials in vitro, primary ECs are generally used, although the characteristics of these cells vary among the donors and change with time in culture. Recently, primary cell lines immortalized by transduction of simian vacuolating virus 40 large T antigen or human telomerase reverse transcriptase have been developed. To determine whether immortalized ECs can substitute for primary ECs in material testing, we investigated endothelialization on biocompatible polymers using three lots of primary human umbilical vein endothelial cells (HUVEC and immortalized microvascular ECs, TIME-GFP. Attachment to and growth on polymer surfaces were comparable between cell types, but results were more consistent with TIME-GFP. Our findings indicate that TIME-GFP is more suitable for in vitro endothelialization testing of biomaterials.

  6. Microvascular Recruitment in Insulin Resistance

    DEFF Research Database (Denmark)

    Sjøberg, Kim Anker

    In this PhD work a new method for measuring microvascular recruitment was developed and evaluated, using continues real-time imaging of contrast enhanced ultrasound. Gas-filled microbubbles were infused intravenously and by taking advantage of the echogenic properties of the microbubbles the reso......In this PhD work a new method for measuring microvascular recruitment was developed and evaluated, using continues real-time imaging of contrast enhanced ultrasound. Gas-filled microbubbles were infused intravenously and by taking advantage of the echogenic properties of the microbubbles...... action in the microvasculature and restored normal microvascular function by increasing the microvascular recruitment similar to in control animals. This effect of GLP-1 on microvascular recruitment was associated with a restoration of both whole body insulin sensitivity and muscle glucose uptake when co...

  7. 石杉碱甲保护人脑微血管内皮细胞损伤的体外实验研究%Protect Effects of Huperzine A on Methylglyoxal Induced Injury in the Cultured Human Brain Microvascular Endothelial Cell in Vitro Experimental Study

    Institute of Scientific and Technical Information of China (English)

    姜松国; 徐磊; 柴冬梅; 朱辉武; 林铮

    2015-01-01

    目的 体外实验研究石杉碱甲对人脑微血管内皮细胞(human brain microvascular endothelial cells,HBMEC)损伤的保护作用和机制.方法 在培养的HBMEC上,利用丙酮醛诱导细胞损伤,通过MTT检测细胞活力,LDH、SOD活性试剂盒及caspase-3活性试剂盒检测细胞损伤情况,观察石杉碱甲的作用和机制.结果 石杉碱甲呈浓度依赖地保护MGO诱导的细胞损伤,在10-5 mol·L-1时呈最大保护作用.丙酮醛能诱导HBMEC的SOD活性下降,而石杉碱甲(10-6,10-5mol·L-1)能逆转这种作用.进一步研究发现石杉碱甲能抑制丙酮醛诱导的caspase-3活性上升.结论 石杉碱甲对丙酮醛诱导的HBMEC的损伤具有保护作用,这可能与其抗自由基和抗凋亡作用有关.

  8. Sesamin inhibits macrophage-induced vascular endothelial growth factor and matrix metalloproteinase-9 expression and proangiogenic activity in breast cancer cells.

    Science.gov (United States)

    Lee, Chun-Chung; Liu, Ko-Jiunn; Wu, Yu-Chen; Lin, Sue-Jane; Chang, Ching-Chun; Huang, Tze-Sing

    2011-06-01

    Sesamin is a sesame component with antihypertensive and antioxidative activities and has recently aroused much interest in studying its potential anticancer application. Macrophage is one of the infiltrating inflammatory cells in solid tumor and may promote tumor progression via enhancement of tumor angiogenesis. In this study, we investigated whether sesamin inhibited macrophage-enhanced proangiogenic activity of breast cancer cell lines MCF-7 and MDA-MB-231. Using vascular endothelial cell capillary tube and network formation assays, both breast cancer cell lines exhibited elevated proangiogenic activities after coculture with macrophages or pretreatment with macrophage-conditioned medium. This elevation of proangiogenic activity was drastically suppressed by sesamin. Vascular endothelial growth factor (VEGF) and matrix metalloproteinase-9 (MMP-9) induced by macrophages in both cell lines were also inhibited by sesamin. Nuclear levels of HIF-1α and NF-κB, important transcription factors for VEGF and MMP-9 expression, respectively, were obviously reduced by sesamin. VEGF induction by macrophage in MCF-7 cells was shown to be via ERK, JNK, phosphatidylinositol 3-kinase, and NF-κB-mediated pathways. These signaling molecules and additional p38(MAPK) were also involved in macrophage-induced MMP-9 expression. Despite such diverse pathways were induced by macrophage, only Akt and p38(MAPK) activities were potently inhibited by sesamin. Expression of interleukin (IL)-6, IL-8, and tumor necrosis factor-α were substantially increased and involved in macrophage-induced VEGF and MMP-9 mRNA expression in MCF-7 cells. Sesamin effectively inhibited the expression of these cytokines to avoid the reinforced induction of VEGF and MMP-9. In conclusion, sesamin potently inhibited macrophage-enhanced proangiogenic activity of breast cancer cells via inhibition of VEGF and MMP-9 induction.

  9. Dauricine inhibits insulin-like growth factor-Ⅰ-induced hypoxia inducible factor 1α protein accumulation and vascular endothelial growth factor expression in human breast cancer cells

    Institute of Scientific and Technical Information of China (English)

    Xu-dong TANG; Xin ZHOU; Ke-yuan ZHOU

    2009-01-01

    Aim: To investigate the effects of dauricine (Dau) on insulin-like growth factor-Ⅰ (IGF-Ⅰ)-induced hypoxia inducible factor 1α (HIF-1α) and vascular endothelial growth factor (VEGF) expression in human breast cancer cells (MCF-7).Methods: Serum-starved MCF-7 cells were pretreated for 1 h with different concentrations of Dau, followed by incubation with IGF-Ⅰ for 6 h. HIF-1α and VEGF protein expression levels were analyzed by Western blotting and ELISA, respectively.HIF-1α and VEGF mRNA levels were determined by real-time PCR. In vitro angiogenesis was observed via the human umbilical vein endothelial cell (HUVEC) tube formation assay. An in vitro invasion assay on HUVECs was performed.Results: Dau significantly inhibited IGF-Ⅰ-induced HIF-1α protein expression but had no effect on HIF-1α mRNA expression. However, Dau remarkably suppressed VEGF expression at both protein and mRNA levels in response to IGF-Ⅰ.Mechanistically, Dau suppressed IGF-Ⅰ-induced HIF-1α and VEGF protein expression mainly by blocking the activation of PI-3K/AKT/mTOR signaling pathway. In addition, Dan reduced IGF-Ⅰ-induced HIF-1α protein accumulation by inhibiting its synthesis as well as by promoting its degradation. Functionally, Dau inhibited angiogenesis in vitro. Moreover, Dau had a direct effect on IGF-Ⅰ-induced invasion of HUVECs.Conclusion: Dau inhibits human breast cancer angiogenesis by suppressing HIF-1α protein accumulation and VEGF expression, which may provide a novel potential mechanism for the anticancer activities of Dau in human breast cancer.

  10. Role of NF-κB activation in matrix metalloproteinase 9, vascular endothelial growth factor and interleukin 8 expression and secretion in human breast cancer cells.

    Science.gov (United States)

    Li, Caijuan; Guo, Sufen; Shi, Tiemei

    2013-04-01

    The aims of this study were to assess the effects and potential mechanisms of parthenolide on the expression of vascular endothelial growth factor (VEGF), interleukin 8 (IL-8) and matrix metalloproteinase 9 (MMP-9) in human breast cancer cell line MDA-MB-231. After incubation with different concentrations of parthenolide for 24 h, MDA-MB-231 cells were collected, and the expressions of VEGF, IL-8 and MMP-9 were measured by real-time PCR and Western blot. The secretions of VEGF, IL-8 and MMP-9 in culture supernatant of MDA-MB-231 cells were then measured with ELISA assays. The NF-κB DNA-binding activity of breast cancer cells treated with parthenolide was analyzed using electrophoretic mobility assays. The real-time PCR and Western blot data showed that the expressions of VEGF, IL-8 and MMP-9 were significantly inhibited by parthenolide at both transcription level and protein level in MDA-MB-231 cells. ELISA results also confirmed these effects at a secretion level. The electrophoretic mobility assay results demonstrated that parthenolide can inhibit NF-κB DNA-binding activity of the breast cancer cells. Hence, the expression of VEGF, IL-8 and MMP-9 may be suppressed by parthenolide through the inhibition of NF-κB DNA-binding activity in MDA-MB-231 cells.

  11. Cell proliferation along vascular islands during microvascular network growth

    Directory of Open Access Journals (Sweden)

    Kelly-Goss Molly R

    2012-06-01

    Full Text Available Abstract Background Observations in our laboratory provide evidence of vascular islands, defined as disconnected endothelial cell segments, in the adult microcirculation. The objective of this study was to determine if vascular islands are involved in angiogenesis during microvascular network growth. Results Mesenteric tissues, which allow visualization of entire microvascular networks at a single cell level, were harvested from unstimulated adult male Wistar rats and Wistar rats 3 and 10 days post angiogenesis stimulation by mast cell degranulation with compound 48/80. Tissues were immunolabeled for PECAM and BRDU. Identification of vessel lumens via injection of FITC-dextran confirmed that endothelial cell segments were disconnected from nearby patent networks. Stimulated networks displayed increases in vascular area, length density, and capillary sprouting. On day 3, the percentage of islands with at least one BRDU-positive cell increased compared to the unstimulated level and was equal to the percentage of capillary sprouts with at least one BRDU-positive cell. At day 10, the number of vascular islands per vascular area dramatically decreased compared to unstimulated and day 3 levels. Conclusions These results show that vascular islands have the ability to proliferate and suggest that they are able to incorporate into the microcirculation during the initial stages of microvascular network growth.

  12. A model of physical factors in the structural adaptation of microvascular networks in normotension and hypertension

    DEFF Research Database (Denmark)

    Jacobsen, Jens Christian Brings; Gustafsson, Finn; Holstein-Rathlou, N.-H.

    2003-01-01

    hemodynamic stimuli in the individual vessel. Simulated results show emanating microvascular networks with properties similar to those observed in vivo. The model points to an altered endothelial function as a key factor in the development of vascular changes characteristic of hypertension....

  13. Cytostatic and pro-apoptotic effects of a novel phenylacetate-dextran derivative (NaPaC) on breast cancer cells in interactions with endothelial cells.

    Science.gov (United States)

    Malherbe, Séverine; Crépin, Michel; Legrand, Chantal; Wei, Ming Xing

    2004-11-01

    We have tested a novel hybrid molecule made of carboxymethylbenzylamide dextran (CMDB) and sodium phenylacetate (NaPa) groups, called the CMDB-NaPa ester (NaPaC), on the proliferation of breast cancer and endothelial cells as well as paracrine effects between these two cell types. Our results showed that NaPaC inhibited the proliferation of MDA-MB-231 cells and MCF-7 cells in a dose-dependent manner. NaPaC was 20-fold more active on highly invasive MDA-MB-231 cells than the NaPa parental molecule. On MCF-7 cells, which present a less aggressive phenotype, NaPaC was only 3-fold more active than the NaPa parental molecule. Furthermore, NaPaC had only a slight effect on the proliferation of primary cultured endothelial cells (HUVEC). A cytostatic effect of NaPaC on tumor cells was observed with cells accumulating in G0/G1 phase after 96 h of treatment. In addition, NaPaC induced a strong apoptotic effect on the two breast cancer cell lines. Conditioned media (CM) from tumor cells inhibited HUVEC proliferation, and this effect was enhanced in the presence of NaPaC (6 mM) and NaPa (10 mM). In addition to this cytostatic effect, CM from tumor cells induced a HUVEC early apoptosis which was increased, mainly, in the presence of NaPa (15 mM). Thus, this study shows that NaPaC is a more powerful anti-proliferative molecule than its parental NaPa molecule, with cytostatic and pro-apoptotic effects on MDA-MB-231 and MCF-7 tumor cells. Also, both molecules increased a pro-apoptotic effect of tumor cells on HUVEC.

  14. 小鼠空肠黏膜微血管内皮细胞糖萼的透射电镜观察%Transmission electron microscopic observation of microvascular endothelial glycocalyx in mouse jejunal mucosa

    Institute of Scientific and Technical Information of China (English)

    吴双; 张涛; 冯波; 穆祥

    2013-01-01

    Objective This study aimed to establish a reliable displaying method of the endothelial glycocalyx in small intestinal mucosal microvessels, and to provide a foundation for further studies on its functions. Methods Six mice were divided into a control group and a cardiac perfusion group, three mice each. The mice were anesthetized by inhalation of ether. The jejunum from the cardiac perfusion group was sampled after they were fixed by cardiac perfusion with gradient alcian blue solution and the jejunum from control group was directly sampled. The jejunal mucosa was prepared as ultrathin sections according to the routine method for transmission electron microscopic (TEM) analysis. Results By TEM, the endothelial glycocalyx was shown in jejunal mucosal microvessels of cardiac perfusion group mice, which had two forms, the continued and the discontinued. The continued glycocalyx was 20-30 nm thick, and the discontinued was 50-100 nm. However, the glycocalyx was not found in the control group. Conclusion The endothelial glycocalyx in jejunal mucosal mirovessels is not observaed by TEM in the samples prepared with the traditional method. The cardiac perfusion fixation with alcian blue solution could better preserves the glycocalyx for TEM analysis, and is a reliable method for its study.%目的 建立可靠的小肠黏膜微血管内皮糖萼显示方法,为探讨其生物学功能提供基础.方法 取小鼠6只,分为对照组和心脏灌注组,每组3只.对小鼠行乙醚吸入麻醉,心脏灌注组使用梯度阿尔新蓝溶液心脏灌注固定后取空肠组织,对照组直接取空肠组织,然后按照常规方法对其黏膜层制备超薄切片,透射电镜观察.结果 透射电镜观察显示,心脏灌注组小鼠肠黏膜微血管内皮细胞膜表面有明显的糖萼结构,呈不连续型和连续型两种形态,不连续性糖萼厚度为50~ 100nm,连续型糖萼厚度为20 ~ 30nm;而对照组未能观察到明显的糖萼结构.结论 常规透

  15. Tumor-Endothelial Interaction Links the CD44+/CD24- Phenotype with Poor Prognosis in Early-Stage Breast Cancer

    Directory of Open Access Journals (Sweden)

    Martin Buess

    2009-10-01

    Conclusions Our results suggest that the interaction of endothelial cells with tumor cells that express the CD44+/CD24- signature, which indicates a low proliferative potential, might explain the unexpected and paradoxical association of the CD44+/CD24- signature with highly proliferative tumors that have an unfavorable prognosis.

  16. Prognostic impact of placenta growth factor and vascular endothelial growth factor A in patients with breast cancer

    DEFF Research Database (Denmark)

    Maae, Else; Olsen, Dorte Aalund; Steffensen, Karina Dahl;

    2012-01-01

    Placenta growth factor (PlGF) and vascular endothelial growth factor A (VEGF-A) are angiogenic growth factors interacting competitively with the same receptors. VEGF-A is essential in both normal and pathologic conditions, but the functions of PlGF seem to be restricted to pathologic conditions...

  17. Blood-based biomarkers of microvascular pathology in Alzheimer's disease.

    LENUS (Irish Health Repository)

    Ewers, Michael

    2012-02-01

    Sporadic Alzheimer\\'s disease (AD) is a genetically complex and chronically progressive neurodegenerative disorder with molecular mechanisms and neuropathologies centering around the amyloidogenic pathway, hyperphosphorylation and aggregation of tau protein, and neurofibrillary degeneration. While cerebrovascular changes have not been traditionally considered to be a central part of AD pathology, a growing body of evidence demonstrates that they may, in fact, be a characteristic feature of the AD brain as well. In particular, microvascular abnormalities within the brain have been associated with pathological AD hallmarks and may precede neurodegeneration. In vivo assessment of microvascular pathology provides a promising approach to develop useful biological markers for early detection and pathological characterization of AD. This review focuses on established blood-based biological marker candidates of microvascular pathology in AD. These candidates include plasma concentration of vascular cell adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1) that are increased in AD. Measures of endothelial vasodilatory function including endothelin (ET-1), adrenomedullin (ADM), and atrial natriuretic peptide (ANP), as well as sphingolipids are significantly altered in mild AD or during the predementia stage of mild cognitive impairment (MCI), suggesting sensitivity of these biomarkers for early detection and diagnosis. In conclusion, the emerging clinical diagnostic evidence for the value of blood-based microvascular biomarkers in AD is promising, however, still requires validation in phase II and III diagnostic trials. Moreover, it is still unclear whether the described protein dysbalances are early or downstream pathological events and how the detected systemic microvascular alterations relate to cerebrovascular and neuronal pathologies in the AD brain.

  18. Diagnostic Power of Vascular Endothelial Growth Factor and Macrophage Colony-Stimulating Factor in Breast Cancer Patients Based on ROC Analysis

    Directory of Open Access Journals (Sweden)

    Monika Zajkowska

    2016-01-01

    Full Text Available Breast cancer (BC is the most common malignancy in women. Vascular endothelial growth factor (VEGF has been described as an important regulator of angiogenesis which plays a vital role in the progression of tumor. Macrophage colony-stimulating factor (M-CSF is a cytokine whose functions include regulation of hematopoietic lineages cells growth, proliferation, and differentiation. We investigated the diagnostic significance of these parameters in comparison to CA15-3 in BC patients and in relation to the control group (benign breast tumor and healthy women. Plasma levels of the tested parameters were determined by ELISA and CA15-3 was determined by CMIA. VEGF was shown to be comparable to CA15-3 values of sensitivity in BC group and, what is more important, higher values in early stages of BC. VEGF was also the only parameter which has statistically significant AUC in all stages of cancer. M-CSF has been shown to be comparable to CA15-3 and VEGF, specificity, and AUC values only in stages III and IV of BC. These results indicate the usefulness and high diagnostic power of VEGF in the detection of BC. Also, it occurred to be the best candidate for cancer diagnostics in stages I and II of BC and in the differentiation between BC and benign cases.

  19. Silencing of osteopontin promotes the radiosensitivity of breast cancer cells by reducing the expression of hypoxia inducible factor 1 and vascular endothelial growth factor

    Institute of Scientific and Technical Information of China (English)

    YANG Li; ZHAO Wei; ZUO Wen-shu; WEI Ling; SONG Xian-rang; WANG Xing-wu; ZHENG Gang; ZHENG Mei-zhu

    2012-01-01

    Background Osteopontin (OPN) is a secreted phosphoglycoprotein (SSP) that is overexpressed in a variety of tumors and was regarded as a molecular marker of tumors.In this study,we intended to demonstrate the role of OPN in human breast cancer cell line MDA-MB-231.Methods Recombinant plasmid expressing small interfering RNA (siRNA) specific to OPN mRNA was transfected into MDA-MB-231 cells to generate the stable transfected cell line MDA-MB-343,and the empty plasmid tansfected cells (MDA-MB-neg) or wildtype MDA-MB-231 cells were used as control cells respectively.Expression of OPN,hypoxia inducible factor-1 (HIF-1) and vascular endothelial growth factor (VEGF) proteins was analyzed by Western blotting analysis.The radiosensitivity of cells was determined by detecting cell apoptosis,cell proliferation and cell senescence.Results HIF-1 and VEGF proteins in MDA-MB-343 cells were significantly downregulated upon the efficient knockdown of OPN expression under either hypoxia or normoxia environment.Moreover,expression of OPN protein was upregualted upon hypoxic culture.Stable OPN-silencing also decreased cell invasion,increased cell apoptosis and cell senescence,as well as reduced clonogenic survival,resulting in increase radiation tolerance.Conclusions Suppression of OPN gene expression can enhance radiosensitivity and affect cell apoptosis in breast cancer cells.OPN seems to be an attractive target for the improvement of radiotherapy.

  20. 通心络干预的脑微血管内皮细胞条件液对大鼠脑皮层神经元的影响%Effect of Tongxinluo Intervened Brain Microvascular Endothelial Cells Conditioned Medium on Cortical Neuron of Rats

    Institute of Scientific and Technical Information of China (English)

    盖聪; 李澎涛; 孙红梅; 李卫红; 张振强; 李聪; 于慧玲; 贾静

    2014-01-01

    目的:观察通心络干预的脑微血管内皮细胞条件培养液对大鼠脑皮层神经元氧化应激反应的影响,探讨通心络在脑微血管内皮细胞缺血损伤状态下保护神经元的机制。方法:首先制备4种大鼠脑微血管内皮细胞(BMEC)条件培养液:①正常BMEC 条件液(N-CM);②正常 BMEC 加通心络药物血清条件液(NT-CM);③拟缺血损伤 BMEC 条件液(I-CM);④损伤 BMEC加通心络药物血清条件液(IT-CM)。将它们分别作用于正常和糖氧剥夺损伤(拟缺血)的大鼠皮层神经元后,测定神经元活性、超氧化物歧化酶(SOD)和丙二醛(MDA)含量。结果:① I-CM 可使正常神经元的活性和 SOD 活力下降,MDA 含量增加;与I-CM 相比,IT-CM 可提高神经元活性和 SOD 活力,降低 MDA 的含量。②与正常神经元比较,拟缺血神经元活性和 SOD 活力明显下降,MDA 含量增加;I-CM 进一步使拟缺血损伤神经元的活力下降;NT-CM 和 IT-CM 在一定程度上可阻抑损伤神经元活性和 SOD 活力的下降,并降低 MDA 含量。结论:大鼠脑微血管内皮细胞损伤后可能造成其旁分泌功能紊乱,进一步导致神经元的损伤。通心络可能通过调节内皮细胞的旁分泌功能保护神经元,该保护作用与减少神经元的氧化应激有关。%Objective:To explore the effect of conditioned medium of rat brain microvascular endothelial cells on oxidative stress of corti-cal neurons and the protective effect of Tongxinluo( TXL)on it. Methods:Four kinds of rats brain microvascular endothelial cell (BMEC)conditioned cultured medium were prepared:①conditioned medium of normal BMEC(N-CM);②conditioned medium of nor-mal BMEC with drug treatment( NT-CM);③ conditioned medium of ischemic BMEC( I-CM);④ conditioned medium of ischemic BMEC with drug treatment(IT-CM). Each type of conditioned medium were applied to BMEC of normal and sugar

  1. Correlation between the expression of vascular endothelial growth factor c and C-erbB-2 in human breast cancer

    Institute of Scientific and Technical Information of China (English)

    Shuxian Qu; Zhendong Zheng; Zhaozhe Liu; Liang Liu; Miao Zhang; Yaling Han; Xiaodong Xie

    2014-01-01

    Objective: We aimed to study the transcription level of VEGF-C in human breast cancer tissue, and explore the correlations with the expression of C-erbB-2.Methods: The expression of VEGF-C mRNA in 51 cases of human breast cancer was assessed by hybridization in situ. The expressions of C-erbB-2 was assessed by immunohistochemistry.Results:The positive rate of VEGF-C mRNA was 54.9% in 51 cases of breast cancer. The transcription level had correlation with tumor size and status of lymph nodes (P < 0.05). The expression of VEGF-C mRNA had a positive correlation with the expression of C-erbB-2 (P < 0.05).Conclusion: The up-expression of VEGF-C has a significant correlation with the malignancy level and clinical stage of breast cancer. The combined detection of VEGF-C, C-erbB-2 may help to estimate the prognosis of patients with breast cancer and study on thetherapeutic implications.

  2. Comparison of methods of microvascular staining and quantification in prostate carcinoma

    DEFF Research Database (Denmark)

    Offersen, Birgitte Vrou; Borre, Michael; Sørensen, Flemming Brandt;

    2002-01-01

    High microvascular density in prostate carcinomas may indicate poor prognosis. Our aim was to compare two different anti-endothelial antibodies and two different ways of evaluating microvascular scores in hot spots (microvessel density (MVD) and Chalkley counts). Adjacent serial sections of forma......High microvascular density in prostate carcinomas may indicate poor prognosis. Our aim was to compare two different anti-endothelial antibodies and two different ways of evaluating microvascular scores in hot spots (microvessel density (MVD) and Chalkley counts). Adjacent serial sections...... of formalin-fixed, paraffin-embedded tumor specimens from TURPs on 51 consecutive patients with prostate carcinoma were immunostained for CD34 and von Willebrand Factor (vWF). Estimates of microvascular density were based on projecting a 10 x 10 grid or a Chalkley grid onto a vascular hot spot of the invasive...... prostate carcinoma. Anti-CD34 antibodies stained microvessels in all 51 tumors, whereas anti-vWF antibodies in 6 tumors resulted in intense background staining causing omission of these. Anti-CD34 antibodies highlighted significantly more microvessels than anti-vWF antibodies, and the anti-CD34 vascular...

  3. Modified Continuous Loop Technique for microvascular anastomosis

    Directory of Open Access Journals (Sweden)

    Kumar Pramod

    2001-01-01

    Full Text Available A modified method of continuous loop technique for microvascular anastomosis is described. The handling of loop is easier & even last suture is placed under vision. This makes the microvascular anastomosis easier and simpler.

  4. In vitro inhibition of protease-activated receptors 1, 2 and 4 demonstrates that these receptors are not involved in an Acanthamoeba castellanii keratitis isolate-mediated disruption of the human brain microvascular endothelial cells.

    Science.gov (United States)

    Iqbal, Junaid; Naeem, Komal; Siddiqui, Ruqaiyyah; Khan, Naveed Ahmed

    2014-11-01

    Granulomatous amoebic encephalitis is a rare but serious human disease leading almost always to death. The pathophysiology of amoebic encephalitis is better understood, while events leading to the constitution of brain infection are largely unknown. Traversal of the blood-brain barrier is a key step in amoebae invasion of the central nervous system and facilitated by amoebic extracellular proteases. By using specific inhibitors of protease-activated receptors 1, 2 and 4, here we studied the role of these host receptors in Acanthamoeba castellanii-mediated damage to human brain microvasculature endothelial cells (HBMEC), which constitute the blood-brain barrier. The primary HBMEC were incubated with A. castellanii-conditioned medium in the presence or absence of FR-171113 (selective inhibitor of protease-activated receptor 1), FSLLRY-NH2 (inhibitor of protease-activated receptor 2), and tcY-NH2 (inhibitor of protease-activated receptor 4). The HBMEC monolayer disruptions were assessed by microscopy using Eosin staining, while host cell cytotoxicity was determined by measuring the release of cytoplasmic lactate dehydrogenase. Zymographic assays were performed to determine the effects of inhibitors of protease-activated receptors on the extracellular proteolytic activities of A. castellanii. A. castellanii-conditioned medium produced severe HBMEC monolayer disruptions within 60 min. The selective inhibitors of protease-activated receptors tested did not affect HBMEC monolayer disruptions. On the contrary, pre-treatment of A. castellanii-conditioned medium with phenylmethylsulfonyl fluoride, a serine protease inhibitor, or heating for 10 min at 95°C abolished HBMEC monolayer disruptions. Additionally, inhibitors of protease-activated receptors tested, failed to block A. castellanii-mediated HBMEC cytotoxicity and did not affect extracellular proteolytic activities of A. castellanii. Protease-activated receptors 1, 2 and 4 do not appear to play a role in A. castellanii

  5. Contribution of endothelial precursors of adipose tissue to breast cancer: progression-link with fat graft for reconstructive surgery.

    Science.gov (United States)

    Bertolini, Francesco

    2013-05-01

    Obesity, an excess accumulation of adipose tissue occurring in mammalians when caloric intake exceeds energy expenditure, is associated with an increased frequency and progression of several types of neoplastic diseases including postmenopausal breast cancer. Recent studies have suggested that obesity-related disruption of the energy homeostasis results in inflammation and alterations of adipokine signalling that may foster cancer initiation and progression. Moreover, two populations of human white adipose tissue (WAT) progenitors cooperate in breast cancer angiogenesis, growth and metastatic progression. This raises the issue of lipotransfer in patients undergoing plastic or reconstructive surgery.

  6. Influence of RhoA-Rock pathway inhibitor on the filamentous actin of hypoxia human pulmonary microvascular endothelial cells%RhoA-Rock信号通路抑制剂对缺氧人肺微血管内皮细胞丝状肌动蛋白细胞骨架的影响

    Institute of Scientific and Technical Information of China (English)

    张费通; 崔其亮; 莫镜

    2012-01-01

    Objective To explore the control factor of pulmonary microvascular endothelial cells in pulmonary hemorrhage with the RhoA-Rock pathway inhibitors.Methods Human pulmonary rnicrovascular endothelial cells were conventionally cultured,and were divided into four groups:control group,inhibitor group,hypoxia group and hypoxia group with inhibitor.As different fluorescein lsothiocyanate-phalloidin and filamentous actin (F-actin) in cytoplasm combined,it issued red fluorescence.We observed the dynamic changes of F-actin by laser scanning confocal microscope in hypoxia human pulmonary microvascular endothelial cells and recorded the value of fluorescence.Results The mean fluorescence intensity of F-actin of hypoxia group in 1 h,12h and 24 h was (64.3 ±5.5)%,(60.3±4.2)%,and (47.8 ±4.6)% as compared with the control group;the ratio of hypoxia group with inhibitor was (66.2 ±3.2)%,(67.1 ±6.2)%,and (72.5 ± 6.1 ) % as compared with the control group.The mean fluorescence intensity of F-actin decreased obviously after 1 h hypoxia treated to cells,decreasing to (64.3 ± 5.5 ) % of the control group (P <0.05 ) ;to 24 h,decreasing to (47.8 ±4.6) % of the control group(P <0.05).The mean fluorescence intensity of F-actin decreased to (66.2 ± 3.2) % of the control group after 1 h hypoxia treated in inhibitor group,which was more than 1 h in the hypoxic group.F-actin decreased obviously to (72.5 ± 6.1 ) % of the control group after 24 h hypoxia treated in inhibitor group.There was significant difference comparing with the hypoxia group after 24 h hypoxia(P <0.05).The mean fluorescence intensity of F-actin of inhibitor group without anoxic was invariant comparing with the control group(P >0.05).Cortex-like structure disappeared and the stress fibers arranged disorderly after hypoxia.Actin depolymedzated and broke gradually with the extension of hypoxia time.If to be hypoxic after pretreated with RhoA-Rock pathway inhibitor,cortex-like structure by

  7. Tumor microvascular changes in antiangiogenic treatment : Assessment by magnetic resonance contrast media of different molecular weights

    NARCIS (Netherlands)

    Turetschek, K; Preda, A; Novikov, [No Value; Brasch, RC; Weinmann, HJ; Wunderbaldinger, P; Roberts, TPL

    2004-01-01

    Purpose: To test magnetic resonance (MR) contrast media of different molecular weights (MWs) for their potential to characterize noninvasively microvascular changes in an experimental tumor treatment model. Materials and Methods: MD-MBA-435, a poorly differentiated human breast cancer cell line, was

  8. Lymphatic Vessel Function and Lymphatic Growth Factor Secretion after Microvascular Lymph Node Transfer in Lymphedema Patients

    Directory of Open Access Journals (Sweden)

    Tiina P. Viitanen, MD

    2013-05-01

    Conclusions: Reconstructing the lymphatic anatomy of the axilla with a lymph node flap may offer possibilities that other reconstructive options are lacking. However, we will need further reports and comparative studies about the clinical efficacy of this new promising technique. In addition to the transferred lymph nodes, lymphatic growth factor production may also be induced by other factors related to microvascular breast reconstruction.

  9. Microvascularization on collared peccary placenta

    DEFF Research Database (Denmark)

    Santos, Tatiana Carlesso; Oliveira, Moacir Franco; Dantzer, Vibeke

    2012-01-01

    into a microvascular network wall in a basket-like fashion. At the base of these baskets venules were formed. On the fetal side, arterioles branched centrally in the fetal rugae into a capillary network in a bulbous form, complementary to the opposite maternal depressions forming the baskets. At the base...

  10. Microvascular decompression for trigeminal neuralgia.

    Science.gov (United States)

    Sade, Burak; Lee, Joung H

    2014-10-01

    The microvascular decompression procedure has proven to be a safe and effective option in the surgical management of neurovascular compression syndromes in general and trigeminal neuralgia in particular. This article aims to serve as an overview of the decision-making process, application of the surgical technique, and clinical outcome pertaining to this procedure.

  11. Microvascular Disease After Renal Transplantation

    Directory of Open Access Journals (Sweden)

    Qi Lun Ooi

    2015-11-01

    Full Text Available Background/Aims: Individuals who reach end-stage kidney disease (CKD5 have a high risk of vascular events that persists even after renal transplantation. This study compared the prevalence and severity of microvascular disease in transplant recipients and patients with CKD5. Methods: Individuals with a renal transplant or CKD5 were recruited consecutively from renal clinics, and underwent bilateral retinal photography (Canon CR5-45, Canon. Their retinal images were deidentified and reviewed for hypertensive/microvascular signs by an ophthalmologist and a trained grader (Wong and Mitchell classification, and for vessel caliber at a grading centre using a computer-assisted method and Knudtson's modification of the Parr-Hubbard formula. Results: Ninety-two transplant recipients (median duration 6.4 years, range 0.8 to 28.8 and 70 subjects with CKD5 were studied. Transplant recipients were younger (pConclusions: Hypertensive/microvascular disease occurred just as often and was generally as severe in transplant recipients and subjects with CKD5. Microvascular disease potentially contributes to increased cardiac events post- transplantation.

  12. Prognostic impact of placenta growth factor and vascular endothelial growth factor A in patients with breast cancer

    DEFF Research Database (Denmark)

    Maae, Else; Olsen, Dorte Aalund; Dahl Steffensen, Karina

    2012-01-01

    recurrence-free survival (RFS) in both univariate analysis (PlGF: p = 0.023; VEGF-A: p = 0.047) and in multivariate analysis (PlGF: p = 0.026; VEGF-A: p = 0.036) when entered in the model separately, but neither PlGF (p = 0.089) nor VEGF-A (p = 0.127) turned out as independent prognostic variables, when...... entered in the model together. Neither PlGF nor VEGF-A expression in non-malignant tissue were predictors for RFS. Conclusion: High protein levels of PlGF and VEGF-A seem to be associated with adverse prognosis in breast cancer patients. Our results support the mutual relationship between PlGF and VEGF...... were measured in 229 tumor tissue specimen from primarily operated patients with unilateral breast cancer. Non-malignant breast tissue was also dissected near the tumor and quantitative measurements were available for 211 patients. PlGF and VEGF-A protein levels in homogenized tissue lysates were...

  13. Stable transfection of estrogen receptor-alpha suppresses expression of cyclooxygenase-2 and vascular endothelial growth factor-C in MDA-MB-231 breast cancer cells

    Institute of Scientific and Technical Information of China (English)

    ZHANG Hui; LIN Ying; XIAO Ying; WANG San-ming; LIU Xiang-xia; WANG Shen-ming

    2010-01-01

    Background Estrogen receptor (ER)-negative breast cancer cells are more aggressive than ER-positive cells. Elevated levels of cyclooxygenase-2 (COX-2) and vascular endothelial growth factor-C (VEGF-C) expression have been detected in cultured human breast cancer cells and are associated with negative hormone receptor status. In this study, we created ERα stable transfectants in MDA-MB-231 cells to explore the effect of ERα on cell growth and COX-2 and VEGF-C expression.Methods The green fluorescent protein (GFP)-ERα plasmids were stably transfected into ER-negative MDA-MB-231 cells. The proliferation and migration of untransfected MDA-MB-231 cells, ERα-transfected MDA-MB-231 cells and ER-positive MCF-7 cells were determined. The expression of COX-2, and the levels of VEGF-C mRNA and the VEGF-C secretion concentration were assayed in these cell lines.Results The proliferation and migration capacities of ERα-tranfected MDA-MB-231 cells were significantly decreased (P <0.05). The expression of COX-2 was significantly lower in ERa-tranfected MDA-MB-231 cells than in untranfected MDA-MB-231 cells. The mRNA and protein levels of VEGF-C were lower in ERa-tranfected MDA-MB-231 cells than in untransfected MDA-MB-231 cells (P<0.05).Conclusions ERα stable transfection inhibits proliferation and migration capacities of MDA-MB-231 cells and decreases expression of COX-2 and VEGF-C. The decreases of proliferation and migration capacities may be related to suppression of COX-2 and VEGF-C expression.

  14. 丹皮酚对LLO诱导大鼠肠黏膜微血管内皮细胞分泌NO、ET-1的影响%Effects of Paeonol on the Secretion of NO and ET-1 in Rat Intestinal Mucosa Microvascular Endothelial Cells Induced by LLO

    Institute of Scientific and Technical Information of China (English)

    陈希; 吴启; 顾进华; 许剑琴; 穆祥

    2012-01-01

    To investigate the treatment mechanism of the paeonol (Pae) on the Hsteriosis and the secretion of NO and ET-1 in rat intestinal mucosa microvascular endothelial cells (RIMVECs) induced by listeriolysin O (LLO). RIMVECs were in culture divided into LLO + Pae, LLO, Pae and blank control group. Cells were assessed proliferation by MTT assay. The Changes of NO and ET-1 level were measured by the method of nitratase, ELISA and hybridization in situ. Compared with other groups, proliferation of LLO was very significantly decreased. The production of NO and ET-1 were enhanced for 12 h, and NO/ET-1 ratio was lower. Pae could regulate imbalance of NO/ET-1 induced by LLO and improve the local microcirculation of intestine.%为了研究李斯特菌溶血素(LLO)对大鼠肠黏膜微血管内皮细胞(RIMVECs)分泌一氧化氮(NO)、内皮素-1(ET-1)的影响,阐释作为治疗李斯特菌病方剂中单味中药丹皮的有效成分丹皮酚(Pae)调节LLO诱导细胞分泌紊乱的作用机理.试验将培养的RIMVECs分为LLO+Pae组、LLO组、Pae组、空白组,通过MTT比色法测定细胞生长情况;硝酸还原酶法测定细胞上清波细胞因子NO浓度以及酶联免疫法检测ET-1浓度;并用原位杂交方法对结果进行验证;结果显示LLO组与其他各组相比:细胞增殖显著性下降;12小时内,NO、ET-1分泌量高于正常水平,并导致NO/ET-1比值失衡(下降);而LLO+Pae组NO/ET-1的比值显著上调;由此可见,Pae可以通过改善LLO引起细胞因子NO、ET-1失衡,有效调节细胞微环境,部分解释了中药丹皮治疗李斯特菌病的作用机理.

  15. 肿瘤坏死因子致肺微血管内皮细胞单层通 透性损伤的实验研究%EXPERIMENTAL RESEARCH ON PERMEABILITY INJURY OF PULMONARY MICROVASCULAR ENDOTHELIAL CELLS MONOLAYER INDUCED BY TUMOR NECROSIS FACTOR

    Institute of Scientific and Technical Information of China (English)

    肖贞良; 孙耕耘; 夏前明; 钱桂生

    2001-01-01

    目的和方法: 用针头式滤器检测肿瘤环死因子(TNF)作用前后及三 种药物干预时大鼠肺微血管内皮细胞(RPMVEC)单层通透性的变化,并用免疫组化的方法检测 TNF作用前后细胞F-肌动蛋白的改变。结果:TNF作用30 min、60min、90 min通透系数Kf值较致伤前显著增高;分别加福莫特罗(FOR)、山莨菪碱或霍乱毒素(CTX)干 预时Kf值均显著低于TNF组。而TNF作用90 min,RPMVEC F-肌动蛋白发生明显解聚:分别加 POR、山莨菪硷或CTX干预时F-肌动蛋白无明显变化。结论:TNF诱导RPMVEC 单层通透性增高的机制与细胞F-肌动蛋白解聚有关,FOR、山莨菪碱和CTX可能通过抑制F- 肌动蛋白解聚而抑制TNF诱导的RPMVEC单层通透性增高。%Methods:To investigate the mechanism of tumor necrosis factor( TNF) induced permeability injury of rat pulmonary microvascular endothelial cel ls (RPMVEC) monolayer, the effect of TNF on permeability of RPMVEC monolayer was examined with microfilter and the effect of TNF on RPMVEC F-actin was observed with immunocytochemistry. The interferring action of formoterol, anisodamine an d cholera toxin on permeability and F-actin changes induced by TNF was also obs erved.Results:①TNF induced significant increase in permeability of RPMVEC monolayer 30, 60 and90 minutes after treatment with TNF.②F-actin in RPMVEC depolymerized 90 minutes after treatment with TNF.Permeability and F-ac tin did not change significantly when formoterol, anisodamine or cholera toxin w as added separately. Conclusion: TNF can induce permeability inj ury of RPMVEC monolayer,which is correlated with depolymerization of F-actin. F ormoterol, anisodamine and cholera toxin can inhibit the permeability change ind uced by TNF which may due to their inhibition to the distribution change of F-a ctin.

  16. Nab-paclitaxel Efficacy in the Orthotopic Model of Human Breast Cancer Is Significantly Enhanced By Concurrent Anti–Vascular Endothelial Growth Factor A Therapy

    Directory of Open Access Journals (Sweden)

    Lisa D. Volk

    2008-06-01

    Full Text Available Nab-paclitaxel is an albumin-bound 130-nm particle form of paclitaxel that has shown an improved efficacy in experimental tumor models and clinical studies compared with solvent-based paclitaxel. Anti–vascular endothelial growth factor A (VEGF-A antibody bevacizumab is known to enhance antitumor activity of cytotoxic drugs. This study evaluated the effects of combined nab-paclitaxel and bevacizumab therapy on growth and metastatic spread of orthotopic breast tumors. Cytotoxic and clonogenic assays measured VEGF-A–dependent modulation of nabpaclitaxel toxicity on cultured tumor cells. Antitumor effects were assessed in mice with luciferase-tagged, wellestablished MDA-MB-231 tumors (250–310 mm3 treated with one, two, or three cycles of nab-paclitaxel (10 mg/ kg, daily for five consecutive days, bevacizumab (2–8 mg/kg, twice a week, or with combination of both drugs. VEGF-A protected MDA-MB-231 cells against nab-paclitaxel cytotoxicity, whereas bevacizumab sensitized cells to the effect of the drug. Combined bevacizumab and nab-paclitaxel treatment synergistically inhibited tumor growth and metastasis resulting in up to 40% of complete regressions of well-established tumors. This therapy also decreased the incidence of lymphatic and pulmonary metastases by 60% and 100%, respectively. The significant increase in the cure of tumor-bearing mice in the nab-paclitaxel/bevacizumab combined group compared with mice treated with single drugs strongly advocates for implementing such strategy in clinics.

  17. The prevention of diabetic microvascular complications of diabetes: is there a role for lipid lowering?

    Science.gov (United States)

    Leiter, Lawrence A

    2005-06-01

    The role of hyperglycemia in the development of microvascular complications of diabetes, such as nephropathy, retinopathy and neuropathy, has been well documented. Evidence is accumulating to support the concept that dyslipidemia can also contribute to the development of these complications. Lipid-lowering agents, such as statins, have been shown to prevent cardiovascular events in patients with diabetes. However, in addition to preventing macrovascular diseases, statins may also be able to retard the progression of microvascular complications of diabetes. Indeed, in addition to reducing lipid levels, these agents can improve endothelial function and reduce oxidative stress, which can improve microvascular function. These findings would provide further support for the use of lipid-lowering agents in patients with diabetes.

  18. Brain endothelial dysfunction in cerebral adrenoleukodystrophy.

    Science.gov (United States)

    Musolino, Patricia L; Gong, Yi; Snyder, Juliet M T; Jimenez, Sandra; Lok, Josephine; Lo, Eng H; Moser, Ann B; Grabowski, Eric F; Frosch, Matthew P; Eichler, Florian S

    2015-11-01

    See Aubourg (doi:10.1093/awv271) for a scientific commentary on this article.X-linked adrenoleukodystrophy is caused by mutations in the ABCD1 gene leading to accumulation of very long chain fatty acids. Its most severe neurological manifestation is cerebral adrenoleukodystrophy. Here we demonstrate that progressive inflammatory demyelination in cerebral adrenoleukodystrophy coincides with blood-brain barrier dysfunction, increased MMP9 expression, and changes in endothelial tight junction proteins as well as adhesion molecules. ABCD1, but not its closest homologue ABCD2, is highly expressed in human brain microvascular endothelial cells, far exceeding its expression in the systemic vasculature. Silencing of ABCD1 in human brain microvascular endothelial cells causes accumulation of very long chain fatty acids, but much later than the immediate upregulation of adhesion molecules and decrease in tight junction proteins. This results in greater adhesion and transmigration of monocytes across the endothelium. PCR-array screening of human brain microvascular endothelial cells after ABCD1 silencing revealed downregulation of both mRNA and protein levels of the transcription factor c-MYC (encoded by MYC). Interestingly, MYC silencing mimicked the effects of ABCD1 silencing on CLDN5 and ICAM1 without decreasing the levels of ABCD1 protein itself. Together, these data demonstrate that ABCD1 deficiency induces significant alterations in brain endothelium via c-MYC and may thereby contribute to the increased trafficking of leucocytes across the blood-brain barrier as seen in cerebral adrenouleukodystrophy.

  19. Cancer-associated fibroblast promote transmigration through endothelial brain cells in three-dimensional in vitro models.

    Science.gov (United States)

    Choi, Yoon Pyo; Lee, Joo Hyun; Gao, Ming-Qing; Kim, Baek Gil; Kang, Suki; Kim, Se Hoon; Cho, Nam Hoon

    2014-11-01

    Brain metastases are associated with high morbidity as well as with poor prognosis and survival in breast cancer patients. Despite its clinical importance, metastasis of breast cancer cells through the blood-brain barrier (BBB) is poorly understood. The objective of our study was to investigate whether cancer-associated fibroblasts (CAFs) play crucial roles in breast cancer brain metastasis. Using a cell adhesion assays, in vitro BBB permeability and transmigration assays and soft agar colony formation assays, we investigated the physical roles of CAFs in breast cancer brain metastasis. We also performed immunofluorescence, flow cytometric analysis, Droplet Digital PCR and Simon™ Simple Western System to confirm changes in expression levels. We established two novel three-dimensional (3D) culture systems using a perpendicular slide chamber and applying 3D embedded culture method to reflect brain metastasis conditions. With a newly developed device, CAFs was proven to promote cell adhesion to human brain microvascular endothelial cells, in vitro BBB permeability and transmigration and colony formation of breast cancer cells. Furthermore, CAFs enhanced the invasive migration of breast cancer cells in two kinds of 3D cultures. These 3D models also reliably recapitulate the initial steps of BBB transmigration, micro-metastasis and colonization. Expression of integrin α5β1 and αvβ3, c-MET and α2,6-siayltransferase was increased in breast cancer cells that migrated through the BBB. In conclusion, based on our in vitro BBB and co-culture models, our data suggest that CAFs may play a role in breast cancer brain metastasis.

  20. Inflammation-induced microvascular insulin resistance is an early event in diet-induced obesity.

    Science.gov (United States)

    Zhao, Lina; Fu, Zhuo; Wu, Jing; Aylor, Kevin W; Barrett, Eugene J; Cao, Wenhong; Liu, Zhenqi

    2015-12-01

    Endothelial dysfunction and vascular insulin resistance usually coexist and chronic inflammation engenders both. In the present study, we investigate the temporal relationship between vascular insulin resistance and metabolic insulin resistance. We assessed insulin responses in all arterial segments, including aorta, distal saphenous artery and the microvasculature, as well as the metabolic insulin responses in muscle in rats fed on a high-fat diet (HFD) for various durations ranging from 3 days to 4 weeks with or without sodium salicylate treatment. Compared with controls, HFD feeding significantly blunted insulin-mediated Akt (protein kinase B) and eNOS [endothelial nitric oxide (NO) synthase] phosphorylation in aorta in 1 week, blunted vasodilatory response in small resistance vessel in 4 weeks and microvascular recruitment in as early as 3 days. Insulin-stimulated whole body glucose disposal did not begin to progressively decrease until after 1 week. Salicylate treatment fully inhibited vascular inflammation, prevented microvascular insulin resistance and significantly improved muscle metabolic responses to insulin. We conclude that microvascular insulin resistance is an early event in diet-induced obesity and insulin resistance and inflammation plays an essential role in this process. Our data suggest microvascular insulin resistance contributes to the development of metabolic insulin resistance in muscle and muscle microvasculature is a potential therapeutic target in the prevention and treatment of diabetes and its related complications.

  1. The role of calcineurin/NFAT in SFRP2 induced angiogenesis--a rationale for breast cancer treatment with the calcineurin inhibitor tacrolimus.

    Directory of Open Access Journals (Sweden)

    Sharareh Siamakpour-Reihani

    Full Text Available Tacrolimus (FK506 is an immunosuppressive drug that binds to the immunophilin FKBPB12. The FK506-FKBP12 complex associates with calcineurin and inhibits its phosphatase activity, resulting in inhibition of nuclear translocation of nuclear factor of activated T-cells (NFAT. There is increasing data supporting a critical role of NFAT in mediating angiogenic responses stimulated by both vascular endothelial growth factor (VEGF and a novel angiogenesis factor, secreted frizzled-related protein 2 (SFRP2. Since both VEGF and SFRP2 are expressed in breast carcinomas, we hypothesized that tacrolimus would inhibit breast carcinoma growth. Using IHC (IHC with antibodies to FKBP12 on breast carcinomas we found that FKBP12 localizes to breast tumor vasculature. Treatment of MMTV-neu transgenic mice with tacrolimus (3 mg/kg i.p. daily (n = 19 resulted in a 73% reduction in the growth rate for tacrolimus treated mice compared to control (n = 15, p = 0.003; which was associated with an 82% reduction in tumor microvascular density (p<0.001 by IHC. Tacrolimus (1 µM inhibited SFRP2 induced endothelial tube formation by 71% (p = 0.005 and inhibited VEGF induced endothelial tube formation by 67% (p = 0.004. To show that NFATc3 is required for SFRP2 stimulated angiogenesis, NFATc3 was silenced with shRNA in endothelial cells. Sham transfected cells responded to SFRP2 stimulation in a tube formation assay with an increase in the number of branch points (p<0.003, however, cells transfected with shRNA to NFATc3 showed no increase in tube formation in response to SFRP2. This demonstrates that NFATc3 is required for SFRP2 induced tube formation, and tacrolimus inhibits angiogenesis in vitro and breast carcinoma growth in vivo. This provides a rationale for examining the therapeutic potential of tacrolimus at inhibiting breast carcinoma growth in humans.

  2. Mst1 inhibits CMECs autophagy and participates in the development of diabetic coronary microvascular dysfunction

    Science.gov (United States)

    Lin, Jie; Zhang, Lei; Zhang, Mingming; Hu, Jianqiang; Wang, Tingting; Duan, Yu; Man, Wanrong; Wu, Bin; Feng, Jiaxu; Sun, Lei; Li, Congye; Zhang, Rongqing; Wang, Haichang; Sun, Dongdong

    2016-01-01

    Cardiovascular complications account for a substantial proportion of morbidity and mortality in diabetic patients. Abnormalities of cardiac microvascular endothelial cells (CMECs) lead to impaired cardiac microvascular vessel integrity and subsequent cardiac dysfunction, underlining the importance of coronary microvascular dysfunction. In this study, experimental diabetes models were constructed using Mst1 transgenic, Mst1 knockout and sirt1 knockout mice. Diabetic Mst1 transgenic mice exhibited impaired cardiac microvessel integrity and decreased cardiac function. Mst1 overexpression deceased CMECs autophagy as evidenced by decreased LC3 expression and enhanced protein aggregation when subjected to high glucose culture. Mst1 knockout improved cardiac microvessel integrity and enhanced cardiac functions in diabetic mice. Mst1 knockdown up-regulated autophagy as indicated by more typical autophagosomes and increased LC3 expression in CMECs subjected to high glucose cultures. Mst1 knockdown also promoted autophagic flux in the presence of bafilomycin A1. Mst1 overexpression increased CMECs apoptosis, whereas Mst1 knockout decreased CMECs apoptosis. Sirt1 knockout abolished the effects of Mst1 overexpression in cardiac microvascular injury and cardiac dysfunction. In conclusion, Mst1 knockout preserved cardiac microvessel integrity and improved cardiac functions in diabetic mice. Mst1 decreased sirt1 activity, inhibited autophagy and enhanced apoptosis in CMECs, thus participating in the pathogenesis of diabetic coronary microvascular dysfunction. PMID:27680548

  3. Ascorbic Acid Efflux from Human Brain Microvascular Pericytes: Role of Re-uptake

    OpenAIRE

    May, James M.; Qu, Zhi-chao

    2015-01-01

    Microvascular pericytes take up ascorbic acid on the ascorbate transporter SVCT2. Intracellular ascorbate then protects the cells against apoptosis induced by culture at diabetic glucose concentrations. To investigate whether pericytes might also provide ascorbate to the underlying endothelial cells, we studied ascorbate efflux from human pericytes. When loaded with ascorbate to intracellular concentrations of 0.8–1.0 mM, almost two-thirds of intracellular ascorbate effluxed from the cells ov...

  4. Rickettsiae induce microvascular hyperpermeability via phosphorylation of VE-cadherins: evidence from atomic force microscopy and biochemical studies.

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    Bin Gong

    Full Text Available The most prominent pathophysiological effect of spotted fever group (SFG rickettsial infection of microvascular endothelial cells (ECs is an enhanced vascular permeability, promoting vasogenic cerebral edema and non-cardiogenic pulmonary edema, which are responsible for most of the morbidity and mortality in severe cases. To date, the cellular and molecular mechanisms by which SFG Rickettsia increase EC permeability are largely unknown. In the present study we used atomic force microscopy (AFM to study the interactive forces between vascular endothelial (VE-cadherin and human cerebral microvascular EC infected with R. montanensis, which is genetically similar to R. rickettsii and R. conorii, and displays a similar ability to invade cells, but is non-pathogenic and can be experimentally manipulated under Biosafety Level 2 (BSL2 conditions. We found that infected ECs show a significant decrease in VE-cadherin-EC interactions. In addition, we applied immunofluorescent staining, immunoprecipitation phosphorylation assay, and an in vitro endothelial permeability assay to study the biochemical mechanisms that may participate in the enhanced vascular permeability as an underlying pathologic alteration of SFG rickettsial infection. A major finding is that infection of R. montanensis significantly activated tyrosine phosphorylation of VE-cadherin beginning at 48 hr and reaching a peak at 72 hr p.i. In vitro permeability assay showed an enhanced microvascular permeability at 72 hr p.i. On the other hand, AFM experiments showed a dramatic reduction in VE-cadherin-EC interactive forces at 48 hr p.i. We conclude that upon infection by SFG rickettsiae, phosphorylation of VE-cadherin directly attenuates homophilic protein-protein interactions at the endothelial adherens junctions, and may lead to endothelial paracellular barrier dysfunction causing microvascular hyperpermeability. These new approaches should prove useful in characterizing the antigenically

  5. Feasibility Study of EndoTAG-1, a Tumor Endothelial Targeting Agent, in Combination with Paclitaxel followed by FEC as Induction Therapy in HER2-Negative Breast Cancer.

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    Michail Ignatiadis

    Full Text Available EndoTAG-1, a tumor endothelial targeting agent has shown activity in metastatic triple-negative breast cancer (BC in combination with paclitaxel.HER2-negative BC patients candidates for neoadjuvant chemotherapy were scheduled to receive 12 cycles of weekly EndoTAG-1 22mg/m2 plus paclitaxel 70mg/m2 followed by 3 cycles of FEC (Fluorouracil 500mg/m2, Epirubicin 100mg/m2, Cyclophosphamide 500mg/m2 every 3 weeks followed by surgery. Primary endpoint was percent (% reduction in Magnetic Resonance Imaging (MRI estimated Gadolinium (Gd enhancing tumor volume at the end of EndoTAG-1 plus paclitaxel administration as compared to baseline. Safety, pathological complete response (pCR defined as no residual tumor in breast and axillary nodes at surgery and correlation between % reduction in MRI estimated tumor volume and pCR were also evaluated.Fifteen out of 20 scheduled patients were included: Six patients with estrogen receptor (ER-negative/HER2-negative and 9 with ER-positive/HER2-negative BC. Nine patients completed treatment as per protocol. Despite premedication and slow infusion rates, grade 3 hypersensitivity reactions to EndoTAG-1 were observed during the 1st, 2nd, 3rd and 6th weekly infusion in 4 patients, respectively, and required permanent discontinuation of the EndoTAG-1. Moreover, two additional patients stopped EndoTAG-1 plus paclitaxel after 8 and 9 weeks due to clinical disease progression. Two patients had grade 3 increases in transaminases and 1 patient grade 4 neutropenia. pCR was achieved in 5 of the 6 ER-/HER2- and in none of the 9 ER+/HER2- BC patients. The mean % reduction in MRI estimated tumor volume at the end of EndoTAG-1 plus paclitaxel treatment was 81% (95% CI, 66% to 96%, p<0.001 for the 15 patients that underwent surgery; 96% for patients with pCR and 73% for patients with no pCR (p = 0.04.The EndoTAG-1 and paclitaxel combination showed promising preliminary activity as preoperative treatment, especially in ER-/HER2

  6. A novel effective method for the assessment of microvascular function in male patients with coronary artery disease: a pilot study using laser speckle contrast imaging

    Energy Technology Data Exchange (ETDEWEB)

    Borges, J.P. [Laboratório de Atividade Física e Promoção è Saúde, Departamento de Desporto Coletivo, Instituto de Educação Física e Desportos, Universidade do Estado do Rio de Janeiro, Rio de Janeiro, RJ (Brazil); Lopes, G.O. [Laboratório de Atividade Física e Promoção è Saúde, Departamento de Desporto Coletivo, Instituto de Educação Física e Desportos, Universidade do Estado do Rio de Janeiro, Rio de Janeiro, RJ (Brazil); Instituto Nacional de Cardiologia, Rio de Janeiro, RJ (Brazil); Verri, V.; Coelho, M.P.; Nascimento, P.M.C.; Kopiler, D.A. [Instituto Nacional de Cardiologia, Rio de Janeiro, RJ (Brazil); Tibirica, E. [Instituto Nacional de Cardiologia, Rio de Janeiro, RJ (Brazil); Laboratório de Investigação Cardiovascular, Departamento Osório de Almeida, Instituto Oswaldo Cruz, FIOCRUZ, Rio de Janeiro, RJ (Brazil)

    2016-09-01

    Evaluation of microvascular endothelial function is essential for investigating the pathophysiology and treatment of cardiovascular and metabolic diseases. Although laser speckle contrast imaging technology is well accepted as a noninvasive methodology for assessing microvascular endothelial function, it has never been used to compare male patients with coronary artery disease with male age-matched healthy controls. Thus, the aim of this study was to determine whether laser speckle contrast imaging could be used to detect differences in the systemic microvascular functions of patients with established cardiovascular disease (n=61) and healthy age-matched subjects (n=24). Cutaneous blood flow was assessed in the skin of the forearm using laser speckle contrast imaging coupled with the transdermal iontophoretic delivery of acetylcholine and post-occlusive reactive hyperemia. The maximum increase in skin blood flow induced by acetylcholine was significantly reduced in the cardiovascular disease patients compared with the control subjects (74 vs 116%; P<0.01). With regard to post-occlusive reactive hyperemia-induced vasodilation, the patients also presented reduced responses compared to the controls (0.42±0.15 vs 0.50±0.13 APU/mmHg; P=0.04). In conclusion, laser speckle contrast imaging can identify endothelial and microvascular dysfunctions in male individuals with cardiovascular disease. Thus, this technology appears to be an efficient non-invasive technique for evaluating systemic microvascular and endothelial functions, which could be valuable as a peripheral marker of atherothrombotic diseases in men.

  7. A novel effective method for the assessment of microvascular function in male patients with coronary artery disease: a pilot study using laser speckle contrast imaging

    Directory of Open Access Journals (Sweden)

    J.P. Borges

    2016-01-01

    Full Text Available Evaluation of microvascular endothelial function is essential for investigating the pathophysiology and treatment of cardiovascular and metabolic diseases. Although laser speckle contrast imaging technology is well accepted as a noninvasive methodology for assessing microvascular endothelial function, it has never been used to compare male patients with coronary artery disease with male age-matched healthy controls. Thus, the aim of this study was to determine whether laser speckle contrast imaging could be used to detect differences in the systemic microvascular functions of patients with established cardiovascular disease (n=61 and healthy age-matched subjects (n=24. Cutaneous blood flow was assessed in the skin of the forearm using laser speckle contrast imaging coupled with the transdermal iontophoretic delivery of acetylcholine and post-occlusive reactive hyperemia. The maximum increase in skin blood flow induced by acetylcholine was significantly reduced in the cardiovascular disease patients compared with the control subjects (74 vs 116%; P<0.01. With regard to post-occlusive reactive hyperemia-induced vasodilation, the patients also presented reduced responses compared to the controls (0.42±0.15 vs 0.50±0.13 APU/mmHg; P=0.04. In conclusion, laser speckle contrast imaging can identify endothelial and microvascular dysfunctions in male individuals with cardiovascular disease. Thus, this technology appears to be an efficient non-invasive technique for evaluating systemic microvascular and endothelial functions, which could be valuable as a peripheral marker of atherothrombotic diseases in men.

  8. A novel effective method for the assessment of microvascular function in male patients with coronary artery disease: a pilot study using laser speckle contrast imaging.

    Science.gov (United States)

    Borges, J P; Lopes, G O; Verri, V; Coelho, M P; Nascimento, P M C; Kopiler, D A; Tibirica, E

    2016-09-01

    Evaluation of microvascular endothelial function is essential for investigating the pathophysiology and treatment of cardiovascular and metabolic diseases. Although laser speckle contrast imaging technology is well accepted as a noninvasive methodology for assessing microvascular endothelial function, it has never been used to compare male patients with coronary artery disease with male age-matched healthy controls. Thus, the aim of this study was to determine whether laser speckle contrast imaging could be used to detect differences in the systemic microvascular functions of patients with established cardiovascular disease (n=61) and healthy age-matched subjects (n=24). Cutaneous blood flow was assessed in the skin of the forearm using laser speckle contrast imaging coupled with the transdermal iontophoretic delivery of acetylcholine and post-occlusive reactive hyperemia. The maximum increase in skin blood flow induced by acetylcholine was significantly reduced in the cardiovascular disease patients compared with the control subjects (74 vs 116%; PAPU/mmHg; P=0.04). In conclusion, laser speckle contrast imaging can identify endothelial and microvascular dysfunctions in male individuals with cardiovascular disease. Thus, this technology appears to be an efficient non-invasive technique for evaluating systemic microvascular and endothelial functions, which could be valuable as a peripheral marker of atherothrombotic diseases in men.

  9. The effect of exercise training on the level of tissue IL-6 and vascular endothelial growth factor in breast cancer bearing mice

    Directory of Open Access Journals (Sweden)

    Sadegh Amani Shalamzari

    2014-04-01

    Full Text Available Objective(s: The goal of this study was assessing the prophylactic effect of exercise and its role as an adjuvant therapy on level of cytokines involved in angiogenesis in estrogen-dependent breast cancer. Materials and Methods: Forty female BALB/c mice were randomly assigned to exercise-tumor-exercise (ETE, exercise-tumor-rest (ETR, rest-tumor-exercise (RTE and rest-tumor-rest (RTR groups. After orientation in the environment, two groups of mice performed continuous endurance exercise for 8 weeks, and thereafter estrogen-dependent MC4L2 cancer cells were injected to them. Then, one group of each of trained and non-trained mice performed endurance exercise 5 days per week for 6 weeks. Tumor volume was measured by a digital caliper weekly. Finally, the mice were sacrificed; tumor tissue was removed, immediately frozen and kept in              -70°C. Tumor sample was homogenized; levels of cytokines were measured and quantified using ELISA. Results: There was significant reduction in the level of interlukin-6 (IL-6 (P=0.001, Vascular endothelial growth factor (VEGF (P=0.0001 and tumor volume (P=0.0001 among the groups performing endurance exercise after malignancy (RTE and ETE in comparison with groups not performing endurance exercise (ETR and RTR, and these results were in agreement with tumor growth rate. Conclusion: Exercise can cause reduction in levels of pro-inflammatory cytokines in tumor tissue. Decreased IL-6 production could reduce the generation of VEGF, resulting in reduced intra-tumor angiogenesis. Due to reduction of the level of these cytokines in groups doing exercise before and after malignancy, exercise is presumed to be an adjuvant therapy in estrogen-receptor dependent tumors in addition to its effective prophylactic role.

  10. Effect of HIV-1gp41 ectodomain on Cryptococcus neoformans-induced cytoskeletal changes in human brain microvascular endothelial cells%HIV-1gp41胞外域对新生隐球菌致人脑微血管内皮细胞骨架改变的影响

    Institute of Scientific and Technical Information of China (English)

    龙敏; 曹虹; Ambrose Jong

    2011-01-01

    To study the effect of HIV-1 gp41 ectodomain (gp41-I90) on the cytoskeletal changes in human brain microvascular endothelial cells (HBMECs) induced by Cryptococcus neoformans. Methods HBMECs were cultured on collagen-coated chamber slide or transwell to allow the formation of cell monolayers. After pre-treatment with gp41-I90 and infection with Cryptococcus neoformans, the HBMECs were examined for the expression of actin or filamin by immunofluorescence assay. HRP permeability of the HBMECs treated with gp41-I90 was detected by ELISA. Transcytosis of Cryptococcus neoformans through the gp41-I90-treated HBMECs was detected by direct counting from a hemocytometer. Results gp41-I90 obviously enhanced the cytoskeletal changes of the HBMECs infected by Cryptococcus neoformans, causing curved and sparse filamentous arrangement of actin and filamin. Gp41-I90 treatment also resulted in obviously increased HRP permeability of the cells and transcytosis of Cryptococcus neoformans. Conclusion gp41-190 enhances Cryptococcus neoformans binding to HBMECs, which is related to its effect in enhancing Cryptococcus neoformans-induced cytoskeletal changes of the cells.%目的 探讨HIV-1 gp41胞外域gp41-I90肽对新生隐球菌致人脑微血管内皮细胞骨架改变的影响.方法 以人脑微血管内皮细胞作为体外血脑屏障模型,在细胞培养玻片或transwell小室上长成单层,用HIV-1 gp41-I90预处理后,再用新生隐球菌感染细胞单层,免疫荧光方法检测细胞骨架蛋白actin(肌动蛋白)和filamin(肌动蛋白连接蛋白)的形态变化,并测定gp41-I90处理的人脑微血管内皮细胞单层对辣根过氧化物酶和新生隐球菌的通过率.结果 gp41-I90能明显增强新生隐球菌所致的人脑微血管内皮细胞骨架蛋白actin和filamin的改变,actin和filamin的丝状排列扭曲,纹理稀疏;gp41-I90处理的人脑微血管内皮细胞单层通透性增强,对辣根过氧化物酶和新生隐球菌的

  11. Aminguanidine suppresses methylglyoxal-mediated oxygen-glucose deprivation injury in human brain microvascular endothelial cells%氨基胍抑制丙酮醛介导的脑微血管内皮细胞缺糖缺氧损伤

    Institute of Scientific and Technical Information of China (English)

    李汶潞; 胡全; 任夏; 何萍; 除慧敏; 戴海斌; 陈忠

    2013-01-01

    Objective:To evaluate the effects of aminguanidine on methylglyoxal-mediated oxygenglucose deprivation (OGD) injury in the cultured human brain microvascular endothelial cells (HBMEC).Methods:Cultured HBMEC cells were pretreated with methylglyoxal before oxygen-glucose deprivation injury.Cell vitality was determined by MTT method,cell mortality was assessed by LDH release method,cell apoptosis was examined by AnnexinV/PI formation method,and the advanced glycation end products (AGEs) were detected by Western-blot.Results:Methylglyoxal induced HBMEC injury in a dose-dependent manner.At 2 mmol/L of methylglyoxa,the cell viability was 56.1% ; when methylglyoxa-pretreated cells exposed to oxygen-glucose deprivation,the cell inhibition rate was 90.0%.Aminguanidine (1 mmol/L) inhibited methylglyoxa and OGD induced LDH release and AnnexinV/PI formation.Furthermore,aminguanidine (1 mmol/L) also decreased advanced glycation end products (AGEs) formation induced by methylglyoxal and oxygen-glucose deprivation.Conclusion:Aminguanidine protected methylglyoxal mediated-oxygen-glucose deprivation injury in the cultured HBMEC,which may be associated with anti-glycation activity.%目的:研究氨基胍对丙酮醛加重脑微血管内皮细胞(HBMEC)缺糖缺氧损伤的保护作用.方法:在培养的HBMEC上,利用丙酮醛加重缺糖缺氧诱导的损伤,通过MTT检测细胞活力,LDH释放检测细胞死亡,流式细胞仪检测细胞凋亡,Western-blot检测晚期糖基化终产物的形成,观察氨基胍的作用和机制.结果:丙酮醛呈浓度依赖地诱导细胞损伤,在2 mmol/L时细胞的存活率为56.1%,而丙酮醛合并缺糖缺氧后,细胞的损伤率增加到90.0%.氨基胍(l mmol/L)能抑制丙酮醛和缺糖缺氧诱导的LDH释放和AnnexinV/PI的形成.进一步研究发现氨基胍能抑制丙酮醛和缺糖缺氧诱导晚期糖基化终产物的形成.结论:氨基胍对丙酮醛加重HBMEC的缺糖缺氧损伤具有保护作用,这可能与其抗糖基化作用有关.

  12. Rab5-mediated VE-cadherin internalization regulates the barrier function of the lung microvascular endothelium.

    Science.gov (United States)

    Yang, Junjun; Yao, Wei; Qian, Guisheng; Wei, Zhenghua; Wu, Guangyu; Wang, Guansong

    2015-12-01

    The small GTPase Rab5 has been well defined to control the vesicle-mediated plasma membrane protein transport to the endosomal compartment. However, its function in the internalization of vascular endothelial (VE)-cadherin, an important component of adherens junctions, and as a result regulating the endothelial cell polarity and barrier function remain unknown. Here, we demonstrated that lipopolysaccharide (LPS) simulation markedly enhanced the activation and expression of Rab5 in human pulmonary microvascular endothelial cells (HPMECs), which is accompanied by VE-cadherin internalization. In parallel, LPS challenge also induced abnormal cell polarity and dysfunction of the endothelial barrier in HPMECs. LPS stimulation promoted the translocation of VE-cadherin from the plasma membrane to intracellular compartments, and intracellularly expressed VE-cadherin was extensively colocalized with Rab5. Small interfering RNA (siRNA)-mediated depletion of Rab5a expression attenuated the disruption of LPS-induced internalization of VE-cadherin and the disorder of cell polarity. Furthermore, knockdown of Rab5 inhibited the vascular endothelial hyperpermeability and protected endothelial barrier function from LPS injury, both in vitro and in vivo. These results suggest that Rab5 is a critical mediator of LPS-induced endothelial barrier dysfunction, which is likely mediated through regulating VE-cadherin internalization. These findings provide evidence, implicating that Rab5a is a potential therapeutic target for preventing endothelial barrier disruption and vascular inflammation.

  13. Dysregulation of coronary microvascular reactivity in asymptomatic patients with type 2 diabetes mellitus

    Energy Technology Data Exchange (ETDEWEB)

    Momose, Mitsuru; Neverve, Jodi; Nekolla, Stephan G.; Schwaiger, Markus; Bengel, Frank M. [Nuklearmedizinische Klinik und Poliklinik der Technischen Universitaet Muenchen, Klinikum rechts der Isar, Ismaninger Strasse 22, 81675 Munich (Germany); Abletshauser, Claudia [Department of Medicine, Novartis Pharma GmbH, Nuernberg (Germany); Schnell, Oliver; Standl, Eberhard [Institut fuer Diabetesforschung, Munich (Germany)

    2002-12-01

    In diabetic patients, a number of studies have suggested an impairment of vascular reactivity in response to vasodilatory stimuli. The pattern of dysregulation at the coronary microcirculatory level, however, has not been clearly defined. Thus, it was the aim of this study to characterise coronary microvascular function non-invasively in a homogeneous group of asymptomatic type 2 diabetic patients. In 46 patients with type 2 diabetes, myocardial blood flow (MBF) was quantified at baseline, in response to cold pressor test (CPT) and during adenosine-mediated vasodilation using positron emission tomography and nitrogen-13 ammonia. None of the patients had been treated with insulin, and none had symptoms of cardiac disease. Decreased MBF during CPT, indicating microvascular dysregulation, was observed in 16 patients (CPT-), while 30 patients demonstrated increased MBF during CPT (CPT+). Response to CPT was mildly, but significantly correlated with response to adenosine (r=0.44, P=0.0035). There was no difference in HbA1c, serum lipid levels or serum endothelial markers between the groups. Microvascular dysregulation in the CPT- group was associated with elevated baseline MBF (P<0.0001), reduced baseline vascular resistance (P=0.0026) and an abnormal increase in resistance during CPT (P=0.0002). In conclusion, coronary microvascular dysregulation is present in approximately one-third of asymptomatic, non-insulin-treated type 2 diabetic patients. Elevated baseline blood flow and reduced microvascular resistance at rest are characteristics of this dysregulation. These data suggest a state of activation of endothelial-dependent vasodilation at baseline which appears to limit the flow response to stress conditions. (orig.)

  14. Hyperosmolality-mediated peritoneal microvascular vasodilation is linked to aquaporin function.

    Science.gov (United States)

    Zakaria, El Rasheid; Althani, Asma; Fawzi, Ashraf A; Fituri, Omar M

    2014-01-01

    Glucose-based peritoneal dialysis (PD) solutions dilate the parietal and visceral peritoneal microvasculature by endothelium-dependent mechanisms that primarily involve hyperosmolality. This PD-mediated dilation occurs by active intracellular glucose uptake and adenosine Al receptor activation, and by hyperosmolality-stimulated glibenclamide-sensitive potassium channels. Both pathways invoke NO as a second messenger for vasodilation. We hypothesized that during crystalloid-induced osmosis, the osmotic water flux through the transendothelial water-exclusive aquaporin 1 (AQP1) channels is the primary mechanism whereby the endothelium is being stimulated to instigate hyperosmolality-driven vasodilation. Four microvascular levels (diameters in the range 6 - 100 microm) were visualized by intravital videomicroscopy of the terminal ileum in anesthetized rats. Microvascular diameters and flow were measured after topical exposure to a 5% hypertonic mannitol or 2.5% glucose-based PD solution, at baseline and after brief tissue pre-treatment (with 0.1% glutaraldehyde for 10 seconds) or after combined tissue pre-treatment and pharmacologic blockade of AQP1 with HgCl2 (100 micromol/L). Vascular endothelial integrity was verified by the response to acetylcholine (10(-4) mol/L) and sodium nitroprusside (10(-4) mol/L). The hyperosmolar solutions both caused rapid and sustained vasodilation at all microvascular levels, which was not altered by tissue pre-treatment. Inhibition of AQP1 completely abolished the mannitol-induced vasodilation and markedly attenuated the PD fluid-mediated vasodilation. Neither glutaraldehyde pre-treatment nor HgCl2 affected tissue integrity or endothelial cell function. We conclude that the peritoneal microvascular vasodilation caused by hyperosmolar PD fluid is instigated by the osmotic water flux through AQP1. Clinical PD solutions have components other than hyperosmolality that can induce endothelium-dependent peritoneal microvascular vasodilation

  15. Cryptogenic stroke following abdominal free flap breast reconstruction surgery

    Directory of Open Access Journals (Sweden)

    Huizhuang Xie

    2014-01-01

    CONCLUSION: Surgeons and cardiologists should be aware of this cerebrovascular complication secondary to PFO following major reconstructive surgery such as microvascular breast reconstruction. It also serves to challenge microvascular surgeons to reconsider routine use of central venous pressure lines in free flap patients who might otherwise have good peripheral vessels for postoperative fluid and antibiotic administration.

  16. Diabetic microvascular complications: possible targets for improved macrovascular outcomes

    Science.gov (United States)

    D’Elia, John A; Bayliss, George; Roshan, Bijan; Maski, Manish; Gleason, Ray E; Weinrauch, Larry A

    2011-01-01

    The results of recent outcome trials challenge hypotheses that tight control of both glycohemoglobin and blood pressure diminishes macrovascular events and survival among type 2 diabetic patients. Relevant questions exist regarding the adequacy of glycohemoglobin alone as a measure of diabetes control. Are we ignoring mechanisms of vasculotoxicity (profibrosis, altered angiogenesis, hypertrophy, hyperplasia, and endothelial injury) inherent in current antihyperglycemic medications? Is the polypharmacy for lowering cholesterol, triglyceride, glucose, and systolic blood pressure producing drug interactions that are too complex to be clinically identified? We review angiotensin–aldosterone mechanisms of tissue injury that magnify microvascular damage caused by hyperglycemia and hypertension. Many studies describe interruption of these mechanisms, without hemodynamic consequence, in the preservation of function in type 1 diabetes. Possible interactions between the renin–angiotensin–aldosterone system and physiologic glycemic control (through pulsatile insulin release) suggest opportunities for further clinical investigation. PMID:21694944

  17. The association of systemic microvascular changes with lung function and lung density: a cross-sectional study.

    Directory of Open Access Journals (Sweden)

    Bianca Harris

    Full Text Available Smoking causes endothelial dysfunction and systemic microvascular disease with resultant end-organ damage in the kidneys, eyes and heart. Little is known about microvascular changes in smoking-related lung disease. We tested if microvascular changes in the retina, kidneys and heart were associated with obstructive spirometry and low lung density on computed tomography. The Multi-Ethnic Study of Atherosclerosis recruited participants age 45-84 years without clinical cardiovascular disease. Measures of microvascular function included retinal arteriolar and venular caliber, urine albumin-to-creatinine ratio and, in a subset, myocardial blood flow on magnetic resonance imaging. Spirometry was measured following ATS/ERS guidelines. Low attenuation areas (LAA were measured on lung fields of cardiac computed tomograms. Regression models adjusted for pulmonary and cardiac risk factors, medications and body size. Among 3,397 participants, retinal venular caliber was inversely associated with forced expiratory volume in one second (FEV(1 (P<0.001 and FEV(1/forced vital capacity (FVC ratio (P = 0.04. Albumin-to-creatinine ratio was inversely associated with FEV(1 (P = 0.002 but not FEV(1/FVC. Myocardial blood flow (n = 126 was associated with lower FEV(1 (P = 0.02, lower FEV(1/FVC (P = 0.001 and greater percentage LAA (P = 0.04. Associations were of greater magnitude among smokers. Low lung function was associated with microvascular changes in the retina, kidneys and heart, and low lung density was associated with impaired myocardial microvascular perfusion. These cross-sectional results suggest that microvascular damage with end-organ dysfunction in all circulations may pertain to the lung, that lung dysfunction may contribute to systemic microvascular disease, or that there may be a shared predisposition.

  18. My First 100 Consecutive Microvascular Free Flaps: Pearls and Lessons Learned in First Year of Practice

    OpenAIRE

    Edward I. Chang, MD

    2013-01-01

    Background: Microvascular reconstruction for oncologic defects is a challenging and rewarding endeavor, and successful outcomes are dependent on a multitude of factors. This study represents lessons learned from a personal prospective experience with 100 consecutive free flaps. Methods: All patients’ medical records were reviewed for demographics, operative notes, and complications. Results: Overall 100 flaps were performed in 84 consecutive patients for reconstruction of breast, head a...

  19. 水蛭提取液对培养的大鼠脑皮质微血管内皮细胞分泌组织型纤溶酶原激活物和纤溶酶原激活剂抑制物1的影响%Effects of hirudo extract liquor on tissue-type plasminogen activator and plasminogen activator inhibitor-1 in microvascular endothelial cells from rat cerebral cortex

    Institute of Scientific and Technical Information of China (English)

    吴文斌; 胡长林; 董凌琳; 余能伟; 孙红斌; 郭富强

    2011-01-01

    目的 探讨水蛭提取液( HEL)对培养的大鼠脑皮质微血管内皮细胞分泌组织型纤溶酶原激活物(tPA)、纤溶酶原激活剂抑制物1( PAI-1)的影响.方法 建立大鼠大脑皮质微血管内皮细胞培养实验模型.MTT法筛选HEL的有效浓度.检测培养上清液的tPA、PAI-1含量与活性变化,RT-PCR检测经HEL治疗组与生理盐水对照组处理后的微血管内皮细胞tPA与PAI-1的表达,免疫组化检测两组微血管内皮细胞tPA的表达.结果 HEL在一定浓度范围内(0.25~1mg/μl)可促进微血管内皮细胞的生长,有剂量依赖关系(P<0.05).HEL治疗组较生理盐水对照组能促进培养的大鼠脑皮质微血管内皮细胞分泌tPA,同时提高其活性,促进tPA mRNA的表达及tPA免疫活性表达,且呈剂量依赖性表达增强(P<0.01).结论 HEL在体外能激活内源性纤溶系统.%Objective To study the effect of hirudo extract liquor (HEL) on activities of tissue-type plasminogen activator (tPA) and plasminogen activator inhibitor-1 (PAI-1), and levels of tPA and PAI-1 in microvascular endothelial cells of the rat cerebral cortex. Methods The experimental model of brain microvascular endothelial cells ( BMEC) of Wistar rat cerebral cortex was prepared in vitro. Cell morphology was observed under the inverted phase contrast microscope and cell activity was measured with MTT assay after BMEC exposure to the concentrations of HEL ranging from 0.0625 to 8 mg/μl. The biochemical index, including activitives and leveb of tPA and PAI-1 in cultured supernatants, as well as variation of semi-quantification of tPA, PAI-1 mRNA levels were measured in BMEC by reverse transcription polymerase chain reaction (RT-PCR) in the HEL treatment group and the control group normal saline treatment. The activities of tPA and PAI were measured by colorimetric assay. The contents of tPA and PAI-1 were determined using specific ELISA. The expression of tPA protein in BMEC was measured by

  20. Intravascular Stenting in Microvascular Anastomoses

    DEFF Research Database (Denmark)

    Assersen, Kristine; Sørensen, Jens

    2015-01-01

    Background The effect of intravascular stenting (IVaS) on microvascular anastomoses has given adverse results. For experienced microsurgeons the benefit of IVaS is doubtful. We have investigated the potential benefit of the IVaS technique for two groups of inexperienced microsurgeons with different...... surgical levels of experience (medical students and young residents). Experienced microsurgeons acted as a control group. Materials and Methods In an experimental crossover study, 139 microsurgical anastomoses were performed on the femoral artery in 70 rats by 10 surgeons. On one side of the rat, the IVaS...... spent on the anastomosis. Results No significant difference in patency rates was seen between the stenting and conventional technique in all three groups. The experienced microsurgeons had 100% patency rate with both techniques. The medical students had 20/28 in the IVaS and 19/28 conventional group...

  1. Coronary microvascular dysfunction in overt diabetic cardiomyopathy

    Directory of Open Access Journals (Sweden)

    K. Bratis

    2014-11-01

    Conclusion: In patients with DM2 myocardial perfusion reserve is markedly decreased, suggestive of microvascular disease. In this small cohort MPRI impairment did not correlate to the LV EF deterioration.

  2. A biomimetic collagen derived peptide exhibits anti-angiogenic activity in triple negative breast cancer.

    Directory of Open Access Journals (Sweden)

    Elena V Rosca

    Full Text Available We investigated the application of a mimetic 20 amino acid peptide derived from type IV collagen for treatment of breast cancer. We showed that the peptide induced a decrease of proliferation, adhesion, and migration of endothelial and tumor cells in vitro. We also observed an inhibition of triple negative MDA-MB-231 xenograft growth by 75% relative to control when administered intraperitoneally for 27 days at 10 mg/kg. We monitored in vivo the changes in vascular properties throughout the treatment using MRI and found that the vascular volume and permeability surface area product decreased significantly. The treatment also resulted in an increase of caspase-3 activity and in a reduction of microvascular density. The multiple mode of action of this peptide, i.e., anti-angiogenic, and anti-tumorigenic, makes it a viable candidate as a therapeutic agent as a monotherapy or in combination with other compounds.

  3. Effects of vascular endothelial growth factor on angiogenesis of the endothelial cells isolated from cavernous malformations

    Institute of Scientific and Technical Information of China (English)

    TAN YuZhen; ZHAO Yao; WANG HaiJie; ZHOU LiangFu; MAO Ying; LIU Rui; SHU Jia; WANG YongFei

    2008-01-01

    Human cerebral cavernous malformation (CM) is a common vascular malformation of the central nervous system. We have investigated the biological characteristics of CM endothelial cells and the cellular and molecular mechanisms of CM angiogenesis to offer new insights into exploring effective measures for treatment of this disease. The endothelial cells were isolated from CM tissue masses dissected during operation and expanded in vitro. Expression of VEGFR-1 and VEGFR-2 was examined with immunocytochemical staining. Proliferation, migration and tube formation of CM endothelial cells were determined using MTT, wounding and transmigration assays, and three-dimensional collagen type Ⅰ gel respectively. The endothelial cells were successfully isolated from the tissue specimens of 25 CMs dissected without dipolar electrocoagulation. The cells show the general characteristics of the vascular endothelial cells. Expression of VEGFR-1 and VEGFR-2 on the cells is higher than that on the normal cerebral microvascular endothelial cells. After treatment with VEGF, numbers of the proliferated and migrated cells, the maximal distance of cell migration and the length and area of capillary-like struc-tures formed in the three-dimensional collagen gel increase significantly. These results demonstrate that expression of VEGFR-1 and VEGFR-2 on CM endothelial cells is up-regulated. By binding to re-ceptors, VEGF may activate the downstream signaling pathways and promote proliferation, migration and tube formation of CM endothelial cells. VEGF/VEGFR signaling pathways play important regulating roles in CM angiogenesis.

  4. Effect of silencing Fas expression by RNA interference on mice brain microvascular endothelial cell line bEnd.3%RNA干扰抑制Fas基因的表达对小鼠脑微血管内皮细胞bEnd.3的影响

    Institute of Scientific and Technical Information of China (English)

    张春兵; 高峰; 张明顺; 滕凤猛

    2013-01-01

    Objective To investigate the impact of reduced Fas expression by RNA interference on the proliferation of mice brain microvascular endothelial cell line bend.3 and the signal transduction pathway of Fas-associated death domain-containing protein (FADD),FADD-like IL-1β-converting enzyme (FLIP),tumor necrosis factor receptor-associated factor (TRAF) and nuclear factor κB (NF-κB).Methods The siRNA fragment targeted to mice Fas gene was designed and synthesized,and transfected into bEnd.3 cells by Lipofection 2000.The cell proliferation was measured using cell counting kit-8 (CCK-8).The expression levels of Fas mRNA and Fas protein were measured by real-time quantitative PCR and western blot respectively.The levels of FADD,FLIP and TRAF protein were measured by western blot simultaneously.Results CCK-8 assay demonstrated that no significantly difference of cell proliferation was found between Fas-siRNA group and blank control group (t =1.805,P > 0.05).Quantification analysis showed that Fas mRNA expression levels were markedly decreased in Fas-228-,Fas-310-,Fas-427-and Fas-891-siRNA group compared with the blank control group (F =123.127,P < 0.05).Western blot indicated that Fas-siRNA significantly reduced the expression of Fas protein (t =12.101,P < 0.01).The relative expression levels of FADD,FLIP and TRAF were significantly decreased as compared with blank group(t =28.315,17.563 and 8.903,P < 0.05).Conclusion Fas-siRNA can effectively block the expression of Fas gene in bEnd.3 cell and decrease the protein levels of Fas downstream signal molecules FADD,FLIP and TRAF.%目的 用RNA干扰(RNAi)技术抑制小鼠Fas基因表达,观察其对脑微血管内皮细胞bEnd.3增殖及FADD-FLIP-TRAFNF-κB信号传导途径的影响,为后续FasL低剂量兴奋效应研究提供实验基础.方法 设计合成靶向小鼠Fas基因的小干扰RNA (siRNA)片段,用脂质体包埋转染bEnd.3细胞;CCK-8法检测细胞增殖;RT-PCR检测bEnd.3细胞Fas m

  5. Higher vascular endothelial growth factor-C concentration in plasma is associated with increased forearm capillary filtration capacity in breast cancer-related lymphedema

    DEFF Research Database (Denmark)

    Jensen, Mads Radmer; Simonsen, Lene; Karlsmark, Tonny

    2015-01-01

    Breast cancer-related lymphedema (BCRL) is a frequent, chronic and debilitating swelling that mainly affects the ipsilateral arm and develops as a complication to breast cancer treatment. The pathophysiology is elusive opposing development of means for prediction and treatment. We have earlier...

  6. Endothelial Genes

    Science.gov (United States)

    2005-06-01

    8217Department of Surgery, Division of Oncology , and 2Department of BRCA-l and BRCA-2 (breast cancer susceptibility genes), Pathology, University of...Suppression subtractive hybridization re- Cancer: principles and practice of oncology . Philadelphia: Lippincott- vealed an RNA sequence (GenBank accession...Lippman ME. Cancer of the breast: molecular biology angiogenesis in sarcomas and carcinomas. Clin Cancer Res 1999;5: of breast cancer. In: DeVita VT

  7. Glycogen Synthase Kinase-3 Inhibitor Protects Against Microvascular Hyperpermeability Following Hemorrhagic Shock

    Science.gov (United States)

    Sawant, Devendra A.; Tharakan, Binu; Hunter, Felicia A.; Childs, Ed W.

    2015-01-01

    Background Hemorrhagic shock (HS)-induce microvascular hyperpermeability involves disruption of endothelial cell adherens junctions leading to increase in paracellular permeability. β-Catenin, an integral component of the adherens junctional complex and Wnt pathway, and caspase-3 via its apoptotic signaling regulate endothelial cell barrier integrity. We have hypothesized that inhibiting phosphorylation of β-catenin and caspase-3 activity using glycogen synthase kinase-3 (GSK-3) specific inhibitor SB216763, would attenuate microvascular hyperpermeability following HS. Methods In Sprague-Dawley rats, HS was induced by withdrawing blood to reduce mean arterial pressure to 40 mmHg for 60 minutes followed by resuscitation. Rats were given SB216763 (600 μg/kg) intravenously 10 minutes prior to shock. To study microvascular permeability, the rats were injected intravenously with FITC-albumin (50 mg/kg) and its flux across the mesenteric post-capillary venules was determined using intravital microscopy. In cell-culture studies, rat lung microvascular endothelial cell (RLMEC) monolayers grown on Transwell plates were pre-treated with SB216763 (5 μM) followed by BAK (5 μg/mL) and caspase-3 (5 μg/mL) protein transfection. FITC-albumin (5 mg/mL) flux across cell monolayers indicates change in monolayer permeability. Activity of canonical Wnt pathway was determined by luciferase assay. Caspase-3 enzyme activity was assayed fluorometrically. Results The HS group showed significant increase in FITC-albumin extravasation (p<0.05) compared with sham. SB216763 significantly decrease HS-induced FITC-albumin extravasation (p<0.05). Pre-treatment with SB216763, protected against a BAK-induced increase in RLMEC monolayer permeability and caspase-3 activity, but failed to show similar results with a caspase-3-induced increase in monolayer permeability. Wnt3a treatment showed an increase in β-catenin dependent TCF-mediated transcription. Conclusion Inhibiting phosphorylation of

  8. Physiological hydrostatic pressure protects endothelial monolayer integrity.

    Science.gov (United States)

    Müller-Marschhausen, K; Waschke, J; Drenckhahn, D

    2008-01-01

    Endothelial monolayer integrity is required to maintain endothelial barrier functions and has found to be impaired in several disorders like inflammatory edema, allergic shock, or artherosclerosis. Under physiologic conditions in vivo, endothelial cells are exposed to mechanical forces such as hydrostatic pressure, shear stress, and cyclic stretch. However, insight into the effects of hydrostatic pressure on endothelial cell biology is very limited at present. Therefore, in this study, we tested the hypothesis that physiological hydrostatic pressure protects endothelial monolayer integrity in vitro. We investigated the protective efficacy of hydrostatic pressure in microvascular myocardial endothelial (MyEnd) cells and macrovascular pulmonary artery endothelial cells (PAECs) by the application of selected pharmacological agents known to alter monolayer integrity in the absence or presence of hydrostatic pressure. In both endothelial cell lines, extracellular Ca(2+) depletion by EGTA was followed by a loss of vascular-endothelial cadherin (VE-caherin) immunostaining at cell junctions. However, hydrostatic pressure (15 cmH(2)O) blocked this effect of EGTA. Similarly, cytochalasin D-induced actin depolymerization and intercellular gap formation and cell detachment in response to the Ca(2+)/calmodulin antagonist trifluperazine (TFP) as well as thrombin-induced cell dissociation were also reduced by hydrostatic pressure. Moreover, hydrostatic pressure significantly reduced the loss of VE-cadherin-mediated adhesion in response to EGTA, cytochalasin D, and TFP in MyEnd cells as determined by laser tweezer trapping using VE-cadherin-coated microbeads. In caveolin-1-deficient MyEnd cells, which lack caveolae, hydrostatic pressure did not protect monolayer integrity compromised by EGTA, indicating that caveolae-dependent mechanisms are involved in hydrostatic pressure sensing and signaling.

  9. Uterine microvascular sensitivity to nanomaterial inhalation: An in vivo assessment☆

    Science.gov (United States)

    Stapleton, P.A.; McBride, C.R.; Yi, J.; Nurkiewicz, T.R.

    2015-01-01

    With the tremendous number and diverse applications of engineered nanomaterials incorporated in daily human activity, exposure can no longer be solely confined to occupational exposures of healthy male models. Cardiovascular and endothelial cell dysfunction have been established using in vitro and in situ preparations, but the translation to intact in vivo models is limited. Intravital microscopy has been used extensively to understand microvascular physiology while maintaining in vivo neurogenic, humoral, and myogenic control. However, a tissue specific model to assess the influences of nanomaterial exposure on female reproductive health has not been fully elucidated. Female Sprague Dawley (SD) rats were exposed to nano-TiO2 aerosols (171 ± 6 nm, 10.1 ± 0.39 mg/m3, 5 h) 24-hours prior to experimentation, leading to a calculated deposition of 42.0 ± 1.65 μg. After verifying estrus status, vital signs were monitored and the right horn of the uterus was exteriorized, gently secured over an optical pedestal, and enclosed in a warmed tissue bath using intravital microscopy techniques. After equilibration, significantly higher leukocyte-endothelium interactions were recorded in the exposed group. Arteriolar responsiveness was assessed using ionophoretically applied agents: muscarinic agonist acetylcholine (0.025 M; ACh; 20, 40, 100, and 200 nA), and nitric oxide donor sodium nitroprusside (0.05 M; SNP; 20, 40, and 100 nA), or adrenergic agonist phenylephrine (0.05 M; PE; 20, 40, and 100 nA) using glass micropipettes. Passive diameter was established by tissue superfusion with 10−4 M adenosine. Similar to male counterparts, female SD rats present systemic microvascular dysfunction; however the ramifications associated with female health and reproduction have yet to be elucidated. PMID:26375943

  10. MicroRNA-223-3p inhibits the angiogenesis of ischemic myocardial microvascular endothelial cells via modulating Rps6kb1/HIF-1α signal pathway%微小RNA-223-3p通过调节Rps6kb1/HIF-1α信号通路抑制缺血心肌微血管内皮细胞血管新生

    Institute of Scientific and Technical Information of China (English)

    戴国华; 宋宪波; 马培泽; 刘宁; 姚静

    2014-01-01

    Objective To explore the role of microRNA on the myocardial microvascular endothelial cells (CMECs) of ischemic heart rats in the process of angiogenesis and related regulation mechanism.Methods Myocardial ischemic rats model was established by coronary ligation.Seven days after operation,the ischemic CMECs were cultured by the method of planting myocardium tissue and identified by immunocytochemistry to observe the biological characteristics of ischemic CMECs angiogenesis,to determine the window period of migration,proliferation,tube formation in the process of its angiogenesis.Dynamic expression changes of microRNA in the process of ischemic CMECs angiogenesis was detected using microRNA chip and further verified by real-time PCR,the core microRNA of the ischemic CMECs was defined and the predicted target genes of core microRNA were determined by bioinformatics methods and real-time PCR.At the same time,the protein expression of target gene and angiogenesis related genes of p38MAPK,PI3K,Akt,VEGF were measured by Western blot.Results The CMECs of rats presented typical characteristics of microvascular endothelial cells,and factor Ⅷ,CD3 1 related antigens were all positively stained by immunocytochemical analysis.The migration window period was on the first day,and the tube formation window period was on the second day of both control and ischemic groups,while the proliferation window period was on the third day for the normal group,and the sixth day for ischemic group.According to the expressional difference and their relationship with angiogenesis,miRNA-223-3p was ultimately determined as the core microRNA in the process of ischemic CMECs angiogenesis,real-time PCR verified this hypothesis.Bioinformatics methods predicted that Rps6kb1 is the target genes of miRNA-223-3p,the pathway analysis showed that Rps6kb1 could regulate angiogenesis via HIF-1α signal pathway.Moreover,the mRNA and protein expression of VEGF,p38MAPK,PI3K,Akt,which were the downstream

  11. Tumor endothelial expression of P-glycoprotein upon microvesicular transfer of TrpC5 derived from adriamycin-resistant breast cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Dong, YePing; Pan, QiongXi; Jiang, Li; Chen, Zhen; Zhang, FangFang; Liu, YanJun; Xing, Hui; Shi, Mei; Li, Jiao; Li, XiYuan; Zhu, YaoDan; Chen, Yun; Bruce, Iain C.; Jin, Jian, E-mail: jinjian31@126.com; Ma, Xin, E-mail: maxin@jiangnan.edu.cn

    2014-03-28

    Highlights: • TrpC5 was mainly accumulated in microvesicles of drug-resistant MCF-7/ADM cells. • Microvesicles from MCF-7/ADM transferred TrpC5 to endothelial cells. • TrpC5 inhibition reduced P-glycoprotein accumulation on tumor blood vessels in vivo. - Abstract: Treatment of carcinoma commonly fails due to chemoresistance. Studies have shown that endothelial cells acquire resistance via the tumor microenvironment. Microvesicle (MV) shedding from the cell membrane to the microenvironment plays an important role in communication between cells. The aim of the present study was to determine whether MCF-7 adriamycin-resistant cells (MCF-7/ADM) shed MVs that alter the characteristics of human microvessel endothelial cells (HMECs). MVs from tumor cells transferred a Ca{sup 2+}-permeable channel TrpC5 to HMECs, inducing the expression of P-glycoprotein (P-gp) by activation of the transcription factor NFATc3 (nuclear factor of activated T cells isoform c3). Expression of the mdr1 gene was blocked by the TrpC5-blocking antibody T5E3, and the production of P-gp in HMECs was reduced by blockade of TrpC5. Thus, we postulate that endothelial cells acquire the resistant protein upon exposure to TrpC5-containg MVs in the microenvironment, and express P-gp in the TrpC5–NFATc3 signal pathway.

  12. Coronary microvascular obstruction in acute myocardial infarction.

    Science.gov (United States)

    Niccoli, Giampaolo; Scalone, Giancarla; Lerman, Amir; Crea, Filippo

    2016-04-01

    The success of a primary percutaneous intervention (PCI) in the setting of ST elevation myocardial infarction depends on the functional and structural integrity of coronary microcirculation. Coronary microvascular dysfunction and obstruction (CMVO) occurs in up to half of patients submitted to apparently successful primary PCI and is associated to a much worse outcome. The current review summarizes the complex mechanisms responsible for CMVO, including pre-existing coronary microvascular dysfunction, and highlights the current limitations in the assessment of microvascular function. More importantly, at the light of the substantial failure of trials hitherto published on the treatment of CMVO, this review proposes a novel integrated therapeutic approach, which should overcome the limitations of previous studies.

  13. 腺病毒介导的Slit2及Slit2 ShRNA转染缺氧诱导的人RPE细胞对人脉络膜微血管内皮细胞增殖的影响%Effects of the hypoxia-induced human retinal pigment epithelial cells which transfected by adenovirus-mediated Slit2 and adenovirus-mediated Slit2 ShRNA on the proliferation of human choroidal microvascular endothelial cells

    Institute of Scientific and Technical Information of China (English)

    汤艳玲; 周希瑗

    2014-01-01

    目的:观察腺病毒介导的Slit2及Slit2 ShRNA转染缺氧诱导的人视网膜色素上皮(retinal pigment epithelial cells,RPE)细胞对人脉络膜微血管内皮细胞(human choroidal microvascular endothelial cell,HCMEC)增殖的影响,探讨Slit2在脉络膜新生血管中的可能作用,为脉络膜新生血管(choroidal neovascularization,CNV)提供新的治疗思路.方法:体外培养并鉴定人RPE细胞、HCMEC;200 μmol/L氯化钴建立化学缺氧模型,Transwell小室建立细胞共培养模型;将缺氧的RPE细胞随机分为Slit2组(加入Slit2)、Slit2 ShRNA组(加入Slit2 ShRNA)、空腺病毒组(加入空腺病毒)、缺氧组,12、24、48 h后采用CCK 8(Cell Counting Kit-8,CCK 8)法检测HCMEC的增殖.结果:不同组别存在组间差别,差异均有统计学意义(F=98.122,P=0.000),不同时间点存在差别(F=3388.913,P=0.000),组别与时间点的交互作用(F=82.863,P=0.000).Slit2组吸光度(absorbance,A)值在24 h、48 h均高于其他组(与缺氧组P=0.001,其余P=0.000),Slit2 ShRNA组A值在24 h、48 h均低于其他组(48 h与缺氧组P=0.003,与空腺病毒组P=0.008,其余P=0.000).结论:Slit2的高表达可明显促进HCMEC的增殖,沉默RPE细胞中的Slit2的表达后,会明显抑制HCMEC的增殖.

  14. Early nuclear alterations and immunohistochemical expression of Ki-67, Erb-B2, vascular endothelial growth factor (VEGF), transforming growth factor (TGF-beta1) and integrine-linked kinase (ILK) two days after tamoxifen in breast carcinoma.

    Science.gov (United States)

    Morena, A M L; Oshima, C T F; Gebrim, L H; Egami, M I; Silva, M R R; Segreto, R A; Giannotti Filho, O; Teixeira, V P C; Segreto, H R C

    2004-01-01

    The purpose of the present study was to evaluate breast carcinoma samples before and two days after treatment with tamoxifen in order to analyse early histopathological alterations--particularlynuclear alterations-- as well as immunohistochemical expression of Ki-67, Erb-B2, VEGF, TGF-beta1 and ILK proteins. Twenty one cases of invasive ductal and lobular breast carcinoma were studied. Patients were submitted to biopsy of the lesion and, after confirmation of the diagnosis, they received 20 mg of tamoxifen a day, beginning two days before surgery. The samples obtained during biopsy and after surgery were stained with HE for histopathological diagnosis. Estrogen receptor was positive in 18 cases and negative in 3. The immunohistochemical method was applied for the detection of Ki-67, Erb-B2, protein, vascular endothelial growth factor (VEGF), transforming growth factor beta (TGF-beta1) and integrin linked kinase (ILK). Two days after tamoxifen treatment, the following results were observed: 1) decrease in the cell volume, chomatine condensation, nucleoli less evident and clearly defined nuclear limits; 2) significant reduction in the expression of Erb-B2 protein and significant increase in the expression of TGF-beta1 protein; 3) expression of others proteins (Ki-67, VEGF and ILK) was not altered during the indicated time frame. Our results suggest that analyzing nuclear alterations and expression of Erb-B2 and TGF-beta1 proteins would be useful to assess the initial response to tamoxifen.

  15. Fat embolism syndrome and pulmonary microvascular cytology.

    Science.gov (United States)

    Castella, X; Vallés, J; Cabezuelo, M A; Fernandez, R; Artigas, A

    1992-06-01

    Pulmonary microvascular cytology consists of analysis of capillary blood sampled while a Swan-Ganz catheter is in the wedge position. This technique has proved to be useful in the diagnosis of lymphangitic spread of carcinoma in the lungs and there are case reports of their use in amniotic fluid embolism. Its usefulness in diagnosing fat embolism syndrome has been shown only rarely. We report a new case in which pulmonary microvascular cytologic study allowed a definite diagnosis of fat embolism syndrome. We suggest obtaining routinely samples of capillary blood when a pulmonary catheter is in place and fat embolism is suspected on a clinical basis.

  16. Role of genetic polymorphisms of ion channels in the pathophysiology of coronary microvascular dysfunction and ischemic heart disease.

    Science.gov (United States)

    Fedele, Francesco; Mancone, Massimo; Chilian, William M; Severino, Paolo; Canali, Emanuele; Logan, Suzanna; De Marchis, Maria Laura; Volterrani, Maurizio; Palmirotta, Raffaele; Guadagni, Fiorella

    2013-11-01

    Conventionally, ischemic heart disease (IHD) is equated with large vessel coronary disease. However, recent evidence has suggested a role of compromised microvascular regulation in the etiology of IHD. Because regulation of coronary blood flow likely involves activity of specific ion channels, and key factors involved in endothelium-dependent dilation, we proposed that genetic anomalies of ion channels or specific endothelial regulators may underlie coronary microvascular disease. We aimed to evaluate the clinical impact of single-nucleotide polymorphisms in genes encoding for ion channels expressed in the coronary vasculature and the possible correlation with IHD resulting from microvascular dysfunction. 242 consecutive patients who were candidates for coronary angiography were enrolled. A prospective, observational, single-center study was conducted, analyzing genetic polymorphisms relative to (1) NOS3 encoding for endothelial nitric oxide synthase (eNOS); (2) ATP2A2 encoding for the Ca²⁺/H⁺-ATPase pump (SERCA); (3) SCN5A encoding for the voltage-dependent Na⁺ channel (Nav1.5); (4) KCNJ8 and KCNJ11 encoding for the Kir6.1 and Kir6.2 subunits of K-ATP channels, respectively; and (5) KCN5A encoding for the voltage-gated K⁺ channel (Kv1.5). No significant associations between clinical IHD manifestations and polymorphisms for SERCA, Kir6.1, and Kv1.5 were observed (p > 0.05), whereas specific polymorphisms detected in eNOS, as well as in Kir6.2 and Nav1.5 were found to be correlated with IHD and microvascular dysfunction. Interestingly, genetic polymorphisms for ion channels seem to have an important clinical impact influencing the susceptibility for microvascular dysfunction and IHD, independent of the presence of classic cardiovascular risk factors.

  17. Microvascular angina in postmenopausal women.

    Directory of Open Access Journals (Sweden)

    Alexander Valdés Martín

    2011-10-01

    Full Text Available Fundamento: la angina microvascular es frecuente en mujeres posmenopáusicas. La isquemia miocárdica ha sido inducida mediante pruebas de estrés, en las que se ha comprobado una relación entre la disfunción endotelial y los defectos de perfusión miocárdica. Objetivo: determinar si la isquemia miocárdica puede evidenciarse por anormalidades de la perfusión y de la función detectadas por gammagrafía miocárdica en mujeres con angina típica, angiografía coronaria normal y disfunción endotelial. Métodos: estudio descriptivo realizado en el Instituto de Cardiología y Cirugía Cardiovascular de La Habana que incluyó 59 mujeres. Se les realizó lipidograma, se les midió función endotelial de la arteria braquial mediante ultrasonido, y estudio electrocardiográfico de 24 horas (Holter. Se aplicó un protocolo de estrés-reposo durante la gammagrafía. Las pacientes fueron divididas en dos grupos, acorde a la presencia (grupo I o ausencia (grupo II de defectos de perfusión miocárdica. Resultados: mostraron defectos de la perfusión 21 pacientes. El 57 % de las pacientes del grupo I exhibió más disfunción endotelial. Sólo doce pacientes mostraron defectos reversibles de la perfusión y en el 75 % de los casos se asoció a una reducción de la fracción de eyección ventricular izquierda postestrés mayor de un 5 % y a anormalidades regionales de la motilidad de la pared. Tres pacientes en el grupo I mostraron evidencia de isquemia comparado con cuatro en el grupo II. Conclusiones: la isquemia inducida por estrés se asocia a una reducción de la fracción de eyección ventricular izquierda postestrés y a una vasodilatación anormal dependiente del endotelio.

  18. Endothelial dysfunction in diabetes mellitus

    Directory of Open Access Journals (Sweden)

    Hadi AR Hadi

    2008-01-01

    Full Text Available Hadi AR Hadi, Jassim Al SuwaidiDepartment of Cardiology and Cardiovascular Surgery, Hamad General Hospital – Hamad Medical Corporation, Doha, State of Qatar; Department of Cardioscience, Sheikh Khalifa Medical City, Abu Dhabi, UAEAbstract: Diabetes mellitus is associated with an increased risk of cardiovascular disease, even in the presence of intensive glycemic control. Substantial clinical and experimental evidence suggest that both diabetes and insulin resistance cause a combination of endothelial dysfunctions, which may diminish the anti-atherogenic role of the vascular endothelium. Both insulin resistance and endothelial dysfunction appear to precede the development of overt hyperglycemia in patients with type 2 diabetes. Therefore, in patients with diabetes or insulin resistance, endothelial dysfunction may be a critical early target for preventing atherosclerosis and cardiovascular disease. Microalbuminuria is now considered to be an atherosclerotic risk factor and predicts future cardiovascular disease risk in diabetic patients, in elderly patients, as well as in the general population. It has been implicated as an independent risk factor for cardiovascular disease and premature cardiovascular mortality for patients with type 1 and type 2 diabetes mellitus, as well as for patients with essential hypertension. A complete biochemical understanding of the mechanisms by which hyperglycemia causes vascular functional and structural changes associated with the diabetic milieu still eludes us. In recent years, the numerous biochemical and metabolic pathways postulated to have a causal role in the pathogenesis of diabetic vascular disease have been distilled into several unifying hypotheses. The role of chronic hyperglycemia in the development of diabetic microvascular complications and in neuropathy has been clearly established. However, the biochemical or cellular links between elevated blood glucose levels, and the vascular lesions remain

  19. Reduction of neutrophil activity decreases early microvascular injury after subarachnoid haemorrhage

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    Bi Weina

    2011-08-01

    Full Text Available Abstract Background Subarachnoid haemorrhage (SAH elicits rapid pathological changes in the structure and function of parenchymal vessels (≤ 100 μm. The role of neutrophils in these changes has not been determined. This study investigates the role of neutrophils in early microvascular changes after SAH Method Rats were either untreated, treated with vinblastine or anti-polymorphonuclear (PMN serum, which depletes neutrophils, or treated with pyrrolidine dithiocarbamate (PDTC, which limits neutrophil activity. SAH was induced by endovascular perforation. Neutrophil infiltration and the integrity of vascular endothelium and basement membrane were assessed immunohistochemically. Vascular collagenase activity was assessed by in situ zymography. Results Vinblastine and anti-PMN serum reduced post-SAH accumulation of neutrophils in cerebral vessels and in brain parenchyma. PDTC increased the neutrophil accumulation in cerebral vessels and decreased accumulation in brain parenchyma. In addition, each of the three agents decreased vascular collagenase activity and post-SAH loss of vascular endothelial and basement membrane immunostaining. Conclusions Our results implicate neutrophils in early microvascular injury after SAH and indicate that treatments which reduce neutrophil activity can be beneficial in limiting microvascular injury and increasing survival after SAH.

  20. Changes of microvascular architecture, ultrastructure and permeability of rat jejunal villi at different ages

    Institute of Scientific and Technical Information of China (English)

    Yan-Min Chen; Jin-Sheng Zhang; Xiang-Lin Duan

    2003-01-01

    AIM: To investigate the changes of microvascular architecture, ultrastructure and permeability of rat jejunal villi at different ages.METHODS: Microvascular corrosion casting, scanning electron microscopy, transmission electron microscopy and Evans blue infiltration technique were used in this study.RESULTS: The intestinal villous plexus of adult rats consisted of arterioles, capillary network and venules. The marginal capillary extended to the base part of the villi and connected to the capillary networks of adjacent villi. In newborn rats,the villous plexus was rather simple, and capillary network was not formed. The villous plexus became cone-shaped and was closely arrayed in ablactation rats. In adult rats,the villous plexus became tongue-shaped and was enlarged both in height and width. In aged rats, the villous plexus shrank in volume and became shorter and narrower. The diametral ratio of villous arteriole to villous venule increased as animals became older. The number of endothelial holes,the thickness of basal membrane and the permeability of microvasculature were increased over the entire course of development from newborn period to aged period.CONCLUSION: The digestive and absorptive functions of the rat jejunum at different ages are highly dependent upon the state of villous microvascular architecture and permeability, and blood circulation is enhanced by collateral branches such as marginal capillary, through which blood is drained to the capillary networks of adjacent villi.

  1. Inhibition of autophagy ameliorates pulmonary microvascular dilation and PMVECs excessive proliferation in rat experimental hepatopulmonary syndrome

    Science.gov (United States)

    Xu, Duo; Chen, Bing; Gu, Jianteng; Chen, Lin; Belguise, Karine; Wang, Xiaobo; Yi, Bin; Lu, Kaizhi

    2016-01-01

    Hepatopulmonary syndrome (HPS) is a defective liver-induced pulmonary vascular disorder with massive pulmonary microvascular dilation and excessive proliferation of pulmonary microvascular endothelial cells (PMVECs). Growing evidence suggests that autophagy is involved in pulmonary diseases, protectively or detrimentally. Thus, it is interesting and important to explore whether autophagy might be involved in and critical in HPS. In the present study, we report that autophagy was activated in common bile duct ligation (CBDL) rats and cultured pulmonary PMVECs induced by CBDL rat serum, two accepted in vivo and in vitro experimental models of HPS. Furthermore, pharmacological inhibition of autophagy with 3-methyladenine (3-MA) significantly alleviated pathological alterations and typical symptom of HPS in CBDL rats in vivo, and consistently 3-MA significantly attenuated the CBDL rat serum-induced excessive proliferation of PMVECs in vitro. All these changes mediated by 3-MA might explain the observed prominent improvement of pulmonary appearance, edema, microvascular dilatation and arterial oxygenation in vivo. Collectively, these results suggest that autophagy activation may play a critical role in the pathogenesis of HPS, and autophagy inhibition may have a therapeutic potential for this disease. PMID:27480323

  2. Microvascular and immunological studies in Raynaud's phenomenon.

    NARCIS (Netherlands)

    Houtman, Pieternella Maria

    1985-01-01

    The purpose of this thesis was to investigate the diagnostic significance of microvascular abnormalities - as observed in the nailfold - in patients with RP with respect to the presence or development of a connective tissue disease. In addition, we investigated whether the observed abnormalities wer

  3. Label-free quantitative cell division monitoring of endothelial cells by digital holographic microscopy

    Science.gov (United States)

    Kemper, Björn; Bauwens, Andreas; Vollmer, Angelika; Ketelhut, Steffi; Langehanenberg, Patrik; Müthing, Johannes; Karch, Helge; von Bally, Gert

    2010-05-01

    Digital holographic microscopy (DHM) enables quantitative multifocus phase contrast imaging for nondestructive technical inspection and live cell analysis. Time-lapse investigations on human brain microvascular endothelial cells demonstrate the use of DHM for label-free dynamic quantitative monitoring of cell division of mother cells into daughter cells. Cytokinetic DHM analysis provides future applications in toxicology and cancer research.

  4. High Ki-67 and Vascular Endothelial Growth Factor (VEGF Protein Expression as Negative Predictive Factor for Combined Neoadjuvant Chemotherapy in Young Age Stage III Breast Cancer

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    I Wayan Sudarsa

    2016-05-01

    Full Text Available Background: Breast cancer was, in general, a heterogeneous disease with diverse biological characteristics, types, subtypes and clinical behavior. Its treatment and management need to be personalized and individualized. Breast cancer in young ages, although rare, is usually a unique and more aggressive cancer associated with poorer prognosis. The combination of young age and advanced stages of breast cancer would make this particular breast cancer harder to treat and cure. Unfortunately, majority of Breast Cancer Patients in Bali were in younger ages, and at advanced stages, that the mainstay of treatment was neo-adjuvant chemotherapy followed by other treatment modalities. Improve prognosis only, those patients who had had a complete pathological response involving primary tumor and regional lymph nodes in the axilla. Several factors had been studied and contributed to breast cancer response to combined neo-adjuvant chemotherapy. Usually, younger patients, was associated with high proliferation rate represented by Ki-67 and early distant metastasis represented by VEGF, which also had role as prognostic markers. The purpose of this study was to determine whether high Ki-67 and VEGF expression correlate with response to NAC and hence, they would be important predictive factors for response to NAC. Method: This study was a cross-sectional and a nested case-control study of stage III breast cancers affecting patients 40 years of age or less, at Sanglah General Hospital and Prima Medika Hospital, conducted from September 1st, 2012 until March 31st, 2014. Clinical and pathology reports were traced and recorded from both hospitals; routine Immunohistochemistry (IHC examinations were performed by both pathology labs. Statistical analysis was performed using Chi-Square test, Odds Ratio (OR, and logistic regression analysis with p<0.05. Results: There were 66 Stage III young breast cancer patients, where 35 (53% showed no or negative response and 31 (47

  5. Diabetic microvascular complications: possible targets for improved macrovascular outcomes

    Directory of Open Access Journals (Sweden)

    Bijan Roshan

    2010-12-01

    Full Text Available John A D’Elia1, George Bayliss1,2, Bijan Roshan1, Manish Maski1, Ray E Gleason1, Larry A Weinrauch11Renal Unit, Joslin Diabetes Center, Department of Medicine, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA, USA; 2Department of Medicine, Rhode Island Hospital, Alpert School of Medicine, Brown University, Providence, RI, USAAbstract: The results of recent outcome trials challenge hypotheses that tight control of both glycohemoglobin and blood pressure diminishes macrovascular events and survival among type 2 diabetic patients. Relevant questions exist regarding the adequacy of glycohemoglobin alone as a measure of diabetes control. Are we ignoring mechanisms of vasculotoxicity (profibrosis, altered angiogenesis, hypertrophy, hyperplasia, and endothelial injury inherent in current antihyperglycemic medications? Is the polypharmacy for lowering cholesterol, triglyceride, glucose, and systolic blood pressure producing drug interactions that are too complex to be clinically identified? We review angiotensin–aldosterone mechanisms of tissue injury that magnify microvascular damage caused by hyperglycemia and hypertension. Many studies describe interruption of these mechanisms, without hemodynamic consequence, in the preservation of function in type 1 diabetes. Possible interactions between the renin–angiotensin–aldosterone system and physiologic glycemic control (through pulsatile insulin release suggest opportunities for further clinical investigation.Keywords: angiotensin-converting enzyme inhibitor, pulsatile insulin, diabetic nephropathy, cardiac autonomic neuropathy, podocytes, beta cells 

  6. Microvascular network topology of the human retinal vessels.

    Science.gov (United States)

    Schröder, S; Brab, M; Schmid-Schönbein, G W; Reim, M; Schmid-Schönbein, H

    1990-01-01

    A quantitative analysis of blood flow in the human retinal vessels requires a detailed picture of the microvascular network topology. In order to lay the foundation for a quantitative microcirculatory network analysis of the human retina, a novel technique for tissue preparation and network characterization was developed. After injection of hydrogen peroxide into the human bulb, the microvasculature was filled with oxygen produced by endothelial catalase and visualized after embedding in a mixture of cedar oil and gum damar. The vessel topology was documented in the form of photomicrographs, which permitted complete reconstruction of the microvasculature on transparent overlays. By considering the complete capillary system it was possible to divide the retinal network into dichotomous, asymmetric arteriolar and venular trees. The Strahler ordering method, which considers both dichotomous and side branching configurations, was selected and applied to analyze the retinal vascular trees, using the capillaries as the zero order reference vessels. The number of vessel segments was found to be an approximate logarithmic function of the order number, in accordance with Horton's law. Vessel lengths within each order were found to be log-normal distributed, and median lengths for different orders could be approximated by a 2nd degree polynomial curve. Diameters within each order could be approximated by a Gaussian distribution, and the mean values for different orders could be expressed by an exponential curve. These data provide the basis for conductance, pressure and flow computations within the retinal microvessels.

  7. The impact of early aerobic exercise on brain microvascular alterations induced by cerebral hypoperfusion.

    Science.gov (United States)

    Leardini-Tristão, Marina; Borges, Juliana Pereira; Freitas, Felipe; Rangel, Raquel; Daliry, Anissa; Tibiriçá, Eduardo; Estato, Vanessa

    2017-02-15

    The therapeutic potential of early exercise training following cerebral hypoperfusion was investigated on brain perfusion and inflammation in rats with permanent bilateral occlusion of the common carotid arteries (2VO). Wistar rats were subjected to 2VO or sham surgery and each group was then subdivided randomly into sedentary or exercise groups. Early exercise training was initiated after three days of 2VO or sham surgery and consisted of seven days of treadmill training (30min/day at ∼60% of maximal exercise test), composing four groups: 1) Sham sedentary (Sham-Sed), 2) Sham exercised (Sham-Ex), 3) 2VO sedentary (2VO-Sed) and 4) 2VO exercised (2VO-Ex). Microvascular cerebral blood flow (MCBF) and NADPH oxidase and eNOS gene expression were evaluated by laser speckle contrast imaging and RT-PCR, respectively, and brain functional capillary density and endothelial-leukocyte interactions were evaluated by fluorescence intravital video-microscopy. The 2VO-Sed group presented a decrease in MCBF (Sham-Sed: 230.9±12.2 vs. 2VO-Sed: 183.6±10.6 arbitrary perfusion units, Pexercise was able to prevent the cerebral microvascular inflammation by decreasing endothelial-leukocyte interactions (2VO-Ex: 0.9±0.3 vs. 2VO-Sed: 5±0.6cells/min/100μm, Pbrain NADPH oxidase gene expression (2VO-Ex: 1.7±0.1 arbitrary units, Pexercise may represent a means of preventing the microvascular alterations induced by chronic cerebral hypoperfusion.

  8. Plasma soluble urokinase-type plasminogen activator receptor level is independently associated with coronary microvascular function in patients with non-obstructive coronary artery disease

    DEFF Research Database (Denmark)

    Mekonnen, Girum; Corban, Michel T; Hung, Olivia Y;

    2015-01-01

    , medications profiles and hs-CRP, suPAR remained an independent predictor of CFR (B = -0.30, p = 0.04), indicating an independent association between suPAR level and coronary microvascular function. CONCLUSIONS: In this cross-sectional study, plasma suPAR level was an independent predictor of coronary......BACKGROUND: Soluble urokinase-type plasminogen activator receptor (suPAR) is a novel biomarker released from leukocytes and endothelial cells that has been associated with atherosclerotic cardiovascular disease. We hypothesized that plasma suPAR level is an independent predictor of coronary...... microvascular function. METHODS: Coronary blood flow velocity and plasma suPAR levels were evaluated in patients with non-obstructive coronary artery disease. Coronary flow reserve (CFR) was calculated as the ratio of hyperemic to basal average peak blood flow velocity and coronary microvascular dysfunction...

  9. Peptides Derived from Type IV Collagen, CXC Chemokines, and Thrombospondin-1 Domain-Containing Proteins Inhibit Neovascularization and Suppress Tumor Growth in MDA-MB-231 Breast Cancer Xenografts

    Directory of Open Access Journals (Sweden)

    Jacob E. Koskimaki

    2009-12-01

    Full Text Available Angiogenesis or neovascularization, the process of new blood vessel formation from preexisting microvasculature, involves interactions among several cell types including parenchymal, endothelial cells, and immune cells. The formation of new vessels is tightly regulated by a balance between endogenous proangiogenic and antiangiogenic factors to maintain homeostasis in tissue; tumor progression and metastasis in breast cancer have been shown to be angiogenesis-dependent. We previously introduced a systematic methodology to identify putative endogenous antiangiogenic peptides and validated these predictions in vitro in human umbilical vein endothelial cell proliferation and migration assays. These peptides are derived from several protein families including type IV collagen, CXC chemokines, and thrombospondin-1 domain-containing proteins. On the basis of the results from the in vitro screening, we have evaluated the ability of one peptide selected from each family named pentastatin-1, chemokinostatin-1, and properdistatin, respectively, to suppress angiogenesis in an MDA-MB-231 human breast cancer orthotopic xenograft model in severe combined immunodeficient mice. Peptides were administered intraperitoneally once per day. We have demonstrated significant suppression of tumor growth in vivo and subsequent reductions in microvascular density, indicating the potential of these peptides as therapeutic agents for breast cancer.

  10. Effect of the glucagon-like peptide-1 analogue liraglutide on coronary microvascular function in patients with type 2 diabetes – a randomized, single-blinded, cross-over pilot study

    DEFF Research Database (Denmark)

    Faber, Rebekka; Zander, Mette; Pena, Adam;

    2015-01-01

    dipyridamole induced stress. Peripheral microvascular endothelial function was assessed by Endo-PAT2000®. Interventions were compared by two-sample t-test after ensuring no carry over effect. RESULTS: A total of 24 patients were included. Twenty patients completed the study (15 male; mean age 57 ± 9; mean BMI...

  11. 超声光散射诊断乳腺癌及其与VEGF表达的相关性研究%Diffused optical tomography with ultrasonography in diagnosis of breast cancer and correlation with vascular endothelial growth factor

    Institute of Scientific and Technical Information of China (English)

    王红磊; 阙燕; 武力; 马大昌; 吴多明; 纪彩玉

    2011-01-01

    Objective: To investigate the correlation between synthesis diagnostic index (SDI) of diffused optical tomography(DOT) with ultrasonography(US) location detected preoperative and Vascular endothelial growth factor expression detected postoperative in breast cancer as well as their relationship with clinicopathological characteristics of breast cancer. Methods: Conventional DOT with US were performed on 131 cases of breast benign mass and 142 cases of breast cancer. SDI was measured. The receiver operating characteristic (ROC) curve analysis was used to estimate the optimal diagnostic threshold of breast cancer. The expression levels of VEGF was detected by SP immunohistochemistry. Analysis the correlation between SDI and the expression levels of VEGF. Analysis the correlation between SDI as well as the expression of HIF-1 α and clinicopathological characteristics of breast cancer. Results: The mean value of SDI was 109.35 ± 48.03 in the breast benign mass and 190.87 ±60.33 in the breast cancer. There were significant difference between them (P<0.01). According to ROC curve analysis, the areas under ROC curve of SDI was 0.889, the SDI value greater than 141.2 was an optimal diagnostic threshold of breast cancer. The SDI was significantly higher in VEGF-positive breast cancer than in VEGF- negative breast cancer(P<0.01).The mean value of SDI and the expression levels of VEGF were significantly correlated with the tumor size, lymph node status ,clinical TNM stage and the expression levels of cerbB-2 (P<0.01), while had no correlation with the expression levels of ER and PR(P>0.05). Conclusion: As a new technology of functional imaging, DOT with US location might play a great role of early diagnosis and distinguishing benign from malignant tumor.%目的:探讨乳腺癌超声光散射断层成像综合诊断指数(SDI)与血管内皮生长因子(VEGF)的关系以及它们与乳腺癌临床病理特征的关系.方法:采用超声光散射

  12. Anesthetic propofol overdose causes endothelial cytotoxicity in vitro and endothelial barrier dysfunction in vivo

    Energy Technology Data Exchange (ETDEWEB)

    Lin, Ming-Chung [Institute of Clinical Medicine, College of Medicine, National Cheng Kung University, Tainan, Taiwan (China); Department of Anesthesiology, Chi Mei Medical Center, Liouying, Tainan, Taiwan (China); Chen, Chia-Ling [Center of Infectious Disease and Signaling Research, College of Medicine, National Cheng Kung University, Tainan, Taiwan (China); Yang, Tsan-Tzu; Choi, Pui-Ching [Institute of Clinical Medicine, College of Medicine, National Cheng Kung University, Tainan, Taiwan (China); Hsing, Chung-Hsi [Department of Anesthesiology, Chi Mei Medical Center, Tainan, Taiwan (China); Department of Anesthesiology, College of Medicine, Taipei Medical University, Taipei, Taiwan (China); Lin, Chiou-Feng, E-mail: cflin@mail.ncku.edu.tw [Institute of Clinical Medicine, College of Medicine, National Cheng Kung University, Tainan, Taiwan (China); Center of Infectious Disease and Signaling Research, College of Medicine, National Cheng Kung University, Tainan, Taiwan (China); Department of Microbiology and Immunology, College of Medicine, National Cheng Kung University, Tainan, Taiwan (China); Institute of Basic Medical Sciences, College of Medicine, National Cheng Kung University, Tainan, Taiwan (China)

    2012-12-01

    An overdose and a prolonged treatment of propofol may cause cellular cytotoxicity in multiple organs and tissues such as brain, heart, kidney, skeletal muscle, and immune cells; however, the underlying mechanism remains undocumented, particularly in vascular endothelial cells. Our previous studies showed that the activation of glycogen synthase kinase (GSK)-3 is pro-apoptotic in phagocytes during overdose of propofol treatment. Regarding the intravascular administration of propofol, we therefore hypothesized that propofol overdose also induces endothelial cytotoxicity via GSK-3. Propofol overdose (100 μg/ml) inhibited growth in human arterial and microvascular endothelial cells. After treatment, most of the endothelial cells experienced caspase-independent necrosis-like cell death. The activation of cathepsin D following lysosomal membrane permeabilization (LMP) determined necrosis-like cell death. Furthermore, propofol overdose also induced caspase-dependent apoptosis, at least in part. Caspase-3 was activated and acted downstream of mitochondrial transmembrane potential (MTP) loss; however, lysosomal cathepsins were not required for endothelial cell apoptosis. Notably, activation of GSK-3 was essential for propofol overdose-induced mitochondrial damage and apoptosis, but not necrosis-like cell death. Intraperitoneal administration of a propofol overdose in BALB/c mice caused an increase in peritoneal vascular permeability. These results demonstrate the cytotoxic effects of propofol overdose, including cathepsin D-regulated necrosis-like cell death and GSK-3-regulated mitochondrial apoptosis, on endothelial cells in vitro and the endothelial barrier dysfunction by propofol in vivo. Highlights: ► Propofol overdose causes apoptosis and necrosis in endothelial cells. ► Propofol overdose triggers lysosomal dysfunction independent of autophagy. ► Glycogen synthase kinase-3 facilitates propofol overdose-induced apoptosis. ► Propofol overdose causes an increase

  13. Treatment of hemimasticatory spasm with microvascular decompression.

    Science.gov (United States)

    Wang, Yong-Nan; Dou, Ning-Ning; Zhou, Qiu-Meng; Jiao, Wei; Zhu, Jin; Zhong, Jun; Li, Shi-Ting

    2013-01-01

    Hemimasticatory spasm is a rare disorder characterized by paroxysmal involuntary contraction of the jaw-closing muscles. As the ideology and pathogenesis of the disease are still unclear, there has been no treatment that could give rise to a good outcome so far. Herein, we tried to use surgical management to cure the disease. Six patients with the disease were included in this study. These patients underwent microvascular decompression of the motor fibers of the trigeminal root. After the operation, all faces of the patients felt relaxed at varied degrees, except for 1 patient. Our study showed that microvascular decompression of the trigeminal nerve could lead to a better outcome. However, a control study with a large sample is needed before this technique is widely used.

  14. Fabrication, characterization, and modeling of microvascular composites

    Science.gov (United States)

    Ryan, Thomas J.

    Composite laminates of glass fiber and epoxy pre-preg were fabricated with microvascular channels. The channels were created using polylactic acid (PLA) filament that evaporates at a temperature of 392 °F (200 °C) above the resin cure temperature of 250 °F (121 °C). After the composite is cured, the panel was removed from the oven and allowed to cool to room temperature. The panel is then reheated to 392 °F to vaporize the filament, leaving a cylindrical channel. A microvascular channel can be used for withdrawing heat, damage detection and self-healing. However, increasing the temperatures of the laminate above the cure temperature of the resin causes excess cross linking, potentially decreasing the mechanical properties. Tensile and flexural mechanical tests were performed on composite specimens and tensile tests were performed on neat resin specimens. A three-dimensional finite element model (FEM) was developed to study the progressive deformation and damage mechanics under tensile loading. The load carrying capacity of the microvascular composite was shown to decrease by 40% from a standard composite material. This paper will present the details of the fabrication, characterization and modeling techniques that were used in this study.

  15. Expression of the pro-angiogenic factors vascular endothelial growth factor and interleukin-8/CXCL8 by human breast carcinomas is responsive to nutrient deprivation and endoplasmic reticulum stress

    Directory of Open Access Journals (Sweden)

    Abcouwer Steve F

    2004-01-01

    Full Text Available Abstract Background The expression of pro-angiogenic cytokines, such as vascular endothelial growth factor (VEGF and interleukin-8/CXCL8 (IL-8, plays an important role in tumor growth and metastasis. Low oxygen tension within poorly-vascularized tumors is thought to be the prime stimulus causing the secretion of VEGF. The expression of IL-8 by solid tumors is thought to be primarily due to intrinsic influences, such as constitutive activation of nuclear factor kappa B (NF-κB. However, VEGF expression is responsive to glucose deprivation, suggesting that low concentrations of nutrients other than oxygen may play a role in triggering the pro-angiogenic phenotype. Glucose deprivation causes endoplasmic reticulum (ER stress and alters gene expression through the unfolded protein response (UPR signaling pathway. A branch of the UPR, known as the ER overload response (EOR, can cause NF-κB activation. Thus, we hypothesized that treatments that cause ER stress and deprivation of other nutrients, such as amino acids, would trigger the expression of angiogenic cytokines by breast cancer cell lines. Results We found that glutamine deprivation and treatment with a chemical inducer of ER stress (tunicamycin caused a marked induction of the secretion of both VEGF and IL-8 protein by a human breast adenocarcinoma cell line (TSE cells. Glutamine deprivation, glucose deprivation and several chemical inducers of ER stress increased VEGF and IL-8 mRNA expression in TSE and other breast cancer cell lines cultured under both normoxic and hypoxic conditions, though hypoxia generally diminished the effects of glucose deprivation. Of all amino acids tested, ambient glutamine availability had the largest effect on VEGF and IL-8 mRNA expression. The induction of VEGF mRNA expression, but not IL-8, was sustained and closely corresponded with the upregulated expression of the ER stress-responsive genes glucose-regulated protein 78 (GRP78 and growth arrest and DNA damage

  16. Cryptogenic stroke following abdominal free flap breast reconstruction surgery

    OpenAIRE

    Huizhuang Xie; Malata, Charles M.

    2014-01-01

    INTRODUCTION: Abdominal free flap breast reconstruction is regarded as the gold standard method of post-mastectomy breast reconstruction by many. It is a major surgery which can be associated with varied systemic complications. To date, there have been no reports of cerebrovascular complications in the literature which examine the possible relation between thromboembolism and patent foramen ovale (PFO) in patients undergoing microvascular breast reconstruction. PRESENTATION OF CASE: A 54-y...

  17. Globotriaosylsphingosine accumulation and not alpha-galactosidase-A deficiency causes endothelial dysfunction in Fabry disease.

    Directory of Open Access Journals (Sweden)

    Mehdi Namdar

    Full Text Available BACKGROUND: Fabry disease (FD is caused by a deficiency of the lysosomal enzyme alpha-galactosidase A (GLA resulting in the accumulation of globotriaosylsphingosine (Gb3 in a variety of tissues. While GLA deficiency was always considered as the fulcrum of the disease, recent attention shifted towards studying the mechanisms through which Gb3 accumulation in vascular cells leads to endothelial dysfunction and eventually multiorgan failure. In addition to the well-described macrovascular disease, FD is also characterized by abnormalities of microvascular function, which have been demonstrated by measurements of myocardial blood flow and coronary flow reserve. To date, the relative importance of Gb3 accumulation versus GLA deficiency in causing endothelial dysfunction is not fully understood; furthermore, its differential effects on cardiac micro- and macrovascular endothelial cells are not known. METHODS AND RESULTS: In order to assess the effects of Gb3 accumulation versus GLA deficiency, human macro- and microvascular cardiac endothelial cells (ECs were incubated with Gb3 or silenced by siRNA to GLA. Gb3 loading caused deregulation of several key endothelial pathways such as eNOS, iNOS, COX-1 and COX-2, while GLA silencing showed no effects. Cardiac microvascular ECs showed a greater susceptibility to Gb3 loading as compared to macrovascular ECs. CONCLUSIONS: Deregulation of key endothelial pathways as observed in FD vasculopathy is likely caused by intracellular Gb3 accumulation rather than deficiency of GLA. Human microvascular ECs, as opposed to macrovascular ECs, seem to be affected earlier and more severely by Gb3 accumulation and this notion may prove fundamental for future progresses in early diagnosis and management of FD patients.

  18. Early systemic microvascular damage in pigs with atherogenic diabetes mellitus coincides with renal angiopoietin dysbalance

    NARCIS (Netherlands)

    M. Khairoun (Meriem); M.M. van den Heuvel (Mieke); B. van den Berg (Bernard); O. Sorop (Oana); De Boer, R. (Rients); N.S. van Ditzhuijzen (Nienke); I.M. Bajema (Ingeborg); H.J. Baelde; M. Zandbergen; D.J.G.M. Duncker (Dirk); T. Rabelink (Ton); M.E. Reinders (Marlies); W.J. van der Giessen (Wim); J.I. Rotmans (Joris)

    2015-01-01

    textabstractBackground: Diabetes mellitus (DM) is associated with a range of microvascular complications including diabetic nephropathy (DN). Microvascular abnormalities in the kidneys are common histopathologic findings in DN, which represent one manifestation of ongoing systemic microvascular dama

  19. Hypoxia-induced reactive oxygen species cause chromosomal abnormalities in endothelial cells in the tumor microenvironment.

    Directory of Open Access Journals (Sweden)

    Miyako Kondoh

    Full Text Available There is much evidence that hypoxia in the tumor microenvironment enhances tumor progression. In an earlier study, we reported abnormal phenotypes of tumor-associated endothelial cells such as those resistant to chemotherapy and chromosomal instability. Here we investigated the role of hypoxia in the acquisition of chromosomal abnormalities in endothelial cells. Tumor-associated endothelial cells isolated from human tumor xenografts showed chromosomal abnormalities, >30% of which were aneuploidy. Aneuploidy of the tumor-associated endothelial cells was also shown by simultaneous in-situ hybridization for chromosome 17 and by immunohistochemistry with anti-CD31 antibody for endothelial staining. The aneuploid cells were surrounded by a pimonidazole-positive area, indicating hypoxia. Human microvascular endothelial cells expressed hypoxia-inducible factor 1 and vascular endothelial growth factor A in response to either hypoxia or hypoxia-reoxygenation, and in these conditions, they acquired aneuploidy in 7 days. Induction of aneuploidy was inhibited by either inhibition of vascular endothelial growth factor signaling with vascular endothelial growth factor receptor 2 inhibitor or by inhibition of reactive oxygen species by N-acetyl-L-cysteine. These results indicate that hypoxia induces chromosomal abnormalities in endothelial cells through the induction of reactive oxygen species and excess signaling of vascular endothelial growth factor in the tumor microenvironment.

  20. Hypoxia-Induced Reactive Oxygen Species Cause Chromosomal Abnormalities in Endothelial Cells in the Tumor Microenvironment

    Science.gov (United States)

    Hida, Yasuhiro; Maishi, Nako; Towfik, Alam Mohammad; Inoue, Nobuo; Shindoh, Masanobu; Hida, Kyoko

    2013-01-01

    There is much evidence that hypoxia in the tumor microenvironment enhances tumor progression. In an earlier study, we reported abnormal phenotypes of tumor-associated endothelial cells such as those resistant to chemotherapy and chromosomal instability. Here we investigated the role of hypoxia in the acquisition of chromosomal abnormalities in endothelial cells. Tumor-associated endothelial cells isolated from human tumor xenografts showed chromosomal abnormalities, >30% of which were aneuploidy. Aneuploidy of the tumor-associated endothelial cells was also shown by simultaneous in-situ hybridization for chromosome 17 and by immunohistochemistry with anti-CD31 antibody for endothelial staining. The aneuploid cells were surrounded by a pimonidazole-positive area, indicating hypoxia. Human microvascular endothelial cells expressed hypoxia-inducible factor 1 and vascular endothelial growth factor A in response to either hypoxia or hypoxia-reoxygenation, and in these conditions, they acquired aneuploidy in 7 days. Induction of aneuploidy was inhibited by either inhibition of vascular endothelial growth factor signaling with vascular endothelial growth factor receptor 2 inhibitor or by inhibition of reactive oxygen species by N-acetyl-L-cysteine. These results indicate that hypoxia induces chromosomal abnormalities in endothelial cells through the induction of reactive oxygen species and excess signaling of vascular endothelial growth factor in the tumor microenvironment. PMID:24260373

  1. Association of Maternal Antiangiogenic Profile at Birth With Early Postnatal Loss of Microvascular Density in Offspring of Hypertensive Pregnancies

    Science.gov (United States)

    Yu, Grace Z.; Aye, Christina Y.L.; Lewandowski, Adam J.; Davis, Esther F.; Khoo, Cheen P.; Newton, Laura; Yang, Cheng T.; Al Haj Zen, Ayman; Simpson, Lisa J.; O’Brien, Kathryn; Cook, David A.; Granne, Ingrid; Kyriakou, Theodosios; Channon, Keith M.; Watt, Suzanne M.

    2016-01-01

    Offspring of hypertensive pregnancies are more likely to have microvascular rarefaction and increased blood pressure in later life. We tested the hypothesis that maternal angiogenic profile during a hypertensive pregnancy is associated with fetal vasculogenic capacity and abnormal postnatal microvascular remodeling. Infants (n=255) born after either hypertensive or normotensive pregnancies were recruited for quantification of postnatal dermal microvascular structure at birth and 3 months of age. Vasculogenic cell potential was assessed in umbilical vein endothelial cells from 55 offspring based on in vitro microvessel tube formation and proliferation assays. Maternal angiogenic profile (soluble fms-like tyrosine kinase-1, soluble endoglin, vascular endothelial growth factor, and placental growth factor) was measured from postpartum plasma samples to characterize severity of pregnancy disorder. At birth, offspring born after hypertensive pregnancy had similar microvessel density to those born after a normotensive pregnancy, but during the first 3 postnatal months, they had an almost 2-fold greater reduction in total vessel density (−17.7±16.4% versus −9.9±18.7%; P=0.002). This postnatal loss varied according to the vasculogenic capacity of the endothelial cells of the infant at birth (r=0.49; P=0.02). The degree of reduction in both in vitro and postnatal in vivo vascular development was proportional to levels of antiangiogenic factors in the maternal circulation. In conclusion, our data indicate that offspring born to hypertensive pregnancies have reduced vasculogenic capacity at birth that predicts microvessel density loss over the first 3 postnatal months. Degree of postnatal microvessel reduction is proportional to levels of antiangiogenic factors in the maternal circulation at birth. PMID:27456522

  2. Direct ink writing of microvascular networks

    Science.gov (United States)

    Wu, Willie

    Nature is replete with examples of embedded microvascular systems that enable efficient fluid flow and distribution for autonomic healing, cooling, and energy harvesting. The ability to incorporate microvascular networks in functional materials systems is therefore both scientifically and technologically important. In this PhD thesis, the direct-write assembly of planar and 3D biomimetic microvascular networks within polymer and hydrogel matrices is demonstrated. In addition, the influence of network design of fluid transport efficiency is characterized. Planar microvascular networks composed of periodic lattices of uniformal microchannels and hierarchical, branching architectures are constructed by direct-write assembly of a fugitive organic ink. Several advancements are required to facilitate their patterning, including pressure valving, dual ink printing, and dynamic pressure variation to allow tunable control of ink deposition. The hydraulic conductance is measured using a high pressure flow meter as a function of network design. For a constant vascular volume and areal coverage, 2- and 4-generation branched architectures that obey Murray's Law exhibited the highest hydraulic conductivity. These experimental observations are in good agreement with predictions made by analytic models. 3D microvascular networks are fabricated by omnidirectional printing a fugitive organic ink into a photopolymerizable hydrogel matrix that is capped with fluid filler of nearly identical composition. Using this approach, 3D networks of arbitrary design can be patterned. After ink deposition is complete, the matrix and fluid filler are chemically cross-linked via UV irradiation, and the ink is removed by liquefication. Aqueous solutions composed of a triblock copolymer of polyethylene oxide (PEO)-polypropylene oxide (PPO)-PEO constitute the materials system of choice due to their thermal- and concentration-dependent phase behavior. Specifically, the fugitive ink consists of a 23 w

  3. Glyoxalase-1 overexpression reduces endothelial dysfunction and attenuates early renal impairment in a rat model of diabetes

    DEFF Research Database (Denmark)

    Brouwers, Olaf; Niessen, Petra M G; Miyata, Toshio;

    2014-01-01

    AIMS/HYPOTHESIS: In diabetes, advanced glycation end-products (AGEs) and the AGE precursor methylglyoxal (MGO) are associated with endothelial dysfunction and the development of microvascular complications. In this study we used a rat model of diabetes, in which rats transgenically overexpressed...

  4. Advances in microvascular decompression for hemifacial spasm

    Institute of Scientific and Technical Information of China (English)

    Zhiqiang Cui; Zhipei Ling

    2015-01-01

    Primary hemifacial spasm (HFS) is a disorder that causes frequent involuntary contractions in the muscles on one side of the face, due to a blood vessel compressing the nerve at its root exit zone (REZ) from the brainstem. Numerous prospective and retrospective case series have confirmed the efficacy of microvascular decompression (MVD) of the facial nerve in patients with HFS. However, while MVD is effective, there are still significant postoperative complications. In this paper, recent technological advances related to MVD (such as lateral spread response, brainstem auditory evokes potential, three dimensional time of flight magnetic resonance angiography, intraoperative neuroendoscopy) are reviewed for the purposes of improving MVD treatment efficacy and reducing postoperative complications.

  5. Endothelial activation, lymphangiogenesis, and humoral rejection of kidney transplants.

    Science.gov (United States)

    Phillips, Sharon; Kapp, Meghan; Crowe, Deborah; Garces, Jorge; Fogo, Agnes B; Giannico, Giovanna A

    2016-05-01

    Antibody-mediated rejection (ABMR) is implicated in 45% of renal allograft failure and 57% of late allograft dysfunction. Peritubular capillary C4d is a specific but insensitive marker of ABMR. The 2013 Banff Conference ABMR revised criteria included C4d-negative ABMR with evidence of endothelial-antibody interaction. We hypothesized that endothelial activation and lymphangiogenesis are increased with C4d-negative ABMR and correlate with intragraft T-regulatory cells and T-helper 17. Seventy-four renal transplant biopsies were selected to include (a) ABMR with C4d Banff scores ≥2 (n = 35), (b) variable microvascular injury and C4d score 0-1 (n = 24), and (c) variable microvascular injury and C4d score = 0 (n = 15). Controls included normal preimplantation donor kidneys (n = 5). Immunohistochemistry for endothelial activation (P- and E-selectins [SEL]), lymphangiogenesis (D2-40), T-regulatory cells (FOXP3), and T-helper 17 (STAT3) was performed. Microvessel and inflammatory infiltrate density was assessed morphometrically in interstitium and peritubular capillaries. All transplants had significantly higher microvessel and lymph vessel density compared with normal. Increased expression of markers of endothelial activation predicted transplant glomerulopathy (P-SEL, P = .003). Increased P-SEL and D2-40 were associated with longer interval from transplant to biopsy (P = .005). All 3 markers were associated with increased interstitial fibrosis, tubular atrophy, and graft failure (P-SEL, P < .001; E-SEL, P = .0011; D2-40, P = .012). There was no association with the intragraft FOXP3/STAT3 ratio. We conclude that endothelial activation and lymphangiogenesis could represent a late response to injury leading to fibrosis and progression of kidney damage, and are independent of the intragraft FOXP3/STAT3 ratio. Our findings support the therapeutic potential of specifically targeting endothelial activation.

  6. Total ‘rib’-preservation technique of internal mammary vessel exposure for free flap breast reconstruction: A 5-year prospective cohort study and instructional video

    Directory of Open Access Journals (Sweden)

    Anais Rosich-Medina

    2015-09-01

    Discussion & conclusion: The total ‘rib’-preservation technique of IMV exposure is a safe, reliable and versatile method for microvascular breast reconstruction and should be considered as a valid alternative to the ‘rib’-sacrificing techniques.

  7. 糖基化终产物对脉络膜微血管内皮细胞增生和管腔形成的影响%Advanced glycation endproducts enhance proliferation, but not tube formation in choroidal microvascular endothelial cells

    Institute of Scientific and Technical Information of China (English)

    王雨生; Ulrike Friedrichs; Wolfram Eichler; Stephan Hoffmann; Peter Wiedemann

    2005-01-01

    ● AIM: To investigate the role of advanced glycation endproducts (AGEs) in the pathogenesis of age-related macular degeneration (AMD).● METHODS: Bovine choroidal endothelial cells (CEC:)were isolated by the modified protocol using lycopersicon esculentum agglutinin coated Dynabeads, and identiffed by immunocytochemical staining with anti-Factor Ⅷ antibody and uptaking of dil-acetylated low-density lipoprotein (dil-ac-LDL). AGEs were prepared by incubating 50g/L bovine serum albumin and 150g/L glucose at 37℃ for 6wk, which were characterized by dot blot assay with anti-AGEs antibody. CEC proliferation was evaluated using 3,(4,5-dimethylthiazol-2-yl)-2,5- diphenyl tetrazolium bromide (MTT) assay, and tube formation in CEC was determined by a Vitrogen system.● RESULTS: More than 90% of the cultured cells were positive to Factor V immunostaining and had the ability to uptake dil-ac-LDL, which were the features of endothelial cells. AGEs we prepared were affinitive to anti-AGEs antibody. After treatment with AGEs for a time course of 3d, CEC proliferation was significantly increased in a dose-dependent manner by AGEs at concentrations between 62.5 and 500mg/L. The cytokine,basic fibroblast growth factor (bFGF), enhanced strongly tube-like structure formation in CEC to 124% (P<0.05)above that of untreated controls. In this condition, AGEs at the concentrations of 500 and 50mg/L showed no effect on CEC tube formation (P>0.05).● CONCLUSION: The present study demonstrated that CEC proliferation was increased by AGEs, however, regarding there was no statistically effect on CEC tube formation. These findings confirm and extend that AGEs could be a potential initiator in the pathogenesis of choroidal neovascularization in exudative AMD, at least in part, through enhancement of CEC proliferation.%目的:研究糖基化终产物(advanced glycationendproducts,AGEs)在年龄相关性黄斑变性发生中的作用.方法:使用lycopersicon esculentum agglutinin包

  8. Pathophysiological roles of microvascular alterations in pulmonary inflammatory diseases: possible implications of tumor necrosis factor-alpha and CXC chemokines

    Directory of Open Access Journals (Sweden)

    Kanami Orihara

    2008-10-01

    Full Text Available Kanami Orihara, Akio MatsudaDepartment of Allergy and Immunology, National Research Institute for Child Health and Development, Tokyo, JapanAbstract: Chronic obstructive pulmonary disease (COPD and bronchial asthma are common respiratory diseases that are caused by chronic infl ammation of the airways. Although these diseases are mediated by substantially distinct immunological reactions, especially in mild cases, they both show increased numbers of neutrophils, increased production of tumor necrosis factor-alpha (TNF-α and poor responses to corticosteroids, particularly in patients with severe diseases. These immunological alterations may contribute strongly to airway structural changes, commonly referred to as airway remodeling. Microvascular alterations, a component of airway remodeling and caused by chronic inflammation, are observed and appear to be clinically involved in both diseases. It has been well established that vascular endothelial growth factor (VEGF plays important roles in the airway microvascular alterations in mild and moderate cases of both diseases, but any role that VEGF might play in severe cases of these diseases remains unclear. Here, we review recent research findings, including our own data, and discuss the possibility that TNF-α and its associated CXC chemokines play roles in microvascular alterations that are even more crucial than those of VEGF in patients with severe COPD or asthma.Keywords: TNF-α, CXC chemokines, corticosteroid, pulmonary microvessels, COPD, asthma

  9. Novel biomarkers of arterial and venous ischemia in microvascular flaps.

    Directory of Open Access Journals (Sweden)

    Gerard K Nguyen

    Full Text Available The field of reconstructive microsurgery is experiencing tremendous growth, as evidenced by recent advances in face and hand transplantation, lower limb salvage after trauma, and breast reconstruction. Common to all of these procedures is the creation of a nutrient vascular supply by microsurgical anastomosis between a single artery and vein. Complications related to occluded arterial inflow and obstructed venous outflow are not uncommon, and can result in irreversible tissue injury, necrosis, and flap loss. At times, these complications are challenging to clinically determine. Since early intervention with return to the operating room to re-establish arterial inflow or venous outflow is key to flap salvage, the accurate diagnosis of early stage complications is essential. To date, there are no biochemical markers or serum assays that can predict these complications. In this study, we utilized a rat model of flap ischemia in order to identify the transcriptional signatures of venous congestion and arterial ischemia. We found that the critical ischemia time for the superficial inferior epigastric fasciocutaneus flap was four hours and therefore performed detailed analyses at this time point. Histolgical analysis confirmed significant differences between arterial and venous ischemia. The transcriptome of ischemic, congested, and control flap tissues was deciphered by performing Affymetrix microarray analysis and verified by qRT-PCR. Principal component analysis revealed that arterial ischemia and venous congestion were characterized by distinct transcriptomes. Arterial ischemia and venous congestion was characterized by 408 and 1536>2-fold differentially expressed genes, respectively. qRT-PCR was used to identify five candidate genes Prol1, Muc1, Fcnb, Il1b, and Vcsa1 to serve as biomarkers for flap failure in both arterial ischemia and venous congestion. Our data suggests that Prol1 and Vcsa1 may be specific indicators of venous congestion and

  10. Endothelial nitric oxide synthase in the microcirculation.

    Science.gov (United States)

    Shu, Xiaohong; Keller, T C Stevenson; Begandt, Daniela; Butcher, Joshua T; Biwer, Lauren; Keller, Alexander S; Columbus, Linda; Isakson, Brant E

    2015-12-01

    Endothelial nitric oxide synthase (eNOS, NOS3) is responsible for producing nitric oxide (NO)--a key molecule that can directly (or indirectly) act as a vasodilator and anti-inflammatory mediator. In this review, we examine the structural effects of regulation of the eNOS enzyme, including post-translational modifications and subcellular localization. After production, NO diffuses to surrounding cells with a variety of effects. We focus on the physiological role of NO and NO-derived molecules, including microvascular effects on vessel tone and immune response. Regulation of eNOS and NO action is complicated; we address endogenous and exogenous mechanisms of NO regulation with a discussion of pharmacological agents used in clinical and laboratory settings and a proposed role for eNOS in circulating red blood cells.

  11. Establishment of an in vitro blood-brain barrier model by co-culturing rat brain microvascular endothelial cells,pericytes and astrocytes%大鼠脑微血管内皮细胞与周细胞、星形胶质细胞共培养建立体外血脑屏障模型

    Institute of Scientific and Technical Information of China (English)

    查雨锋; 傅晓钟; 张顺; 罗敏; 欧瑜; 董永喜; 王爱民; 王永林

    2015-01-01

    目的:应用原代培养的大鼠脑微血管内皮细胞(brain-microvessel endothelial cells,BMECs )与脑微血管周细胞(brain-microvessel pericytes,BMPC )、星形胶质细胞(astro-cytes,AS)共培养建立可模拟在体状态的体外血脑屏障(blood-brain barrier,BBB)模型。方法原代分离、纯化和培养大鼠BMECs、BMPC和AS,通过细胞形态学和免疫细胞化学染色方法鉴定原代培养的细胞,应用Millicell细胞培养插(孔径0.4μm)建立5种不同类型的体外BBB模型,经跨内皮电阻值(transendothelial electrical resistance,TEER)、荧光素钠通透性(sodium fluorescent,Na-FLU )、碱性磷酸酶(AKP)和γ-谷氨酰转肽酶(γ-GT1)的表达测定以及阳性药在体内和体外BBB通透量的相似性,比较评价其屏障功能。结果原代培养的BMECs呈典型的铺路卵石样结构,BMPC胞体较大且呈分枝状,AS 有细长突触,胞质较浅;免疫细胞化学染色证实原代细胞为目标细胞;BMECs与BMPC、AS共培养后TEER值可达(478±25)Ω·cm2,Na-FLU 的表观渗透系数为[(8.23±0.78)×10-6]cm·s-1,AKP和γ-GT1表达分别为(6.90±0.27)金氏单位· g-1 Pro,(4.39±0.32)μg·g-1 Pro;阳性药在体外BBB的表观渗透系数(apparent permeability coefficient,Papp )与在体数据具有较好的相关性(R2=0.92)。结论原代培养的大鼠BMECs与BMPC、AS共培养建立的体外BBB模型在形态、结构及屏障功能方面具备BBB的基本特征,为研究BBB的生理学、病理学以及筛选化合物提供了一种有用工具。%Aim To establish in vitro blood-brain barrier (BBB) model with characteristics of simulation of in vivo BBB by primi-tive co-culture of brain-microvessel endothelial cells (BMECs) with brain-microvessel pericytes (BMPC)and astrocytes (AS). Methods BMECs,BMPC and AS from SD rats were primitively isolated,purified and cultured,and then

  12. Microvascularization on collared peccary placenta: a microvascular cast study [corrected] in late pregnancy.

    Science.gov (United States)

    Santos, Tatiana Carlesso; Oliveira, Moacir Franco; Dantzer, Vibeke; Miglino, Maria Angélica

    2012-07-01

    The microvascularization of the collared peccary (Tayassu tajacu) placenta was studied by vascular casts and immunolocalization of α-smooth muscle actin and vimentin, to identify the three dimensional organization and vascular flow interrelation in the microvasculature between the maternal and fetal compartments of the placentae. The immunolocalization of vimentin in the vascular endothelium and in the smooth muscle cells of blood vessels showed indented capillaries along the uterine epithelium and the trophoblast at the sides of complementary maternal and fetal microfolds, or rugae. This confers the three-dimensional structure observed in vascular casts. On the maternal side, casts demonstrated uterine folds coated by with primary and secondary ridges, and by areolae dispersed between these ridges. The arteriole runs through the center/middle of ridges, branching at the top into a microvascular network wall in a basket-like fashion. At the base of these baskets venules were formed. On the fetal side, arterioles branched centrally in the fetal rugae into a capillary network in a bulbous form, complementary to the opposite maternal depressions forming the baskets. At the base of the bulbous protrusions, the fetal venules arise. The blood vessel orientation in the materno-fetal interface of the placentae of collared peccaries suggests a blood flow pattern of the type countercurrent to cross current. The same pattern has been reported in domestic swine demonstrating that, even after 38 million years, the Tayassuidae and Suidae families exhibit similar placental morphology, which is here characterized at the microvascular level.

  13. Phase transition of the microvascular network architecture in human pathologies.

    Science.gov (United States)

    Bianciardi, Giorgio; Traversi, Claudio; Cattaneo, Ruggero; De Felice, Claudia; Monaco, Annalisa; Tosi, Gianmarco; Parrini, Stefano; Latini, Giuseppe

    2012-01-01

    We have investigated the microvascular pattern in acquired or genetic diseases in humans. The lower gingival and vestibular oral mucosa, as well as the optic nerve head, was chosen to characterize the vascular pattern complexity due to the simple accessibility and visibility Local fractal dimensions, fractal dimension of the minimum path and Lempel-Ziv complexity have been used as operational numerical tools to characterize the microvascular networks. In the normal healthy subjects microvascular networks show nonlinear values corresponding to the complexity of a diffusion limited aggregation (DLA) model, while in several acquired or genetic diseases they are approaching the ones of an invasion percolation model.

  14. Self-healing materials with microvascular networks.

    Science.gov (United States)

    Toohey, Kathleen S; Sottos, Nancy R; Lewis, Jennifer A; Moore, Jeffrey S; White, Scott R

    2007-08-01

    Self-healing polymers composed of microencapsulated healing agents exhibit remarkable mechanical performance and regenerative ability, but are limited to autonomic repair of a single damage event in a given location. Self-healing is triggered by crack-induced rupture of the embedded capsules; thus, once a localized region is depleted of healing agent, further repair is precluded. Re-mendable polymers can achieve multiple healing cycles, but require external intervention in the form of heat treatment and applied pressure. Here, we report a self-healing system capable of autonomously repairing repeated damage events. Our bio-inspired coating-substrate design delivers healing agent to cracks in a polymer coating via a three-dimensional microvascular network embedded in the substrate. Crack damage in the epoxy coating is healed repeatedly. This approach opens new avenues for continuous delivery of healing agents for self-repair as well as other active species for additional functionality.

  15. Mandibular reconstruction with composite microvascular tissue transfer

    Energy Technology Data Exchange (ETDEWEB)

    Coleman, J.J. III; Wooden, W.A. (Emory Univ. School of Medicine, Atlanta, GA (USA))

    1990-10-01

    Microvascular free tissue transfer has provided a variety of methods of restoring vascularized bone and soft tissue to difficult defects created by tumor resection and trauma. Over 7 years, 26 patients have undergone 28 free flaps for mandibular reconstruction, 15 for primary squamous cell carcinoma of the floor of the mouth or tongue, 7 for recurrent tumor, and 6 for other reasons (lymphangioma (1), infection (1), gunshot wound (1), and osteoradionecrosis (3)). Primary reconstruction was performed in 19 cases and secondary in 9. All repairs were composite flaps including 12 scapula, 5 radial forearm, 3 fibula, 2 serratus, and 6 deep circumflex iliac artery. Mandibular defects included the symphysis alone (7), symphysis and body (5), symphysis-body-ramus condyle (2), body or ramus (13), and bilateral body (1). Fourteen patients had received prior radiotherapy to adjuvant or curative doses. Eight received postoperative radiotherapy. All patients had initially successful vascularized reconstruction by clinical examination (28) and positive radionuclide scan (22 of 22). Bony stability was achieved in 25 of 26 patients and oral continence in 24 of 26. One complete flap loss occurred at 14 days. Complications of some degree developed in 22 patients including partial skin necrosis (3), orocutaneous fistula (3), plate exposure (1), donor site infection (3), fracture of reconstruction (1), and fracture of the radius (1). Microvascular transfer of bone and soft tissue allows a reliable reconstruction--despite previous radiotherapy, infection, foreign body, or surgery--in almost every situation in which mandible and soft tissue are absent. Bony union, a healed wound, and reasonable function and appearance are likely despite early fistula, skin loss, or metal plate or bone exposure.

  16. Acute cocoa flavanol supplementation improves muscle macro- and microvascular but not anabolic responses to amino acids in older men.

    Science.gov (United States)

    Phillips, Bethan E; Atherton, Philip J; Varadhan, Krishna; Limb, Marie C; Williams, John P; Smith, Kenneth

    2016-05-01

    The anabolic effects of nutrition on skeletal muscle may depend on adequate skeletal muscle perfusion, which is impaired in older people. Cocoa flavanols have been shown to improve flow-mediated dilation, an established measure of endothelial function. However, their effect on muscle microvascular blood flow is currently unknown. Therefore, the objective of this study was to explore links between the consumption of cocoa flavanols, muscle microvascular blood flow, and muscle protein synthesis (MPS) in response to nutrition in older men. To achieve this objective, leg blood flow (LBF), muscle microvascular blood volume (MBV), and MPS were measured under postabsorptive and postprandial (intravenous Glamin (Fresenius Kabi, Germany), dextrose to sustain glucose ∼7.5 mmol·L(-1)) conditions in 20 older men. Ten of these men were studied with no cocoa flavanol intervention and a further 10 were studied with the addition of 350 mg of cocoa flavanols at the same time that nutrition began. Leg (femoral artery) blood flow was measured by Doppler ultrasound, muscle MBV by contrast-enhanced ultrasound using Definity (Lantheus Medical Imaging, Mass., USA) perflutren contrast agent and MPS using [1, 2-(13)C2]leucine tracer techniques. Our results show that although older individuals do not show an increase in LBF or MBV in response to feeding, these absent responses are apparent when cocoa flavanols are given acutely with nutrition. However, this restoration in vascular responsiveness is not associated with improved MPS responses to nutrition. We conclude that acute cocoa flavanol supplementation improves muscle macro- and microvascular responses to nutrition, independently of modifying muscle protein anabolism.

  17. Very rapid effect of pitavastatin on microvascular function in comparison to rosuvastatin: reactive hyperemia peripheral arterial tonometric study

    Directory of Open Access Journals (Sweden)

    Kono Y

    2013-05-01

    Full Text Available Yasushi Kono,1 Shota Fukuda,1 Kenei Shimada,2 Koki Nakanishi,2 Kenichiro Otsuka,2 Tomoichiro Kubo,1 Satoshi Jissho,1 Haruyuki Taguchi,1 Junichi Yoshikawa,3 Minoru Yoshiyama21Department of Medicine, Osaka Ekisaikai Hospital, Nishi-ku, Osaka, Japan; 2Department of Internal Medicine and Cardiology, Osaka City University School of Medicine, Abeno-ku, Osaka, Japan; 3Nishinomiya Watanabe Cardiovascular Center, Nishinomiya, JapanBackground: It has been reported that pitavastatin improves endothelial function faster than other statins. Recently introduced reactive hyperemia peripheral arterial tonometry (RH-PAT provides objective and quantitative assessment of peripheral microvascular function.Purpose: This study aimed to investigate whether peripheral microvascular function improved 2 hours after pitavastatin in subjects with coronary artery disease (CAD using RH-PAT, and the results were compared with those of rosuvastatin.Methods: This study included 94 subjects with CAD, assigned to a group given 2 mg of pitavastatin (n = 36, a group given 2.5 mg of rosuvastatin (n = 38, and a control group (n = 20. RH-PAT examinations were performed before and 2 hours after statin administration.Results: The RH-PAT index increased 2 hours after pitavastatin administration from 1.82 ± 0.45 to 2.16 ± 0.62 (P = 0.02, whereas there were no differences in the RH-PAT index in the rosuvastatin group (1.79 ± 0.71 to 1.91 ± 0.53, P = 0.09 and the control group (1.68 ± 0.36 to 1.84 ± 0.58, P = 0.4. No significant changes were observed at 2 hours in serum cholesterol levels in each group.Conclusion: The present study demonstrated that peripheral microvascular function improved 2 hours after a single clinical dose of pitavastatin, but not after rosuvastatin.Keywords: coronary artery disease, statin, microvascular function

  18. The endothelial glycocalyx promotes homogenous blood flow distribution within the microvasculature.

    Science.gov (United States)

    McClatchey, P Mason; Schafer, Michal; Hunter, Kendall S; Reusch, Jane E B

    2016-07-01

    Many common diseases involve impaired tissue perfusion, and heterogeneous distribution of blood flow in the microvasculature contributes to this pathology. The physiological mechanisms regulating homogeneity/heterogeneity of microvascular perfusion are presently unknown. Using established empirical formulations for blood viscosity modeling in vivo (blood vessels) and in vitro (glass tubes), we showed that the in vivo formulation predicts more homogenous perfusion of microvascular networks at the arteriolar and capillary levels. Next, we showed that the more homogeneous blood flow under simulated in vivo conditions can be explained by changes in red blood cell interactions with the vessel wall. Finally, we demonstrated that the presence of a space-filling, semipermeable layer (such as the endothelial glycocalyx) at the vessel wall can account for the changes of red blood cell interactions with the vessel wall that promote homogenous microvascular perfusion. Collectively, our results indicate that the mechanical properties of the endothelial glycocalyx promote homogeneous microvascular perfusion. Preservation or restoration of normal glycocalyx properties may be a viable strategy for improving tissue perfusion in a variety of diseases.

  19. Divergent responses of different endothelial cell types to infection with Candida albicans and Staphylococcus aureus.

    Directory of Open Access Journals (Sweden)

    Kati Seidl

    Full Text Available Endothelial cells are important in the pathogenesis of bloodstream infections caused by Candida albicans and Staphylococcus aureus. Numerous investigations have used human umbilical vein endothelial cells (HUVECs to study microbial-endothelial cell interactions in vitro. However, the use of HUVECs requires a constant supply of umbilical cords, and there are significant donor-to-donor variations in these endothelial cells. The use of an immortalized endothelial cell line would obviate such difficulties. One candidate in this regard is HMEC-1, an immortalized human dermal microvascular endothelial cell line. To determine if HMEC-1 cells are suitable for studying the interactions of C. albicans and S. aureus with endothelial cells in vitro, we compared the interactions of these organisms with HMEC-1 cells and HUVECs. We found that wild-type C. albicans had significantly reduced adherence to and invasion of HMEC-1 cells as compared to HUVECs. Although wild-type S. aureus adhered to and invaded HMEC-1 cells similarly to HUVECs, an agr mutant strain had significantly reduced invasion of HMEC-1 cells, but not HUVECs. Furthermore, HMEC-1 cells were less susceptible to damage induced by C. albicans, but more susceptible to damage caused by S. aureus. In addition, HMEC-1 cells secreted very little IL-8 in response to infection with either organism, whereas infection of HUVECs induced substantial IL-8 secretion. This weak IL-8 response was likely due to the anatomic site from which HMEC-1 cells were obtained because infection of primary human dermal microvascular endothelial cells with C. albicans and S. aureus also induced little increase in IL-8 production above basal levels. Thus, C. albicans and S. aureus interact with HMEC-1 cells in a substantially different manner than with HUVECs, and data obtained with one type of endothelial cell cannot necessarily be extrapolated to other types.

  20. Effects of intracellular acidosis on endothelial function: an overview.

    Science.gov (United States)

    Crimi, Ettore; Taccone, Fabio Silvio; Infante, Teresa; Scolletta, Sabino; Crudele, Valeria; Napoli, Claudio

    2012-04-01

    The endothelium represents the largest functional organ in the human body playing an active role in vasoregulation, coagulation, inflammation, and microvascular permeability. Endothelium contributes to maintain vascular integrity, intravascular volume, and tissue oxygenation promoting inflammatory network response for local defense and repair. Acid-basis homeostasis is an important physiologic parameter that controls cell function, and changes in pH can influence vascular tone by regulating endothelium and vascular smooth muscle cells. This review presents a current perspective of the effects of intracellular acidosis on the function and the basic regulatory mechanisms of endothelial cells.

  1. Selenate Enhances STAT3 Transcriptional Activity in Endothelial Cells: Differential Actions of Selenate and Selenite on LIF Cytokine Signaling and Cell Viability

    OpenAIRE

    Alturkmani, Hani J; Zgheib, Carlos; Zouein, Fouad A.; Alshaaer, Nour Eddin F.; Kurdi, Mazen; Booz, George W.

    2012-01-01

    Sodium selenate may have utility in treating Alzheimer’s disease and diabetes; however, its impact on the associated proinflammatory cytokine signaling of endothelial cells has not been investigated. We report that treatment of human microvascular endothelial cells with sodium selenate at a pharmacological dose (100 μM) enhanced tyrosine phosphorylation of nuclear STAT3 on Y705 in response to IL-6-type cytokine, leukemia inhibitory factor (LIF), indicative of enhanced STAT3 activity. Accordin...

  2. Effects of sodium and potassium supplementation on endothelial function: a fully controlled dietary intervention study.

    Science.gov (United States)

    Gijsbers, Lieke; Dower, James I; Schalkwijk, Casper G; Kusters, Yvo H A M; Bakker, Stephan J L; Hollman, Peter C H; Geleijnse, Johanna M

    2015-11-14

    High Na and low K intakes have adverse effects on blood pressure, which increases the risk for CVD. The role of endothelial dysfunction and inflammation in this pathophysiological process is not yet clear. In a randomised placebo-controlled cross-over study in untreated (pre)hypertensives, we examined the effects of Na and K supplementation on endothelial function and inflammation. During the study period, subjects were provided with a diet that contained 2·4 g/d of Na and 2·3 g/d of K for a 10 460 kJ (2500 kcal) intake. After 1-week run-in, subjects received capsules with supplemental Na (3·0 g/d), supplemental K (2·8 g/d) or placebo, for 4 weeks each, in random order. After each intervention, circulating biomarkers of endothelial function and inflammation were measured. Brachial artery flow-mediated dilation (FMD) and skin microvascular vasomotion were assessed in sub-groups of twenty-two to twenty-four subjects. Of thirty-seven randomised subjects, thirty-six completed the study. Following Na supplementation, serum endothelin-1 was increased by 0·24 pg/ml (95 % CI 0·03, 0·45), but no change was seen in other endothelial or inflammatory biomarkers. FMD and microvascular vasomotion were unaffected by Na supplementation. K supplementation reduced IL-8 levels by 0·28 pg/ml (95 % CI 0·03, 0·53), without affecting other circulating biomarkers. FMD was 1·16 % (95% CI 0·37, 1·96) higher after K supplementation than after placebo. Microvascular vasomotion was unaffected. In conclusion, a 4-week increase in Na intake increased endothelin-1, but had no effect on other endothelial or inflammatory markers. Increased K intake had a beneficial effect on FMD and possibly IL-8, without affecting other circulating endothelial or inflammatory biomarkers.

  3. Sulfhydryl angiotensin-converting enzyme inhibitor promotes endothelial cell survival through nitric-oxide synthase, fibroblast growth factor-2, and telomerase cross-talk.

    Science.gov (United States)

    Donnini, Sandra; Terzuoli, Erika; Ziche, Marina; Morbidelli, Lucia

    2010-03-01

    The protective effect exerted by angiotensin-converting enzyme inhibitors (ACEI) in cardiovascular diseases caused by endothelial injury and aging has been attributed to the restoration of endothelial cell functions. Recently, we demonstrated a central role of the fibroblast growth factor-2 (FGF-2)/FGF receptor-1 system in mediating the acquisition of an angiogenic phenotype in coronary microvascular endothelium exposed to ACEI. Here, we report on the rescuing effect of ACEI on impaired endothelium and the intracellular signaling mechanisms that lead endothelial cells to enter apoptosis and to senesce. Conditions mimicking pathological cell damage (serum deprivation) lead to endothelial apoptosis as evidenced by increased caspase-3 activity. ACEI enhanced cell survival through activation of prosurvival and antiaging signals involving Akt phosphorylation, endothelial nitric-oxide synthase (eNOS) expression and activation, FGF-2 and telomerase catalytic subunit (TERT) up-regulation, and delayed senescence. In microvascular endothelial cells exposed to ACEI, Akt/eNOS pathway-dependent FGF-2 was necessary for gene transcription of TERT. These protective effects were particularly evident for sulfhydryl-containing ACEI (zofenoprilat), which were reported to exhibit potent antioxidant effects. In conclusion, ACEI with antioxidant properties up-regulate eNOS, FGF-2, and TERT mRNA, which favor endothelial cell survival and prolong their lifespan, thus restoring endothelial cell functions after vascular damage. These effects could explain the beneficial effects of these drugs in various cardiovascular diseases associated with endothelial injury and aging.

  4. Clinical outcome following micro-vascular decompression for trigeminal neuralgia

    Directory of Open Access Journals (Sweden)

    Godugu Bhaskar Rao

    2015-07-01

    Conclusion: Micro-vascular decompression is safe and effective in producing good pain relief over a long term in patients with Trigeminal neuralgias refractive to medical treatment. [Int J Res Med Sci 2015; 3(7.000: 1741-1744

  5. Fibroblast nemosis induces angiogenic responses of endothelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Enzerink, Anna, E-mail: anna.enzerink@helsinki.fi [Haartman Institute, University of Helsinki, P.O. BOX 21, FIN-00014 Helsinki (Finland); Rantanen, Ville, E-mail: ville.rantanen@helsinki.fi [Computational Systems Biology Laboratory, Institute of Biomedicine and Genome-Scale Biology Research Program, University of Helsinki, P.O. BOX 63, 00014 Helsinki (Finland); Vaheri, Antti, E-mail: antti.vaheri@helsinki.fi [Haartman Institute, University of Helsinki, P.O. BOX 21, FIN-00014 Helsinki (Finland)

    2010-03-10

    Increasing evidence points to a central link between inflammation and activation of the stroma, especially of fibroblasts therein. However, the mechanisms leading to such activation mostly remain undescribed. We have previously characterized a novel type of fibroblast activation (nemosis) where clustered fibroblasts upregulated the production of cyclooxygenase-2, secretion of prostaglandins, proteinases, chemotactic cytokines, and hepatocyte growth factor (HGF), and displayed activated nuclear factor-{kappa}B. Now we show that nemosis drives angiogenic responses of endothelial cells. In addition to HGF, nemotic fibroblasts secreted vascular endothelial growth factor (VEGF), and conditioned medium from spheroids promoted sprouting and networking of human umbilical venous endothelial cells (HUVEC). The response was partly inhibited by function-blocking antibodies against HGF and VEGF. Conditioned nemotic fibroblast medium promoted closure of HUVEC and human dermal microvascular endothelial cell monolayer wounds, by increasing the motility of the endothelial cells. Wound closure in HUVEC cells was partly inhibited by the antibodies against HGF. The stromal microenvironment regulates wound healing responses and often promotes tumorigenesis. Nemosis offers clues to the activation process of stromal fibroblasts and provides a model to study the part they play in angiogenesis-related conditions, as well as possibilities for therapeutical approaches desiring angiogenesis in tissue.

  6. Loss of the endothelial glycocalyx links albuminuria and vascular dysfunction.

    Science.gov (United States)

    Salmon, Andrew H J; Ferguson, Joanne K; Burford, James L; Gevorgyan, Haykanush; Nakano, Daisuke; Harper, Steven J; Bates, David O; Peti-Peterdi, Janos

    2012-08-01

    Patients with albuminuria and CKD frequently have vascular dysfunction but the underlying mechanisms remain unclear. Because the endothelial surface layer, a meshwork of surface-bound and loosely adherent glycosaminoglycans and proteoglycans, modulates vascular function, its loss could contribute to both renal and systemic vascular dysfunction in proteinuric CKD. Using Munich-Wistar-Fromter (MWF) rats as a model of spontaneous albuminuric CKD, multiphoton fluorescence imaging and single-vessel physiology measurements revealed that old MWF rats exhibited widespread loss of the endothelial surface layer in parallel with defects in microvascular permeability to both water and albumin, in both continuous mesenteric microvessels and fenestrated glomerular microvessels. In contrast to young MWF rats, enzymatic disruption of the endothelial surface layer in old MWF rats resulted in neither additional loss of the layer nor additional changes in permeability. Intravenous injection of wheat germ agglutinin lectin and its adsorption onto the endothelial surface layer significantly improved glomerular albumin permeability. Taken together, these results suggest that widespread loss of the endothelial surface layer links albuminuric kidney disease with systemic vascular dysfunction, providing a potential therapeutic target for proteinuric kidney disease.

  7. OK432优化的内皮细胞疫苗抗小鼠乳腺癌作用研究%Anti-tumor effects of OK432 optimized umbilical vein endothelial cell vaccine in murine breast cancer

    Institute of Scientific and Technical Information of China (English)

    徐茂磊; 周玲; 杨小平

    2014-01-01

    目的:探讨 OK432优化的人脐静脉内皮细胞( HU-VECs)疫苗对小鼠EAC乳腺癌的生长抑制作用。方法体外培养HUVECs,与佐剂OK432混合,制备HUVECs-OK432疫苗,以小鼠 EAC 乳腺癌皮下移植瘤模型考查 HUVECs-OK432疫苗的抗肿瘤效应,并通过ELISA、脾细胞增殖及细胞毒性T淋巴细胞( CTL)杀伤实验检测疫苗免疫后体液及细胞免疫应答水平。结果在预防性免疫中, HUVECs-OK432疫苗可以明显抑制EAC乳腺癌生长;HUVECs-OK432疫苗诱导小鼠产生了高滴度的特异性 HUVEC 抗体;HU-VECs-OK432疫苗能有效刺激免疫小鼠脾淋巴细胞的增殖;HUVECs-OK432组小鼠脾细胞诱导产生了明显的靶向HU-VEC细胞的CTL杀伤作用。结论 HUVECs-OK432疫苗可以有效诱导机体产生靶向HUVEC的特异性体液及细胞免疫应答,从而有效抑制了小鼠EAC乳腺癌的生长。%Aim To investigate the anti-EAC breast cancer effects of viable human umbilical vein endothe-lial cells ( HUVECs) vaccine. Methods Subconflu-ent HUVECs were mixed with OK432 to prepare HU-VECs-OK432 vaccine, and the anti-tumor efficacy of HUVECs-OK432 was investigated using a subcutaneous tumor model of EAC breast cancer. ELISA, splenic lymphocyte proliferation assay and cytotoxic T lympho-cytes ( CTL ) killing assay were adopted to detect the humoral and cellular immune responses after HUVECs-OK432 immunization. Results HUVECs-OK432 im-munization significantly inhibited the growth of EAC tumor in mice in the prophylactic procedures. High ti-ter of anti-HUVEC antibody was elicited by HUVECs-OK432 immunization. The proliferation activity of splenocytes from mice immunized with HUVECs-OK432 was significantly increased. T lymphocytes iso-lated from HUVECs-OK432-immunized mice were dose dependently killing HUVECs in vitro. Conclusion Strong humoral and cellular immune responses targeting HUVEC are elicited by HUVECs-OK432 immuniza-tion, which results in a significant inhibition on the growth

  8. Gene delivery into primary brain capillary endothelial cells for protein secretion

    DEFF Research Database (Denmark)

    Larsen, Annette Burkhart; Thomsen, Louiza Bohn; Lichota, Jacek;

    model was established by co-culturing primary BCECs together with primary astrocytes, both of which were isolated from rats. This was done in order to study the possibility of using gene transfection in an environment closer to the in-vivo BBB situation. The in-vitro BBB barrier model showed trans...... for recombinant protein production. We have previously shown that it is possible to transfect human microvascular endothelial cells (HBMEC) and rat brain endothelial (RBE4) cells with genetic material encoding the human growth hormone (GH1) (Thomsen et al 2011). In the present study, however, an in-vitro BBB...

  9. Gene delivery of therapeutic polypeptides into brain capillary endothelial cells for protein secretion

    DEFF Research Database (Denmark)

    Larsen, Annette Burkhart; Thomsen, Louiza Bohn; Moos, Torben;

    has been to investigate the usage of BCEC as factories for recombinant protein production. A non-viral gene carrier was prepared from pullulan-spermine conjugated with plasmid DNA (Thomsen et al., 2011). In vitro transfection of Rat Brain Endothelial Cells (RBE4) and Human Brain Microvascular...... Endothelial cells (HBMECs) were conducted with three plasmids bearing cDNA encoding human BDNF, EPO or the FGL peptide. Results revealed a high expression of BDNF, EPO and FGL transcripts in transfected cells compared to the non-transfected cells, which strongly suggest that transfection were successful...

  10. MALT1 Protease Activation Triggers Acute Disruption of Endothelial Barrier Integrity via CYLD Cleavage.

    Science.gov (United States)

    Klei, Linda R; Hu, Dong; Panek, Robert; Alfano, Danielle N; Bridwell, Rachel E; Bailey, Kelly M; Oravecz-Wilson, Katherine I; Concel, Vincent J; Hess, Emily M; Van Beek, Matthew; Delekta, Phillip C; Gu, Shufang; Watkins, Simon C; Ting, Adrian T; Gough, Peter J; Foley, Kevin P; Bertin, John; McAllister-Lucas, Linda M; Lucas, Peter C

    2016-09-27

    Microvascular endothelial cells maintain a tight barrier to prevent passage of plasma and circulating immune cells into the extravascular tissue compartment, yet endothelial cells respond rapidly to vasoactive substances, including thrombin, allowing transient paracellular permeability. This response is a cornerstone of acute inflammation, but the mechanisms responsible are still incompletely understood. Here, we demonstrate that thrombin triggers MALT1 to proteolytically cleave cylindromatosis (CYLD). Fragmentation of CYLD results in microtubule disruption and a cascade of events leading to endothelial cell retraction and an acute permeability response. This finding reveals an unexpected role for the MALT1 protease, which previously has been viewed mostly as a driver of pro-inflammatory NF-κB signaling in lymphocytes. Thus, MALT1 not only promotes immune cell activation but also acutely regulates endothelial cell biology, actions that together facilitate tissue inflammation. Pharmacologic inhibition of MALT1 may therefore have synergistic impact by targeting multiple disparate steps in the overall inflammatory response.

  11. MALT1 Protease Activation Triggers Acute Disruption of Endothelial Barrier Integrity via CYLD Cleavage

    Directory of Open Access Journals (Sweden)

    Linda R. Klei

    2016-09-01

    Full Text Available Microvascular endothelial cells maintain a tight barrier to prevent passage of plasma and circulating immune cells into the extravascular tissue compartment, yet endothelial cells respond rapidly to vasoactive substances, including thrombin, allowing transient paracellular permeability. This response is a cornerstone of acute inflammation, but the mechanisms responsible are still incompletely understood. Here, we demonstrate that thrombin triggers MALT1 to proteolytically cleave cylindromatosis (CYLD. Fragmentation of CYLD results in microtubule disruption and a cascade of events leading to endothelial cell retraction and an acute permeability response. This finding reveals an unexpected role for the MALT1 protease, which previously has been viewed mostly as a driver of pro-inflammatory NF-κB signaling in lymphocytes. Thus, MALT1 not only promotes immune cell activation but also acutely regulates endothelial cell biology, actions that together facilitate tissue inflammation. Pharmacologic inhibition of MALT1 may therefore have synergistic impact by targeting multiple disparate steps in the overall inflammatory response.

  12. The angiogenic gene profile of circulating endothelial progenitor cells from ischemic stroke patients

    Directory of Open Access Journals (Sweden)

    Navarro-Sobrino Míriam

    2013-02-01

    Full Text Available Abstract Background The identification of circulating endothelial progenitor cells (EPCs has introduced new possibilities for cell-based treatments for stroke. We tested the angiogenic gene expression of outgrowth endothelial cells (OECs, an EPC subtype capable to shape vessel structures. Methods OECs (at colony or mature stages from ischemic stroke patients (n=8 were characterized using the RT2 ProfilerTM human angiogenesis PCR Array, and human microvascular endothelial cells (hCMEC/D3 were used as an expression reference of endothelial cells. Results Colony-OECs showed higher expression of CCL2, ID3, IGF-1, MMP9, TGFBR1, TNFAIP2, TNF and TGFB1. However, BAI-1, NRP2, THBS1, MMP2 and VEGFC expression was increased in mature-OECs (p Conclusion Our study shows that OECs from stroke patients present higher levels of pro-angiogenic factors at early stages, decreasing in mature OECs when they become more similar to mature microvascular endothelial cells.

  13. IL-17A potentiates TNFα-induced secretion from human endothelial cells and alters barrier functions controlling neutrophils rights of passage

    DEFF Research Database (Denmark)

    Bosteen, Markus H; Tritsaris, Katerina; Hansen, Anker J

    2014-01-01

    Interleukin-17A (IL-17A) is an important pro-inflammatory cytokine that regulates leukocyte mobilization and recruitment. To better understand how IL-17A controls leukocyte trafficking across capillaries in the peripheral blood circulation, we used primary human dermal microvascular endothelial...

  14. [Vascular endothelial Barrier Function].

    Science.gov (United States)

    Ivanov, A N; Puchinyan, D M; Norkin, I A

    2015-01-01

    Endothelium is an important regulator of selective permeability of the vascular wall for different molecules and cells. This review summarizes current data on endothelial barrier function. Endothelial glycocalyx structure, its function and role in the molecular transport and leukocytes migration across the endothelial barrier are discussed. The mechanisms of transcellular transport of macromolecules and cell migration through endothelial cells are reviewed. Special section of this article addresses the structure and function of tight and adherens endothelial junction, as well as their importance for the regulation of paracellular transport across the endothelial barrier. Particular attention is paid to the signaling mechanism of endothelial barrier function regulation and the factors that influence on the vascular permeability.

  15. Histamine Induces Vascular Hyperpermeability by Increasing Blood Flow and Endothelial Barrier Disruption In Vivo.

    Science.gov (United States)

    Ashina, Kohei; Tsubosaka, Yoshiki; Nakamura, Tatsuro; Omori, Keisuke; Kobayashi, Koji; Hori, Masatoshi; Ozaki, Hiroshi; Murata, Takahisa

    2015-01-01

    Histamine is a mediator of allergic inflammation released mainly from mast cells. Although histamine strongly increases vascular permeability, its precise mechanism under in vivo situation remains unknown. We here attempted to reveal how histamine induces vascular hyperpermeability focusing on the key regulators of vascular permeability, blood flow and endothelial barrier. Degranulation of mast cells by antigen-stimulation or histamine treatment induced vascular hyperpermeability and tissue swelling in mouse ears. These were abolished by histamine H1 receptor antagonism. Intravital imaging showed that histamine dilated vasculature, increased blood flow, while it induced hyperpermeability in venula. Whole-mount staining showed that histamine disrupted endothelial barrier formation of venula indicated by changes in vascular endothelial cadherin (VE-cadherin) localization at endothelial cell junction. Inhibition of nitric oxide synthesis (NOS) by L-NAME or vasoconstriction by phenylephrine strongly inhibited the histamine-induced blood flow increase and hyperpermeability without changing the VE-cadherin localization. In vitro, measurements of trans-endothelial electrical resistance of human dermal microvascular endothelial cells (HDMECs) showed that histamine disrupted endothelial barrier. Inhibition of protein kinase C (PKC) or Rho-associated protein kinase (ROCK), NOS attenuated the histamine-induced barrier disruption. These observations suggested that histamine increases vascular permeability mainly by nitric oxide (NO)-dependent vascular dilation and subsequent blood flow increase and maybe partially by PKC/ROCK/NO-dependent endothelial barrier disruption.

  16. [Microvascular anastomoses in cervical esophageal reconstruction].

    Science.gov (United States)

    Takushima, A; Harii, K; Asato, H

    2001-09-01

    Ischemia or hemostasis in the gastric, jejunal, and colonic pedicle after esophagectomy is believed to contribute significantly to postoperative complications. With the advent of microvascular anastomoses, many surgeons have adopted vascular augmentation (supercharge) as a means of avoiding these difficulties. Microsurgical free tissue transfer represented by the free jejunum and forearm flap also plays an important role in esophageal reconstruction. In this paper, the authors introduce the technical points important for successful revascularization including the choice of recipient vessels, setting up of the reconstructive materials, and postoperative monitoring. In cases of gastric pull-up elevated via posterior mediastinum, the left gastroduodenal vessels are anastomosed to the cervical transverse or superior thyroidal vessels. In cases of duodenal or colonic pull-up elevated via the anterosternal route, the vascular pedicles are anastomosed to the internal mammary vessels which are dissected by resecting the costal cartilage. When the free jejunum flap is used, the cervical transverse or superior thyroidal vessels are most frequently used as recipients. Postoperative monitoring of free flaps is performed using Doppler ultrasound or through a small skin incision made above the transferred tissue. Although gastric or colonic pull-up is difficult to monitor, color Doppler sonography permits quantitative analysis of blood flow and may be a useful option.

  17. Nox2-dependent glutathionylation of endothelial NOS leads to uncoupled superoxide production and endothelial barrier dysfunction in acute lung injury.

    Science.gov (United States)

    Wu, Feng; Szczepaniak, William S; Shiva, Sruti; Liu, Huanbo; Wang, Yinna; Wang, Ling; Wang, Ying; Kelley, Eric E; Chen, Alex F; Gladwin, Mark T; McVerry, Bryan J

    2014-12-15

    Microvascular barrier integrity is dependent on bioavailable nitric oxide (NO) produced locally by endothelial NO synthase (eNOS). Under conditions of limited substrate or cofactor availability or by enzymatic modification, eNOS may become uncoupled, producing superoxide in lieu of NO. This study was designed to investigate how eNOS-dependent superoxide production contributes to endothelial barrier dysfunction in inflammatory lung injury and its regulation. C57BL/6J mice were challenged with intratracheal LPS. Bronchoalveolar lavage fluid was analyzed for protein accumulation, and lung tissue homogenate was assayed for endothelial NOS content and function. Human lung microvascular endothelial cell (HLMVEC) monolayers were exposed to LPS in vitro, and barrier integrity and superoxide production were measured. Biopterin species were quantified, and coimmunoprecipitation (Co-IP) assays were performed to identify protein interactions with eNOS that putatively drive uncoupling. Mice exposed to LPS demonstrated eNOS-dependent increased alveolar permeability without evidence for altered canonical NO signaling. LPS-induced superoxide production and permeability in HLMVEC were inhibited by the NOS inhibitor nitro-l-arginine methyl ester, eNOS-targeted siRNA, the eNOS cofactor tetrahydrobiopterin, and superoxide dismutase. Co-IP indicated that LPS stimulated the association of eNOS with NADPH oxidase 2 (Nox2), which correlated with augmented eNOS S-glutathionylation both in vitro and in vivo. In vitro, Nox2-specific inhibition prevented LPS-induced eNOS modification and increases in both superoxide production and permeability. These data indicate that eNOS uncoupling contributes to superoxide production and barrier dysfunction in the lung microvasculature after exposure to LPS. Furthermore, the results implicate Nox2-mediated eNOS-S-glutathionylation as a mechanism underlying LPS-induced eNOS uncoupling in the lung microvasculature.

  18. UVB therapy decreases the adhesive interaction between peripheral blood mononuclear cells and dermal microvascular endothelium, and regulates the differential expression of CD54, VCAM-1, and E-selectin in psoriatic plaques

    Energy Technology Data Exchange (ETDEWEB)

    Cai, J.-P.; Harris, K.; Chin, Y.H. [Miami Univ., FL (United States). School of Medicine; Falanga, V.; Taylor, J.R. [Miami Univ., FL (United States). School of Medicine]|[Miami Veteran Affairs Medical Center, Miami, FL (United States)

    1996-01-01

    A dermal lymphocytic infiltrate is a characteristic feature of psoriasis, and may be involved in the pathogenesis of the disease. We have previously shown that specialized dermal microvascular endothelial cells (DMEC) in psoriatic lesions promote the selective adherence of the CD4 CD45Ro helper T-cell subset. In this study, we examined the adhesive interaction between peripheral blood mononuclear cells and psoriatic DMEC in patients treated with ultraviolet B light (UVB), and correlated the results with the expression and function of endothelial adhesion molecules on DMEC. (author).

  19. Endothelial cell processing and alternatively spliced transcripts of factor VIII: potential implications for coagulation cascades and pulmonary hypertension.

    Directory of Open Access Journals (Sweden)

    Claire L Shovlin

    Full Text Available BACKGROUND: Coagulation factor VIII (FVIII deficiency leads to haemophilia A. Conversely, elevated plasma levels are a strong predictor of recurrent venous thromboemboli and pulmonary hypertension phenotypes in which in situ thromboses are implicated. Extrahepatic sources of plasma FVIII are implicated, but have remained elusive. METHODOLOGY/PRINCIPAL FINDINGS: Immunohistochemistry of normal human lung tissue, and confocal microscopy, flow cytometry, and ELISA quantification of conditioned media from normal primary endothelial cells were used to examine endothelial expression of FVIII and coexpression with von Willebrand Factor (vWF, which protects secreted FVIII heavy chain from rapid proteloysis. FVIII transcripts predicted from database mining were identified by RT-PCR and sequencing. FVIII mAb-reactive material was demonstrated in CD31+ endothelial cells in normal human lung tissue, and in primary pulmonary artery, pulmonary microvascular, and dermal microvascular endothelial cells. In pulmonary endothelial cells, this protein occasionally colocalized with vWF, centered on Weibel Palade bodies. Pulmonary artery and pulmonary microvascular endothelial cells secreted low levels of FVIII and vWF to conditioned media, and demonstrated cell surface expression of FVIII and vWF Ab-reacting proteins compared to an isotype control. Four endothelial splice isoforms were identified. Two utilize transcription start sites in alternate 5' exons within the int22h-1 repeat responsible for intron 22 inversions in 40% of severe haemophiliacs. A reciprocal relationship between the presence of short isoforms and full-length FVIII transcript suggested potential splice-switching mechanisms. CONCLUSIONS/SIGNIFICANCE: The pulmonary endothelium is confirmed as a site of FVIII secretion, with evidence of synthesis, cell surface expression, and coexpression with vWF. There is complex alternate transcription initiation from the FVIII gene. These findings provide a

  20. Fat Grafting to the Breast, a Simple Procedure for a Very Complex Reconstruction.

    Science.gov (United States)

    Claro, Francisco; Pereira, Fernando Linhares; Pinto-Neto, Aarão M

    2016-01-01

    Usually, complicated reconstructions demand complex procedures. However, we report an unpublished situation where lipofilling was the only effective procedure for breast reconstruction, once 4 previous procedures, including 2 microvascular free flaps, had failed. The reported case describes a woman without subcutaneous tissue in the left breast topography, with radiation sequelae resulting in a fibrotic, hyperchromic, unexpandable skin that was tethered to her costal bone and pleura. The 4 previous attempts of breast reconstruction resulted in unavailable nearby recipient vessels, and this situation appointed breast lipofilling as the most feasible procedure. This report shows the power of breast lipofilling, a simple procedure that can be used even for the more complex reconstructions.

  1. A Novel Microvascular Flow Technique: Initial Results in Thyroids.

    Science.gov (United States)

    Machado, Priscilla; Segal, Sharon; Lyshchik, Andrej; Forsberg, Flemming

    2016-03-01

    To evaluate the flow imaging capabilities of a new prototype ultrasound (US) image processing technique (superb micro-vascular imaging [SMI]; Toshiba Medical Systems, Tokyo, Japan) for depiction of microvascular flow in normal thyroid tissue and thyroid nodules compared with standard color and power Doppler US imaging.Ten healthy volunteers and 22 patients, with a total of 25 thyroid nodules, scheduled for US-guided fine needle aspiration were enrolled in this prospective study. Subjects underwent US examination consisting of grayscale, color and power Doppler imaging (CDI and PDI) followed by color and monochrome SMI and pulsed Doppler. SMI is a novel, microvascular flow imaging mode implemented on the Aplio 500 US system (Toshiba). SMI uses advanced clutter suppression to extract flow signals from large to small vessels and depicts this information at high frame rates as a color overlay image or as a monochrome map of flow. Two radiologists independently scored still images and digital clips for overall flow detection, vessel branching details and noise on a visual-analog scale of 1 (worst) to 10 (best).For the volunteers SMI visualized microvasculature with significantly lower velocity than CDI and PDI (P SMI demonstrated microvascular flow with significantly higher image scores and provided better depiction of the vessel branching details compared with CDI and PDI (P SMI mode than in the other modes, including color SMI (P SMI mode consistently improved the depiction of thyroid microvascular flow compared with standard CDI and PDI.

  2. Endothelial binding of beta toxin to small intestinal mucosal endothelial cells in early stages of experimentally induced Clostridium perfringens type C enteritis in pigs.

    Science.gov (United States)

    Schumacher, V L; Martel, A; Pasmans, F; Van Immerseel, F; Posthaus, H

    2013-07-01

    Beta toxin (CPB) is known to be an essential virulence factor in the development of lesions of Clostridium perfringens type C enteritis in different animal species. Its target cells and exact mechanism of toxicity have not yet been clearly defined. Here, we evaluate the suitability of a neonatal piglet jejunal loop model to investigate early lesions of C. perfringens type C enteritis. Immunohistochemically, CPB was detected at microvascular endothelial cells in intestinal villi during early and advanced stages of lesions induced by C. perfringens type C. This was first associated with capillary dilatation and subsequently with widespread hemorrhage in affected intestinal segments. CPB was, however, not demonstrated on intestinal epithelial cells. This indicates a tropism of CPB toward endothelial cells and suggests that CPB-induced endothelial damage plays an important role in the early stages of C. perfringens type C enteritis in pigs.

  3. Platelets, acting in part via P-selectin, mediate cytomegalovirus-induced microvascular dysfunction.

    Science.gov (United States)

    Khoretonenko, Mikhail V; Brunson, Jerry L; Senchenkov, Evgeny; Leskov, Igor L; Marks, Christian R; Stokes, Karen Y

    2014-12-15

    Cytomegalovirus (CMV) infects a majority of the population worldwide. It has been implicated in cardiovascular disease, induces microvascular dysfunction, and synergizes with hypercholesterolemia to promote leukocyte and platelet recruitment in venules. Although platelets and platelet-associated P-selectin contribute to cardiovascular disease inflammation, their role in CMV-induced vascular responses is unknown. We assessed the role of platelets in CMV-induced microvascular dysfunction by depleting platelets and developing bone marrow chimeric mice deficient in platelet P-selectin. Wild-type and chimeric mice received mock or murine (m)CMV intraperitoneally. Five weeks later, some mice were switched to a high-cholesterol diet (HC) to investigate the synergism between mCMV and HC. Arteriolar vasodilation and recruitment of leukocytes and donor platelets in venules were measured at 11wk. mCMV with or without HC caused significant endothelial dysfunction in arterioles. Platelet depletion restored normal vasodilation in mCMV-HC but not mCMV-ND mice, whereas protection was seen in both groups for platelet P-selectin chimeras. Only mCMV + HC elevated leukocyte and platelet recruitment in venules. Leukocyte adhesion was reduced to mock levels by acute platelet depletion but was only partially decreased in platelet P-selectin chimeras. Platelets from mCMV-HC mice and, to a lesser extent, mCMV-ND but not mock-HC mice showed significant adhesion in mCMV-HC recipients. Our findings implicate a role for platelets, acting through P-selectin, in CMV-induced arteriolar dysfunction and suggest that the addition of HC leads to a platelet-dependent, inflammatory infiltrate that is only partly platelet P-selectin dependent. CMV appeared to have a stronger activating influence than HC on platelets and may represent an additional therapeutic target in vulnerable patients.

  4. Preparation of a designed poly(trimethylene carbonate) microvascular network by stereolithography.

    Science.gov (United States)

    Schüller-Ravoo, Sigrid; Zant, Erwin; Feijen, Jan; Grijpma, Dirk W

    2014-12-01

    Designed flexible and elastic network structures are prepared by stereolithography using a photo-crosslinkable resin based on a poly(trimethylene carbonate) (PTMC) macromer with a molecular weight of 3150 g/mol. Physical properties and the compatibility with human umbilical vein endothelial cells (HUVECs) are evaluated. The hydrophobic networks are found to be flexible and elastic, with an E modulus of 7.9 ± 0.1 MPa, a tensile strength of 3.5 ± 0.1 MPa and an elongation at break of 76.7 ± 0.7%. HUVECs attach and proliferate well on the surfaces of the built structures. A three-dimensional microvascular network is designed to serve as a perfusable scaffold for tissue engineering. In the design, 5 generations of open channels each branch into 4 smaller channels yielding a microvascular region with a high density of capillaries. The overall cross-sectional area through which medium or blood can be perfused remains constant. These structures would ensure efficient nourishment of cells in a large volume of tissue. Built by stereolithography using the PTMC resin, the smallest channels of these structures have square cross-sectional areas, with inner widths of approximately 224 μm and wall thicknesses of approximately 152 μm. The channels are open, allowing water to perfuse the scaffold at 0.279 ± 0.006 mL/s at 80 mmHg and 0.335 ± 0.009 mL/s at 120 mmHg.

  5. [Endothelial cell adhesion molecules].

    Science.gov (United States)

    Ivanov, A N; Norkin, I A; Puchin'ian, D M; Shirokov, V Iu; Zhdanova, O Iu

    2014-01-01

    The review presents current data concerning the functional role of endothelial cell adhesion molecules belonging to different structural families: integrins, selectins, cadherins, and the immunoglobulin super-family. In this manuscript the regulatory mechanisms and factors of adhesion molecules expression and distribution on the surface of endothelial cells are discussed. The data presented reveal the importance of adhesion molecules in the regulation of structural and functional state of endothelial cells in normal conditions and in pathology. Particular attention is paid to the importance of these molecules in the processes of physiological and pathological angiogenesis, regulation of permeability of the endothelial barrier and cell transmigration.

  6. High glucose augments stress-induced apoptosis in endothelial cells

    Institute of Scientific and Technical Information of China (English)

    Wenwen Zhong; Yang Liu; Hui Tian

    2009-01-01

    Hyperglycemia has been identified as one of the important factors involved in the microvascular complications of diabetes, and has been related to increased cardiovascular mortality. Endothelial damage and dysfunction result from diabetes; therefore, the aim of this study was to determine the response of endothelial cells to stressful stimuli, modelled in normal and high glucose concentrations in vitro. Eahy 926 endothelial cells were cultured in 5 mmol/L or 30 mmol/L glucose conditions for a 24 hour period and oxidative stress was induced by exposure to hydrogen peroxide (H2O2) or tumour necrosis factor- α (TNF- α ), following which the protective effect of the glucocorticoid dexamethasone was assessed. Apoptosis, necrosis and cell viability were determined using an ELISA for DNA fragmentation, an enzymatic lactate dehydrogenase assay and an MTT assay, respectively. High glucose significantly increased the susceptibility of Eahy 926 cells to apoptosis in the presence of 500 μmol/L H2O2, above that induced in normal glucose (P<0.02). A reduction of H2O2- and TNF- α -induced apoptosis occurred in both high and low glucose after treatment with dexametha-sone (P<0.05). Conclusion high glucose is effective in significantly augmenting stress caused by H2O2, but not in causing stress alone. These findings suggest a mechanism by which short term hyperglycemia may facilitate and augment endothelial damage.

  7. Retinal Microvascular Abnormalities in Neurofibromatosis Type 1 Associated with Congenital Retinal Macrovessels

    Science.gov (United States)

    Makino, Shinji; Endoh, Katsuhisa; Tampo, Hironobu

    2013-01-01

    Here, we report a case of retinal microvascular abnormalities in a patient with neurofibromatosis type 1 (NF1) associated with congenital retinal macrovessels. An abnormal retinal macrovessel, crossing the macula horizontally, was detected in the right eye. Additionally, retinal microvascular abnormalities were detected. Eight years after the initial visit, the retinal microvascular abnormalities were noted to have changed substantially. We speculate that retinal microvascular abnormalities in NF1 may change dynamically over the years. PMID:23781366

  8. Retinal Microvascular Abnormalities in Neurofibromatosis Type 1 Associated with Congenital Retinal Macrovessels

    Directory of Open Access Journals (Sweden)

    Shinji Makino

    2013-01-01

    Full Text Available Here, we report a case of retinal microvascular abnormalities in a patient with neurofibromatosis type 1 (NF1 associated with congenital retinal macrovessels. An abnormal retinal macrovessel, crossing the macula horizontally, was detected in the right eye. Additionally, retinal microvascular abnormalities were detected. Eight years after the initial visit, the retinal microvascular abnormalities were noted to have changed substantially. We speculate that retinal microvascular abnormalities in NF1 may change dynamically over the years.

  9. Endothelial cell adhesion and growth within a bioassay chamber using microstamped ECM proteins

    Science.gov (United States)

    Rubenstein, David A.; Frame, Mary D.

    2011-06-01

    Our goal was to evaluate microvascular endothelial cell growth on microstamped patterns of extracellular matrix proteins (ECM). A combination of photo- and soft-lithography was used to make features ˜100 μm deep and 150μm wide. Polydimethylsiloxane imprints of features produced positive molds used to stamp collagen I, IV, laminin and fibronectin onto cleaned hydrophilic or hydrophobic glass coverslips. Human dermal microvascular endothelial cells were seeded at an initial density of 800 cells cm-2, and cultured for three days. Explanted murine aortas, serving as an initial source for autologous endothelial cells, were perfused at 240 μL min-1 for 1 day. Cell morphology was also quantified on both the non-patterned glass and within the microstamped patterns. Viability was high (>90%) on all microstamped proteins, regardless of glass hydrophobicity. Viability was reduced on bare hydrophobic glass. Cell density was 4 or 8 fold higher on microstamped ECM proteins compared with hydrophilic or hydrophobic glass, respectively. Confluence was approached more rapidly on microstamped proteins. Thus, rapid concentrated growth of endothelial cells was markedly enhanced within microstamped ECM patterns on hydrophilic and hydrophobic glass.

  10. Endothelial-myofibroblast transition contributes to the early development of diabetic renal interstitial fibrosis in streptozotocin-induced diabetic mice.

    Science.gov (United States)

    Li, Jinhua; Qu, Xinli; Bertram, John F

    2009-10-01

    Diabetic nephropathy is the leading cause of chronic renal failure. Myofibroblasts play a major role in the synthesis and secretion of extracellular matrix in diabetic renal fibrosis. Increasing evidence suggests that endothelial cells may undergo endothelial-myofibroblast transition under physiological and pathophysiological circumstances. Therefore, this study investigates whether endothelial-myofibroblast transition occurs and contributes to the development of diabetic renal interstitial fibrosis. Diabetes was induced by administration of streptozotocin to Tie2-Cre;LoxP-EGFP mice, an endothelial lineage-traceable mouse line generated by crossbreeding B6.Cg-Tg(Tek-cre)12F1v/J mice with B6.Cg-Tg(ACTB-Bgeo/GFP)21Lbe/J mice. The endothelial-myofibroblast transition was also studied in MMECs (a mouse pancreatic microvascular endothelial cell line) and primary cultures of CD31+/EYFP- (enhanced yellow fluorescent protein) endothelial cells isolated from adult normal alpha-smooth muscle actin promoter-driven-EYFP (alpha-SMA/EYFP) mouse kidneys. Confocal microscopy demonstrated that 10.4 +/- 4.2 and 23.5 +/- 7.4% of renal interstitial myofibroblasts (alpha-SMA+) in 1- and 6-month streptozotocin-induced diabetic kidneys were of endothelial origin (EGFP+/alpha-SMA+ cells), compared with just 0.2 +/- 0.1% of myofibroblasts in vehicle-treated Tie2-Cre;LoxP-EGFP mice (P diabetic renal interstitial fibrosis and suggest that the endothelial-myofibroblast transition may be a therapeutic target.

  11. Endothelium dependent vasomotion and in vitro markers of endothelial repair in patients with severe sepsis: an observational study.

    Directory of Open Access Journals (Sweden)

    Sabrina H van Ierssel

    Full Text Available BACKGROUND: Outcome in sepsis is mainly defined by the degree of organ failure, for which endothelial dysfunction at the macro- and microvascular level is an important determinant. In this study we evaluated endothelial function in patients with severe sepsis using cellular endothelial markers and in vivo assessment of reactive hyperaemia. MATERIALS AND METHODS: Patients with severe sepsis (n = 30 and 15 age- and gender- matched healthy volunteers were included in this study. Using flow cytometry, CD34+/KDR+ endothelial progenitor cells (EPC, CD31+ T-cells, and CD31+/CD42b- endothelial microparticles (EMP were enumerated. Migratory capacity of cultured circulating angiogenic cells (CAC was assessed in vitro. Endothelial function was determined using peripheral arterial tonometry at the fingertip. RESULTS: In patients with severe sepsis, a lower number of EPC, CD31+ T-cells and a decreased migratory capacity of CAC coincided with a blunted reactive hyperaemia response compared to healthy subjects. The number of EMP, on the other hand, did not differ. The presence of organ failure at admission (SOFA score was inversely related with the number of CD31+ T-cells. Furthermore, the number of EPC at admission was decreased in patients with progressive organ failure within the first week. CONCLUSION: In patients with severe sepsis, in vivo measured endothelial dysfunction coincides with lower numbers and reduced function of circulating cells implicated in endothelial repair. Our results suggest that cellular markers of endothelial repair might be valuable in the assessment and evolution of organ dysfunction.

  12. Induction of Endothelial Cell Apoptosis by Anti-alpha-enolase Antibody

    Institute of Scientific and Technical Information of China (English)

    Hong-bo Yang; Wen-jie Zheng; Xuan Zhang; Fu-lin Tang

    2011-01-01

    Objective To assess the prevalence of anti-alpha-enolase antibody in systemic autoimmune diseases in Chinese patients and its role in endothelial cell apoptosis.Methods The reactivity of anti-alpha-enolase antibody in a variety of autoimmune disorders in Chinese patients was evaluated by dot blot assay. Endothelial cell apoptosis was investigated by in vitro incubation of endothelial cells with IgG purified from anti-alpha-enolase antibody-positive sera, with or without pre-incubation with recombinant alpha-enolase.Results Anti-alpha-enolase antibody was prevalent in different systemic autoimmune diseases with relatively high reactivity in Chinese patients. In vitro incubation of endothelial cells with IgG containing anti-alpha-enolase antibody induced apoptosis in a time- and dose-dependent manner. Apoptosis was partly inhibited by pre-incubation of the endothelial cells with recombinant alpha-enolase.Conclusions Our data suggest that alpha-enolase is a common auto-antigen recognized by antiendothelial cell antibodies in connective tissue disease. Interaction between alpha-enolase and its autoantibody plays a role in endothelial cell apoptosis. Changes other than cell killing may contribute to the pathogenesis of endothelial damage and microvascular lesions.

  13. Protective effects of quercetin on rat pial microvascular changes during transient bilateral common carotid artery occlusion and reperfusion.

    Directory of Open Access Journals (Sweden)

    Dominga eLapi

    2012-03-01

    Full Text Available The aim of this study was to assess the in vivo effects of quercetin on pial microvascular responses during transient bilateral common carotid artery occlusion (BCCAO and reperfusion. Rat pial microcirculation was visualized by fluorescence microscopy through a closed cranial window. Pial arterioles were classified in five orders of branchings. Capillaries were assigned order 0, the smallest arterioles order 1 and the largest ones order 5. In ischemic rats, 30 min BCCAO and 60 min reperfusion caused arteriolar diameter decrease (by 14.5 ± 3.3% of baseline in order 2, microvascular leakage (0.47 ± 0.04 NGL: Normalized Grey Levels, leukocyte adhesion in venules (9 ± 2/100 µm venular length, v.l./30s and reduction of capillary perfusion (by 40 ± 7% of baseline. Moreover, at the end of BCCAO and reperfusion there was a significant increase in reactive oxygen species formation (ROS when compared with baseline. Quercetin highest dose determined dilation in all arteriolar orders (by 40 ± 4 % of baseline in order 2 and prevented microvascular permeability (0.15 ± 0.02 NGL, leukocyte adhesion (3 ± 1/100 µm v.l./30s as well as ROS formation, while capillary perfusion was protected. Inhibition of endothelial Nitric Oxide Synthase (NOS prior to quercetin reduced arteriolar dilation (order 2 diameter increase by 10.3 ± 2.5% of baseline and caused permeability increase (0.29 ± 0.03 NGL; inhibition of neuronal NOS or inducible NOS did not affect quercetin-induced effects. Inhibition of guanylyl cyclase prior to quercetin reversed the quercetin’s effects on pial arteriolar diameter and leakage. In conclusion, quercetin was able to protect pial microcirculation from ischemia-reperfusion damage inducing arteriolar dilation likely by nitric oxide release. Moreover, quercetin scavenger activity blunted reactive oxygen species formation preserving the blood-brain barrier integrity.

  14. Ultrasound - Breast

    Science.gov (United States)

    ... Physician Resources Professions Site Index A-Z Ultrasound - Breast Ultrasound imaging of the breast uses sound waves ... the Breast? What is Ultrasound Imaging of the Breast? Ultrasound is safe and painless, and produces pictures ...

  15. Breast pain

    Science.gov (United States)

    Pain - breast; Mastalgia; Mastodynia; Breast tenderness ... There are many possible causes for breast pain. For example, hormone level changes from menstruation or pregnancy often cause breast tenderness. Some swelling and tenderness just before your period ...

  16. Nitric oxide-induced changes in endothelial expression of phosphodiesterases 2, 3, and 5

    DEFF Research Database (Denmark)

    Schankin, Christoph J; Kruse, Lars S; Reinisch, Veronika M;

    2010-01-01

    line containing such PDEs. METHODS: Real time polymerase chain reaction and Western blots were used to show expression of PDE2A, PDE3B, and PDE5A in a stable cell line of human brain microvascular endothelial cells. Effects of NO on PDE expression were analyzed at specific time intervals after......OBJECTIVE: To investigate nitric oxide (NO)-mediated changes in expression of cyclic nucleotide degrading phosphodiesterases 2A (PDE2A), PDE3B, and PDE5A in human endothelial cells. BACKGROUND: Nitric oxide induces production of cyclic guanosine monophosphate (cGMP), which along with cyclic...... adenosine monophosphate (cAMP) is degraded by PDEs. NO donors and selective inhibitors of PDE3 and PDE5 induce migraine-like headache and play a role in endothelial dysfunction during stroke. The current study investigates possible NO modulation of cGMP-related PDEs relevant to headache induction in a cell...

  17. Repeat microvascular decompression for recurrent idiopathic trigeminal neuralgia

    NARCIS (Netherlands)

    Bakker, Nicolaas A.; van Dijk, J. Marc C.; Immenga, Steven; Wagemakers, Michiel; Metzemaekers, Jan D. M.

    2014-01-01

    Object. Microvascular decompression (MVD) is considered the method of choice to treat idiopathic trigeminal neuralgia (TN) refractory to medical treatment. However, repeat MVD for recurrent TN is not well established. In this paper, the authors describe a large case series in which patients underwen

  18. Vitamin D and retinal microvascular damage : The Rotterdam Study

    NARCIS (Netherlands)

    Mutlu, Unal; Ikram, M Arfan; Hofman, Albert; de Jong, Paulus T V M; Uitterlinden, Andre G; Klaver, Caroline C W; Ikram, M Kamran

    2016-01-01

    Vitamin D has been linked to various cardiovascular risk factors including indices of large-vessel disease. However, it remains unclear whether vitamin D is also associated with microvascular damage. In a community-dwelling population, we studied associations between vitamin D serum levels and retin

  19. Microvascular networks for continuous self-healing materials

    Science.gov (United States)

    Toohey, Kathleen Suzanne

    Microcapsule-based and hollow glass fiber-based self-healing composites enable a single repair of crack damage in a given location. Re-mendable polymers have demonstrated crack mending for multiple cycles, but only with heat treatment and applied pressure. We demonstrate, for the first time, the autonomic characteristics of microencapsulated systems with the ability to heal repeated damage events in a continuous self-healing coating on a microvascular substrate. A three-dimensional, interconnected microvascular network in the substrate serves as a delivery system for healing materials to heal damage in a polymer coating. Two approaches to microvascular delivery are explored: one using a fluid monomer in the network with solidphase catalyst particles in the coating, and one using two or more separate networks with two fluid healing components. In both systems, the fluid(s) flow from the microchannels into the crack plane through capillary action, and polymerization occurs to heal the crack. In the single-fluid system, a single crack in a brittle epoxy coating is healed as many as seven times, and the ability to heal continuously is limited only by the availability of catalyst. The two-part healing system contained in separate networks in a specimen has significant potential to extend the repeatability of the healing process due to the microvascular supply of both healing components. Preliminary tests of this two-part system demonstrate that multiple healing cycles are possible.

  20. Mouse lung contains endothelial progenitors with high capacity to form blood and lymphatic vessels

    Directory of Open Access Journals (Sweden)

    Barleon Bernhard

    2010-07-01

    Full Text Available Abstract Background Postnatal endothelial progenitor cells (EPCs have been successfully isolated from whole bone marrow, blood and the walls of conduit vessels. They can, therefore, be classified into circulating and resident progenitor cells. The differentiation capacity of resident lung endothelial progenitor cells from mouse has not been evaluated. Results In an attempt to isolate differentiated mature endothelial cells from mouse lung we found that the lung contains EPCs with a high vasculogenic capacity and capability of de novo vasculogenesis for blood and lymph vessels. Mouse lung microvascular endothelial cells (MLMVECs were isolated by selection of CD31+ cells. Whereas the majority of the CD31+ cells did not divide, some scattered cells started to proliferate giving rise to large colonies (> 3000 cells/colony. These highly dividing cells possess the capacity to integrate into various types of vessels including blood and lymph vessels unveiling the existence of local microvascular endothelial progenitor cells (LMEPCs in adult mouse lung. EPCs could be amplified > passage 30 and still expressed panendothelial markers as well as the progenitor cell antigens, but not antigens for immune cells and hematopoietic stem cells. A high percentage of these cells are also positive for Lyve1, Prox1, podoplanin and VEGFR-3 indicating that a considerabe fraction of the cells are committed to develop lymphatic endothelium. Clonogenic highly proliferating cells from limiting dilution assays were also bipotent. Combined in vitro and in vivo spheroid and matrigel assays revealed that these EPCs exhibit vasculogenic capacity by forming functional blood and lymph vessels. Conclusion The lung contains large numbers of EPCs that display commitment for both types of vessels, suggesting that lung blood and lymphatic endothelial cells are derived from a single progenitor cell.

  1. Short-term microvascular response of striated muscle to cp-Ti, Ti-6Al-4V, and Ti-6Al-7Nb.

    Science.gov (United States)

    Pennekamp, Peter H; Gessmann, Jan; Diedrich, Oliver; Burian, Björn; Wimmer, Markus A; Frauchiger, Vinzenz M; Kraft, Clayton N

    2006-03-01

    Due to excellent mechanical properties and good corrosion resistance, titanium-aluminium-vanadium (Ti-6Al-4V) and titanium-aluminium-niobium (Ti-6Al-7Nb) are extensively used for orthopedic surgery. Concern has been voiced concerning the implications of the constituent vanadium in Ti-6Al-4V on the surrounding environment. Particularly in osteosynthesis where the alloys stand in direct contact to skeletal muscle, undesirable biologic reactions may have severe consequences. In a comparative study, we assessed in vivo nutritive perfusion and leukocytic response of striated muscle to the metals Ti-6Al-4V, Ti-6Al-7Nb, and commercially pure titanium (cpTi), thereby drawing conclusions on their short-term inflammatory potential. In 28 hamsters, utilizing the dorsal skinfold chamber preparation and intravital microscopy, we quantified primary and secondary leukocyte-endothelial cell interaction, leukocyte extravasation, microvascular diameter change, and capillary perfusion in collecting and postcapillary venules of skeletal muscle. A manifest discrepancy between the metals concerning impact on local microvascular parameters was not found. All metals induced an only transient and moderate inflammatory response. Only a slight increase in leukocyte recruitment and a more sluggish recuperation of inflammatory parameters in animals treated with Ti-6Al-4V compared to the other two metals suggested a minor, overall not significant discrepancy in biocompatibility. Gross toxicity of bulk Ti-6Al-4V on surrounding tissue could not be found. Conclusively, the commonly used biomaterials Ti-6Al-4V, Ti-6Al-7Nb, and cpTi induce an only transient inflammatory answer of the skeletal muscle microvascular system. Our results indicate that on the microvascular level the tested bulk Ti-alloys and cpTi do not cause adverse biologic reactions in striated muscle.

  2. Phentolamine inhibits angiogenesis in vitro: Suppression of proliferation migration and differentiation of human endothelial cells.

    Science.gov (United States)

    Pan, Liangli; Liu, Chenyang; Kong, Yanan; Piao, Zhengguo; Cheng, Biao

    2016-06-16

    It is widely known that the β-adrenergic receptor (AR) blocker (propranolol) inhibits human endothelial cell (EC) angiogenesis in vitro, but how the α-AR antagonist (phentolamine) affects human EC angiogenesis has not yet been studied. Here, we show for the first time that both human dermal microvascular ECs (HDMECs) and human brain microvascular ECs (HBMECs) express α-ARs. Moreover, our results indicate that phentolamine inhibits the proliferation, migration, and tubulogenesis of HDMECs and HBMECs. Finally, VEGFR-2 and Ang1/2 expression of HDMECs was suppressed by phentolamine. Together, these results indicate that phentolamine impairs several critical events of neovascularization, and α-ARs, as well as the VEGF/VEGFR-2 and Ang/Tie-2 signaling pathways, may be involved in these processes. Our results suggest a novel therapeutic strategy for the use of α-blockers in the treatment of human angiogenesis-dependent diseases.

  3. Intermittent hypoxia-induced endothelial barrier dysfunction requires ROS-dependent MAP kinase activation.

    Science.gov (United States)

    Makarenko, Vladislav V; Usatyuk, Peter V; Yuan, Guoxiang; Lee, May M; Nanduri, Jayasri; Natarajan, Viswanathan; Kumar, Ganesh K; Prabhakar, Nanduri R

    2014-04-15

    The objective of the present study was to determine the impact of simulated apnea with intermittent hypoxia (IH) on endothelial barrier function and assess the underlying mechanism(s). Experiments were performed on human lung microvascular endothelial cells exposed to IH-consisting alternating cycles of 1.5% O2 for 30s followed by 20% O2 for 5 min. IH decreased transendothelial electrical resistance (TEER) suggesting attenuated endothelial barrier function. The effect of IH on TEER was stimulus dependent and reversible after reoxygenation. IH-exposed cells exhibited stress fiber formation and redistribution of cortactin, vascular endothelial-cadherins, and zona occludens-1 junction proteins along with increased intercellular gaps at cell-cell boundaries. Extracellular signal-regulated kinase (ERK) and c-jun NH2-terminal kinase (JNK) were phosphorylated in IH-exposed cells. Inhibiting either ERK or JNK prevented the IH-induced decrease in TEER and the reorganization of the cytoskeleton and junction proteins. IH increased reactive oxygen species (ROS) levels, and manganese (III) tetrakis (1-methyl-4-pyridyl) porphyrin pentachloride, a membrane-permeable antioxidant, prevented ERK and JNK phosphorylation as well as IH-induced changes in endothelial barrier function. These results demonstrate that IH via ROS-dependent activation of MAP kinases leads to reorganization of cytoskeleton and junction proteins resulting in endothelial barrier dysfunction.

  4. Endothelial cells of the blood-brain barrier: a target for glucocorticoids and estrogens?

    Science.gov (United States)

    Dietrich, Jean-Bernard

    2004-01-01

    Adhesion molecules are involved in the leukocyte recruitment of leukocytes at the blood-brain barrier. For this reason, it is important to understand how the regulation of their gene expression controls lymphocyte adhesion to endothelial cells in microvessels. Indeed, due to their specificity and diversity, adhesion molecules involved in extravasation play an essential role in the recruitment of activated leukocytes and activation of inflammation. Multiple sclerosis results from a chronic inflammation of the CNS which is mediated by infiltration of inflammatory cells from the immune system. Administration of glucocorticoids is a routine method to control multiple sclerosis since naturally derived or synthetic glucocorticoids are potent immunosuppressive and anti-inflammatory agents. Glucocorticoids also have beneficial effects in stabilizing the blood-brain barrier, as steroid hormones regulate the expression of adhesion molecule genes in endothelial cells. Other hormones such as estrogens modulate many endothelial cell biological activities, among them adhesion to leukocytes. They regulate expression of adhesion molecules genes on endothelial cells and are useful for the treatment of experimental autoimmune encephalomyelitis, the animal model of multiple sclerosis. The effects of glucocorticoids and estrogens on the expression of adhesion molecules on endothelial cells, including microvascular endothelial cells of the blood-brain barrier, are reviewed in this paper, as well as the involvement of these hormones in the therapy of experimental autoimmune encephalomyelitis and multiple sclerosis.

  5. Angiopoietin-1 protects myocardial endothelial cell function blunted by angiopoietin-2 and high glucose condition

    Institute of Scientific and Technical Information of China (English)

    Qin-hui TUO; Guo-zuo XIONG; Heng ZENG; Hei-di YU; Shao-wei SUN; Hong-yan LING; Bing-yang ZHU; Duan-fang LIAO; Jian-xiong CHEN

    2011-01-01

    Aim:To evaluate the effects of angiopoietin-1 (Ang-1) on myocardial endothelial cell function under high glucose (HG) condition.Methods:Mouse heart myocardial endothelial cells (MHMECs) were cultured and incubated under HG (25 mmol/L) or normal glucose (NG, 5 mmol/L) conditions for 72 h. MTT was used to determine cellular viability, and TUNEL assay and caspase-3 enzyme linked immunosorbent assays were used to assay endothelial apoptosis induced by serum starvation. Immunoprecipitation and Western blot analysis were used to analyze protein phosphorylation and expression. Endothelial tube formation was used as an in vitro assay for angiogenesis.Results:Exposure of MHMECs to HG resulted in dramatic decreases in phosphorylation of the Tie-2 receptor and its downstream signaling partners, Akt/eNOS, compared to that under NG conditions. Ang-1 (250 ng/mL) increased Tie-2 activation, inhibited cell apoptosis, and promoted angiogenesis. Ang-1-mediated protection of endothelial function was blunted by Ang-2 (25 ng/mL).Conclusion:Ang-1 activates the Tie-2 pathway and restores hyperglycemia-induced myocardial microvascular endothelial dysfunction.This suggests a protective role of Ang-1 in the ischemic myocardium, particularly in hearts affected by hyperglycemia or diabetes.

  6. Coniferyl Aldehyde Ameliorates Radiation Intestine Injury via Endothelial Cell Survival

    Energy Technology Data Exchange (ETDEWEB)

    Jeong, Ye Ji; Jung, Myung Gu; Lee, Yoonjin; Lee, Haejune [Korea Institute of Radiological and Medical Sciences, Seoul (Korea, Republic of); Lee, Yunsil [Ewha Woman' s Univ., Seoul (Korea, Republic of); Ko, Younggyu [Korea Univ., Seoul (Korea, Republic of)

    2014-05-15

    Cancer treatments related gastrointestinal toxicity has also been recognized as a significant economic burden. Especially, extensive apoptosis of microvascular endothelial cell of the lamina propria is the primary lesion initiating intestinal radiation damage after abdominal radiation therapy. Coniferyl aldehyde (CA) is phenolic compounds isolated from cork stoppers, and one of the major pyrolysis products of lignin. Shi H. was support for the empirical use of CA as a medicinal food for cardiovascular diseases. CA has positive effect in broad way but there is no consequence in radiation induced intestine damage. Here, we investigate effect of CA on small intestine after abdominal IR to mice in this study. In this study, CA increased the survival rate in C3H mice against 13.5 Gy abdominal IR. We found CA protects small intestine via preventing endothelial cell apoptosis and enhancing their angiogenic activity. CA also showed protective effect on crypt cell survival. Endothelial cell survival may affect crypt cell protection against IR. From this data, we concluded that CA is effective for protection against abdominal radiation injury. CA could ameliorate side-effect of radiation therapy.

  7. Transcriptional Regulation of Cystathionine-γ-Lyase in Endothelial Cells by NADPH Oxidase 4-Dependent Signaling*

    Science.gov (United States)

    Mistry, Rajesh K.; Murray, Thomas V. A.; Prysyazhna, Oleksandra; Martin, Daniel; Burgoyne, Joseph R.; Santos, Celio; Eaton, Philip; Shah, Ajay M.; Brewer, Alison C.

    2016-01-01

    The gasotransmitter, hydrogen sulfide (H2S) is recognized as an important mediator of endothelial cell homeostasis and function that impacts upon vascular tone and blood pressure. Cystathionine-γ-lyase (CSE) is the predominant endothelial generator of H2S, and recent evidence suggests that its transcriptional expression is regulated by the reactive oxygen species, H2O2. However, the cellular source of H2O2 and the redox-dependent molecular signaling pathway that modulates this is not known. We aimed to investigate the role of Nox4, an endothelial generator of H2O2, in the regulation of CSE in endothelial cells. Both gain- and loss-of-function experiments in human endothelial cells in vitro demonstrated Nox4 to be a positive regulator of CSE transcription and protein expression. We demonstrate that this is dependent upon a heme-regulated inhibitor kinase/eIF2α/activating transcription factor 4 (ATF4) signaling module. ATF4 was further demonstrated to bind directly to cis-regulatory sequences within the first intron of CSE to activate transcription. Furthermore, CSE expression was also increased in cardiac microvascular endothelial cells, isolated from endothelial-specific Nox4 transgenic mice, compared with wild-type littermate controls. Using wire myography we demonstrate that endothelial-specific Nox4 transgenic mice exhibit a hypo-contractile phenotype in response to phenylephrine that was abolished when vessels were incubated with a CSE inhibitor, propargylglycine. We, therefore, conclude that Nox4 is a positive transcriptional regulator of CSE in endothelial cells and propose that it may in turn contribute to the regulation of vascular tone via the modulation of H2S production. PMID:26620565

  8. Treatment of Angina Pectoris Associated with Coronary Microvascular Dysfunction.

    Science.gov (United States)

    Ong, Peter; Athanasiadis, Anastasios; Sechtem, Udo

    2016-08-01

    Treatment of angina pectoris associated with coronary microvascular dysfunction is challenging as the underlying mechanisms are often diverse and overlapping. Patients with type 1 coronary microvascular dysfunction (i.e. absence of epicardial coronary artery disease and myocardial disease) should receive strict control of their cardiovascular risk factors and thus receive statins and ACE-inhibitors in most cases. Antianginal medication consists of ß-blockers and/or calcium channel blockers. Second line drugs are ranolazine and nicorandil with limited evidence. Despite individually titrated combinations of these drugs up to 30 % of patients have refractory angina. Rho-kinase inhibitors and endothelin-receptor antagonists represent potential drugs that may prove useful in these patients in the future.

  9. Investigation of the improvement of Salviae miltiorrhiza combined with low molecular heparin sodium on endothelial dysfunctions and hemorheological abnormal after breast cancer surgery%丹参注射液联合低分子肝素改善乳腺癌手术后血管内皮细胞功能和血流变异常的临床研究

    Institute of Scientific and Technical Information of China (English)

    罗静; 黄元坤; 周志兵

    2013-01-01

    目的 观察丹参注射液联合低分子肝素对乳腺癌手术后血管内皮细胞功能和血流变的改善情况.方法 将100例乳腺癌手术患者随机分为两组,其中丹参联合低分子肝素组(观察组)50例静脉给予丹参注射液,并皮下注射低分子肝素钠;低分子肝素组(对照组)50例,只皮下注射低分子肝素钠,于第1天和14天观察两组临床疗效及临床积分、血管内皮功能指标、血浆TXA2和PGI2水平和血流变参数变化.结果 观察组总有效率94%,时照组总有效率86%,两组间差异有统计学意义(x2=8.104,P<0.05);观察组临床积分优于对照组,差异有统计学意义(P<0.01).观察组ET-I水平、tPA水平、TXA2和PGI2水平均较对照组低,差异有统计学意义(均P<0.05);观察组血流变参数红细胞比容%(HCT%)、血浆粘度比(mPa.s)、红细胞电泳时间(s)和纤维蛋白原(g/L)改善情况优于对照组,差异有统计学意义(均P<0.05).两组患者治疗中均未出现严重不良反应.结论 丹参联合低分子肝素可明显改善乳腺癌术后患者血管内皮细胞功能和血流变异常状态,能有效预防下肢深静脉血栓的影成和肺栓塞的发生.%Objective To investigate the effect of salviae miltiorrhiza combined with low molecular heparin sodium on endothelial function and hemorheological parameters in patients after breast cancer surgery.Methods 100 patients after breast cancer surgery were randomly divided into 2 group,salviae miltiorrhiza combined with low molecular heparin sodium treatment group (treatment group) and low molecular heparin sodium treatment group (control group),with 50 patients in each.The curative effect,clinical integrals,endothelial indexes were evaluated.And the serum levels of TXA2 and PGI2 and hemorheological parameters were measured.Results The curative effect and clinical integrals were better in treatment group (P<0.05 in all) on day 14.Meanwhile,ET-1,tPA,TXA2 and PGI2

  10. A macromolecular model for the endothelial surface layer

    Science.gov (United States)

    Harden, James; Danova-Okpetu, Darina; Grest, Gary

    2006-03-01

    The endothelial surface layer (ESL) is a micron-scale macromolecular lining of the luminal side of blood vessels composed of proteoglycans, glycoproteins, polysaccharides and associated plasma proteins all in dynamic equilibrium. It has numerous physiological roles including the regulation of blood flow and microvascular permeability, and active participation in mechanotransduction and stress regulation, coagulation, cell adhesion, and inflammatory response. The dynamic structure and the mechanical properties of the ESL are crucial for many of its physiological properties. We present a topological model for the ESL composed of three basic macromolecular elements: branched proteoglycans, linear polysaccharide chains, and small plasma proteins. The model was studied using non-equilibrium molecular dynamics simulations and compared with scaling theories for associating tethered polymers. We discuss the observed dynamical and mechanical properties of the ESL captured by this model, and the possible physical insight it provides into the physiological behavior of the ESL.

  11. Preparation of chitosan films using different neutralizing solutions to improve endothelial cell compatibility.

    Science.gov (United States)

    He, Qing; Ao, Qiang; Gong, Yandao; Zhang, Xiufang

    2011-12-01

    The development of chitosan-based constructs for application in large-size defects or highly vascularized tissues is still a challenging issue. The poor endothelial cell compatibility of chitosan hinders the colonization of vascular endothelial cells in the chitosan-based constructs, and retards the establishment of a functional microvascular network following implantation. The aim of the present study is to prepare chitosan films with different neutralization methods to improve their endothelial cell compatibility. Chitosan salt films were neutralized with either sodium hydroxide (NaOH) aqueous solution, NaOH ethanol solution, or ethanol solution without NaOH. The physicochemical properties and endothelial cell compatibility of the chitosan films were investigated. Results indicated that neutralization with different solutions affected the surface chemistry, swelling ratio, crystalline conformation, nanotopography, and mechanical properties of the chitosan films. The NaOH ethanol solution-neutralized chitosan film (Chi-NaOH/EtOH film) displayed a nanofiber-dominant surface, while the NaOH aqueous solution-neutralized film (Chi-NaOH/H(2)O film) and the ethanol solution-neutralized film (Chi-EtOH film) displayed nanoparticle-dominant surfaces. Moreover, the Chi-NaOH/EtOH films exhibited a higher stiffness as compared to the Chi-NaOH/H(2)O and Chi-EtOH films. Endothelial cell compatibility of the chitosan films was evaluated with a human microvascular endothelial cell line, HMEC-1. Compared with the Chi-NaOH/H(2)O and Chi-EtOH films, HMECs cultured on the Chi-NaOH/EtOH films fully spread and exhibited significantly higher levels of adhesion and proliferation, with retention of the endothelial phenotype and function. Our findings suggest that the surface nanotopography and mechanical properties contribute to determining the endothelial cell compatibility of chitosan films. The nature of the neutralizing solutions can affect the physicochemical properties and

  12. Early experiences with microvascular free tissue transfer in Lagos, Nigeria

    Directory of Open Access Journals (Sweden)

    Bolaji O Mofikoya

    2014-01-01

    Full Text Available Objectives: Microvascular free tissue transfer within our subregion is fraught with considerable challenges. We aim to highlight our experiences gained with our first fifteen cases of microvascular free tissue transfer at the Lagos University Teaching Hospital. We believe our report will be useful to colleagues embarking on such reconstructions in similar settings. Materials and Methods: The clinical records of the first 15 cases of free flaps done at our center were reviewed. The indications for surgery, choice of flap, recipient vessels, duration of surgery and complications were noted. Results: Fifteen cases were done, 10 flaps survived, ten defects occurred following trauma while remaining five followed cancer resections. Anterolateral thigh and radial forearm flaps were the most common flap used. The mean duration of surgeries was 7.1 hours SD ± 1.10 hours. Our take back rate was 13.3%, with a salvage rate of 50%. Three flaps failed on account venous congestion while remaining two failed due to arterial occlusion. Conclusion: 66.67% free flap success rate recorded reflect our early experiences in our institution. We believe meticulous planning, careful vessel selection, close flap monitoring as well as improved infrastructural support can lead to much better success rates in microvascular reconstruction in our country.

  13. Microfabrication of cylindrical microfluidic channel networks for microvascular research.

    Science.gov (United States)

    Huang, Zhouchun; Li, Xiang; Martins-Green, Manuela; Liu, Yuxin

    2012-10-01

    Current methods for formation of microvascular channel scaffolds are limited with non-circular channel cross-sections, complicated fabrication, and less flexibility in microchannel network design. To address current limitations in the creation of engineered microvascular channels with complex three-dimensional (3-D) geometries in the shape of microvessels, we have developed a reproducible, cost-effective, and flexible micromanufacturing process combined with photolithographic reflowable photoresist and soft lithography techniques to fabricate cylindrical microchannel and networks. A positive reflowable photoresist AZ P4620 was used to fabricate a master microchannel mold with semi-circular cross-sections. By the alignment and bonding of two polydimethylsiloxane (PDMS) microchannels replicated from the master mold together, a cylindrical microchannel or microchannel network was created. Further examination of the channel dimensions and surface profiles at different branching levels showed that the shape of the microfluidic channel was well approximated by a semi-circular surface, and a multi-level, multi-depth channel network was created. In addition, a computational fluidic dynamics (CFD) model was used to simulate shear flows and corresponding pressure distributions inside of the microchannel and channel network based on the dimensions of the fabricated channels. The fabricated multi-depth cylindrical microchannel network can provide platforms for the investigation of microvascular cells growing inside of cylindrical channels under shear flows and lumen pressures, and work as scaffolds for the investigation of morphogenesis and tubulogenesis.

  14. Preventing microvascular complications in type 1 diabetes mellitus

    Directory of Open Access Journals (Sweden)

    Vijay Viswanathan

    2015-01-01

    Full Text Available Patients with complications of diabetes such as retinopathy, nephropathy, and cardiovascular complications have increased hospital stay with greater economic burden. Prevention of complications should be started before the onset of type 1 diabetes mellitus (T1DM by working on risk factors and thereafter by intervention upon confirmatory diagnosis which can prevent further damage to β-cells. The actual risk of getting microvascular complications like microalbuminuria and retinopathy progression starts at glycated hemoglobin (HbA1c level of 7%. As per the American Diabetes Association, a new pediatric glycemic control target of HbA1c 20 years as compared to patients <10 years of age. Screening of these complications should be done regularly, and appropriate preventive strategies should be followed. Angiotensin converting enzyme inhibitors and angiotensin II receptor blocker reduce progression from microalbuminuria to macroalbuminuria and increase the regression rate to normoalbuminuria. Diabetic microvascular complications can be controlled with tight glycemic therapy, dyslipidemia management and blood pressure control along with renal function monitoring, lifestyle changes, including smoking cessation and low-protein diet. An integrated and personalized care would reduce the risk of development of microvascular complications in T1DM patients. The child with diabetes who receives limited care is more likely to develop long-term complications at an earlier age. Screening for subclinical complications and early interventions with intensive therapy is the need of the hour.

  15. Evaluation of gravimetric techniques to estimate the microvascular filtration coefficient.

    Science.gov (United States)

    Dongaonkar, R M; Laine, G A; Stewart, R H; Quick, C M

    2011-06-01

    Microvascular permeability to water is characterized by the microvascular filtration coefficient (K(f)). Conventional gravimetric techniques to estimate K(f) rely on data obtained from either transient or steady-state increases in organ weight in response to increases in microvascular pressure. Both techniques result in considerably different estimates and neither account for interstitial fluid storage and lymphatic return. We therefore developed a theoretical framework to evaluate K(f) estimation techniques by 1) comparing conventional techniques to a novel technique that includes effects of interstitial fluid storage and lymphatic return, 2) evaluating the ability of conventional techniques to reproduce K(f) from simulated gravimetric data generated by a realistic interstitial fluid balance model, 3) analyzing new data collected from rat intestine, and 4) analyzing previously reported data. These approaches revealed that the steady-state gravimetric technique yields estimates that are not directly related to K(f) and are in some cases directly proportional to interstitial compliance. However, the transient gravimetric technique yields accurate estimates in some organs, because the typical experimental duration minimizes the effects of interstitial fluid storage and lymphatic return. Furthermore, our analytical framework reveals that the supposed requirement of tying off all draining lymphatic vessels for the transient technique is unnecessary. Finally, our numerical simulations indicate that our comprehensive technique accurately reproduces the value of K(f) in all organs, is not confounded by interstitial storage and lymphatic return, and provides corroboration of the estimate from the transient technique.

  16. Delayed, bilateral, non-microvascular ear replantation after violent amputation.

    Science.gov (United States)

    García-Murray, E; Adán-Rivas, O; Salcido-Calzadilla, H

    2009-06-01

    Amputation of any body part is undoubtedly a traumatic experience leaving a terrible deformity, especially when the part or parts involved are visible and constitute an essential component of someone's facial whole. Bilateral ear amputation and successful subsequent replantation has been reported historically, but not in the modern surgical literature. We report the case of a 27-year-old female who was abducted and suffered a bilateral ear amputation at the hands of one of her captors to speed delivery of ransom money; the severed parts were sent to the parents approximately 2 hours after the amputation had taken place, and the girl was released some 48 hours after the ears were delivered. Microvascular replantation was attempted immediately after admission to the hospital some 2 hours after her release, but failed, and so a non-microvascular replantation was performed and was successful, after approximately 54 hours of ischaemia time. We consider this the first report of a complete bilateral, delayed, non-microvascular, successful ear replantation in a human being in the modern literature.

  17. Microvascular Abnormality in Schizophrenia as Shown by Retinal Imaging

    Science.gov (United States)

    Meier, Madeline H.; Shalev, Idan; Moffitt, Terrie E.; Kapur, Shitij; Keefe, Richard S.E.; Wong, Tien; Belsky, Daniel W.; Harrington, HonaLee; Hogan, Sean; Houts, Renate; Caspi, Avshalom; Poulton, Richie

    2013-01-01

    Objective Retinal and cerebral microvessels are structurally and functionally homologous, but, unlike cerebral microvessels, retinal microvessels can be noninvasively measured in vivo via retinal imaging. Here we test the hypothesis that individuals with schizophrenia show microvascular abnormality and evaluate the utility of retinal imaging as a tool for future schizophrenia research. Methods Participants were members of the Dunedin Study, a population-representative cohort followed from birth with 95% retention. Study members underwent retinal imaging at age 38 years. We assessed retinal arteriolar and venular caliber for all members of the cohort, including individuals who developed schizophrenia. Results Study members who developed schizophrenia were distinguished by wider retinal venules, suggesting microvascular abnormality reflective of insufficient brain oxygen supply. Analyses that controlled for confounding health conditions suggested that wider retinal venules are not simply an artifact of co-occurring health problems in schizophrenia patients. Wider venules were also associated with a dimensional measure of adult psychosis symptoms and with psychosis symptoms reported in childhood. Conclusions Findings provide initial support for the hypothesis that individuals with schizophrenia show microvascular abnormality. Moreover, results suggest that the same vascular mechanisms underlie subthreshold symptoms and clinical disorder and that these associations may begin early in life. These findings highlight the promise of retinal imaging as a tool for understanding the pathogenesis of schizophrenia. PMID:24030514

  18. [Microvascular anastomoses in reconstructive head and neck surgery].

    Science.gov (United States)

    Remmert, S

    1995-04-01

    At the ENT Department of the University of Lübeck, 57 microvascular tissue transplants with 129 anastomoses (61 arterial and 68 venous) have been performed in the last three years. Arteries have always been anastomosed end to end. The venous anastomoses have primarily been performed as end-to-side unions with the jugular vein. In nine patients, great distances between the donor and recipient vessel had to be connected with venous interponates. In two cases in which veins were lacking in the neck after radical neck dissection or radiation fibrosis, we used the cephalic vein or veins of the capsule of the thyroid gland as recipient vessels. The jejunal or osteomyocutaneous transplants were first fitted into the defect before performing the anastomosis. The jejunal peristalsis and the required freedom of movement in shaping the bone necessitated this technique. Microvascular anastomosis was first performed on the transplant of the radialis flap and the neurovascular infrahyoid muscular flap, and then they were integrated into the defect. We lost two transplants postoperatively because of venous thrombosis. In this article wie describe our anastomosis technique, the frequency distribution of recipient vessels, and the rules and characteristics of microvascular anastomosis after radiation and neck dissection.

  19. Microvascular Architecture of Hepatic Metastases in a Mouse Model

    Directory of Open Access Journals (Sweden)

    Darshini Kuruppu

    1997-01-01

    Full Text Available Development of effective treatment for hepatic metastases can be initiated by a better understanding of tumour vasculature and blood supply. This study was designed to characterise the microvascular architecture of hepatic metastases and observe the source of contributory blood supply from the host. Metastases were induced in mice by an intrasplenic injection of colon carcinoma cells (106 cells/ml. Vascularization of tumours was studied over a three week period by scanning electron microscopy of microvascular corrosion casts. Metastatic liver involvement was observed initially within a week post induction, as areas approximately 100 μm in diameter not perfused by the casting resin. On histology these spaces corresponded to tumour cell aggregates. The following weeks highlighted the angiogenesis phase of these tumours as they received a vascular supply from adjacent hepatic sinusoids. Direct sinusoidal supply of metastases was maintained throughout tumour growth. At the tumour periphery most sinusoids were compressed to form a sheath demarcating the tumour from the hepatic vasculature. No direct supply from the hepatic artery or the portal vein was observed. Dilated vessels termed vascular lakes dominated the complex microvascular architecture of the tumours, most tapering as they traversed towards the periphery. Four vascular branching patterns could be identified as true loops, bifurcations and trifurcations, spirals and capillary networks. The most significant observation in this study was the direct sinusoidal supply of metastases, together with the vascular lakes and the peripheral sinusoidal sheaths of the tumour microculature.

  20. Preventing microvascular complications in type 1 diabetes mellitus.

    Science.gov (United States)

    Viswanathan, Vijay

    2015-04-01

    Patients with complications of diabetes such as retinopathy, nephropathy, and cardiovascular complications have increased hospital stay with greater economic burden. Prevention of complications should be started before the onset of type 1 diabetes mellitus (T1DM) by working on risk factors and thereafter by intervention upon confirmatory diagnosis which can prevent further damage to β-cells. The actual risk of getting microvascular complications like microalbuminuria and retinopathy progression starts at glycated hemoglobin (HbA1c) level of 7%. As per the American Diabetes Association, a new pediatric glycemic control target of HbA1c 20 years as compared to patients <10 years of age. Screening of these complications should be done regularly, and appropriate preventive strategies should be followed. Angiotensin converting enzyme inhibitors and angiotensin II receptor blocker reduce progression from microalbuminuria to macroalbuminuria and increase the regression rate to normoalbuminuria. Diabetic microvascular complications can be controlled with tight glycemic therapy, dyslipidemia management and blood pressure control along with renal function monitoring, lifestyle changes, including smoking cessation and low-protein diet. An integrated and personalized care would reduce the risk of development of microvascular complications in T1DM patients. The child with diabetes who receives limited care is more likely to develop long-term complications at an earlier age. Screening for subclinical complications and early interventions with intensive therapy is the need of the hour.

  1. Fibroadenoma - breast

    Science.gov (United States)

    Breast lump - fibroadenoma; Breast lump - noncancerous; Breast lump - benign ... The cause of fibroadenomas is not known. There may be a connection to a problem with genes. Fibroadenoma is the most common benign ...

  2. Breast Implants

    Science.gov (United States)

    ... Medical Procedures Implants and Prosthetics Breast Implants Breast Implants Share Tweet Linkedin Pin it More sharing options Linkedin Pin it Email Print Breast implants are medical devices that are implanted under the ...

  3. Pericytes contribute to the disruption of the cerebral endothelial barrier via increasing VEGF expression: implications for stroke.

    Directory of Open Access Journals (Sweden)

    Ying Bai

    Full Text Available Disruption of the blood-brain barrier (BBB integrity occurring during the early onset of stroke is not only a consequence of, but also contributes to the further progression of stroke. Although it has been well documented that brain microvascular endothelial cells and astrocytes play a critical role in the maintenance of BBB integrity, pericytes, sandwiched between endothelial cells and astrocytes, remain poorly studied in the pathogenesis of stroke. Our findings demonstrated that treatment of human brain microvascular pericytes with sodium cyanide (NaCN and glucose deprivation resulted in increased expression of vascular endothelial growth factor (VEGF via the activation of tyrosine kinase Src, with downstream activation of mitogen activated protein kinase and PI3K/Akt pathways and subsequent translocation of NF-κB into the nucleus. Conditioned medium from NaCN-treated pericytes led to increased permeability of endothelial cells, and this effect was significantly inhibited by VEGF-neutralizing antibody. The in vivo relevance of these findings was further corroborated in the stroke model of mice wherein the mice, demonstrated disruption of the BBB integrity and concomitant increase in the expression of VEGF in the brain tissue as well as in the isolated microvessel. These findings thus suggest the role of pericyte-derived VEGF in modulating increased permeability of BBB during stroke. Understanding the regulation of VEGF expression could open new avenues for the development of potential therapeutic targets for stroke and other neurological disease.

  4. Differential roles for endothelial ICAM-1, ICAM-2, and VCAM-1 in shear-resistant T cell arrest, polarization, and directed crawling on blood-brain barrier endothelium.

    Science.gov (United States)

    Steiner, Oliver; Coisne, Caroline; Cecchelli, Roméo; Boscacci, Rémy; Deutsch, Urban; Engelhardt, Britta; Lyck, Ruth

    2010-10-15

    Endothelial ICAM-1 and ICAM-2 were shown to be essential for T cell diapedesis across the blood-brain barrier (BBB) in vitro under static conditions. Crawling of T cells prior to diapedesis was only recently revealed to occur preferentially against the direction of blood flow on the endothelial surface of inflamed brain microvessels in vivo. Using live cell-imaging techniques, we prove that Th1 memory/effector T cells predominantly crawl against the direction of flow on the surface of BBB endothelium in vitro. Analysis of T cell interaction with wild-type, ICAM-1-deficient, ICAM-2-deficient, or ICAM-1 and ICAM-2 double-deficient primary mouse brain microvascular endothelial cells under physiological flow conditions allowed us to dissect the individual contributions of endothelial ICAM-1, ICAM-2, and VCAM-1 to shear-resistant T cell arrest, polarization, and crawling. Although T cell arrest was mediated by endothelial ICAM-1 and VCAM-1, T cell polarization and crawling were mediated by endothelial ICAM-1 and ICAM-2 but not by endothelial VCAM-1. Therefore, our data delineate a sequential involvement of endothelial ICAM-1 and VCAM-1 in mediating shear-resistant T cell arrest, followed by endothelial ICAM-1 and ICAM-2 in mediating T cell crawling to sites permissive for diapedesis across BBB endothelium.

  5. Hypoxia/Aglycemia-induced endothelial barrier dysfunction and tight junction protein downregulation can be ameliorated by citicoline.

    Directory of Open Access Journals (Sweden)

    Xiaotang Ma

    Full Text Available This study explores the effect of citicoline on the permeability and expression of tight junction proteins (TJPs in endothelial cells under hypoxia/aglycemia conditions. Hypoxia or oxygen and glucose deprivation (OGD was utilized to induce endothelial barrier breakdown model on human umbilical vein endothelial cells (HUVECs and mouse brain microvascular endothelial cells (bEnd.3s. The effect of citicoline on endothelial barrier breakdown models was determined at either low or high concentrations. FITC-Dextran flux was used to examine the endothelial permeability. The expression of TJPs was measured by immunofluorescence, Real-time PCR and Western Blot methods. Results showed that hypoxia or OGD increased the permeability of HUVECs accompanied with down-regulation of occludens-1 (ZO-1 and occludin at both mRNA and protein levels. Similarly in bEnd.3s, hypoxia increased the permeability and decreased the expression of ZO-1 and claudin-5. Citicoline treatment dose-dependently decreased the permeability in these two models, which paralleled with elevated expression of TJPs. The data demonstrate that citicoline restores the barrier function of endothelial cells compromised by hypoxia/aglycemia probably via up-regulating the expression of TJPs.

  6. Hypoxia/Aglycemia-induced endothelial barrier dysfunction and tight junction protein downregulation can be ameliorated by citicoline.

    Science.gov (United States)

    Ma, Xiaotang; Zhang, Huiting; Pan, Qunwen; Zhao, Yuhui; Chen, Ji; Zhao, Bin; Chen, Yanfang

    2013-01-01

    This study explores the effect of citicoline on the permeability and expression of tight junction proteins (TJPs) in endothelial cells under hypoxia/aglycemia conditions. Hypoxia or oxygen and glucose deprivation (OGD) was utilized to induce endothelial barrier breakdown model on human umbilical vein endothelial cells (HUVECs) and mouse brain microvascular endothelial cells (bEnd.3s). The effect of citicoline on endothelial barrier breakdown models was determined at either low or high concentrations. FITC-Dextran flux was used to examine the endothelial permeability. The expression of TJPs was measured by immunofluorescence, Real-time PCR and Western Blot methods. Results showed that hypoxia or OGD increased the permeability of HUVECs accompanied with down-regulation of occludens-1 (ZO-1) and occludin at both mRNA and protein levels. Similarly in bEnd.3s, hypoxia increased the permeability and decreased the expression of ZO-1 and claudin-5. Citicoline treatment dose-dependently decreased the permeability in these two models, which paralleled with elevated expression of TJPs. The data demonstrate that citicoline restores the barrier function of endothelial cells compromised by hypoxia/aglycemia probably via up-regulating the expression of TJPs.

  7. What Is Breast Cancer?

    Science.gov (United States)

    ... Research? Breast Cancer About Breast Cancer What Is Breast Cancer? Breast cancer starts when cells in the breast ... spread, see our section on Cancer Basics . Where breast cancer starts Breast cancers can start from different parts ...

  8. Rickettsia massiliae and Rickettsia conorii Israeli Spotted Fever Strain Differentially Regulate Endothelial Cell Responses.

    Science.gov (United States)

    Bechelli, Jeremy; Smalley, Claire; Milhano, Natacha; Walker, David H; Fang, Rong

    2015-01-01

    Rickettsiae primarily target microvascular endothelial cells. However, it remains elusive how endothelial cell responses to rickettsiae play a role in the pathogenesis of rickettsial diseases. In the present study, we employed two rickettsial species with high sequence homology but differing virulence to investigate the pathological endothelial cell responses. Rickettsia massiliae is a newly documented human pathogen that causes a mild spotted fever rickettsiosis. The "Israeli spotted fever" strain of R. conorii (ISF) causes severe disease with a mortality rate up to 30% in hospitalized patients. At 48 hours post infection (HPI), R. conorii (ISF) induced a significant elevation of IL-8 and IL-6 while R. massiliae induced a statistically significant elevated amount of MCP-1 at both transcriptional and protein synthesis levels. Strikingly, R. conorii (ISF), but not R. massiliae, caused a significant level of cell death or injury in HMEC-1 cells at 72 HPI, demonstrated by live-dead cell staining, annexin V staining and lactate dehydrogenase release. Monolayers of endothelial cells infected with R. conorii (ISF) showed a statistically significant decrease in electrical resistance across the monolayer compared to both R. massiliae-infected and uninfected cells at 72 HPI, suggesting increased endothelial permeability. Interestingly, pharmacological inhibitors of caspase-1 significantly reduced the release of lactate dehydrogenase by R. conorii (ISF)-infected HMEC-1 cells, which suggests the role of caspase-1 in mediating the death of endothelial cells. Taken together, our data illustrated that a distinct proinflammatory cytokine profile and endothelial dysfunction, as evidenced by endothelial cell death/injury and increased permeability, are associated with the severity of rickettsial diseases.

  9. Dengue Virus NS1 Disrupts the Endothelial Glycocalyx, Leading to Hyperpermeability.

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    Henry Puerta-Guardo

    2016-07-01

    Full Text Available Dengue is the most prevalent arboviral disease in humans and a major public health problem worldwide. Systemic plasma leakage, leading to hypovolemic shock and potentially fatal complications, is a critical determinant of dengue severity. Recently, we and others described a novel pathogenic effect of secreted dengue virus (DENV non-structural protein 1 (NS1 in triggering hyperpermeability of human endothelial cells in vitro and systemic vascular leakage in vivo. NS1 was shown to activate toll-like receptor 4 signaling in primary human myeloid cells, leading to secretion of pro-inflammatory cytokines and vascular leakage. However, distinct endothelial cell-intrinsic mechanisms of NS1-induced hyperpermeability remained to be defined. The endothelial glycocalyx layer (EGL is a network of membrane-bound proteoglycans and glycoproteins lining the vascular endothelium that plays a key role in regulating endothelial barrier function. Here, we demonstrate that DENV NS1 disrupts the EGL on human pulmonary microvascular endothelial cells, inducing degradation of sialic acid and shedding of heparan sulfate proteoglycans. This effect is mediated by NS1-induced expression of sialidases and heparanase, respectively. NS1 also activates cathepsin L, a lysosomal cysteine proteinase, in endothelial cells, which activates heparanase via enzymatic cleavage. Specific inhibitors of sialidases, heparanase, and cathepsin L prevent DENV NS1-induced EGL disruption and endothelial hyperpermeability. All of these effects are specific to NS1 from DENV1-4 and are not induced by NS1 from West Nile virus, a related flavivirus. Together, our data suggest an important role for EGL disruption in DENV NS1-mediated endothelial dysfunction during severe dengue disease.

  10. Dengue Virus NS1 Disrupts the Endothelial Glycocalyx, Leading to Hyperpermeability

    Science.gov (United States)

    Puerta-Guardo, Henry; Glasner, Dustin R.; Harris, Eva

    2016-01-01

    Dengue is the most prevalent arboviral disease in humans and a major public health problem worldwide. Systemic plasma leakage, leading to hypovolemic shock and potentially fatal complications, is a critical determinant of dengue severity. Recently, we and others described a novel pathogenic effect of secreted dengue virus (DENV) non-structural protein 1 (NS1) in triggering hyperpermeability of human endothelial cells in vitro and systemic vascular leakage in vivo. NS1 was shown to activate toll-like receptor 4 signaling in primary human myeloid cells, leading to secretion of pro-inflammatory cytokines and vascular leakage. However, distinct endothelial cell-intrinsic mechanisms of NS1-induced hyperpermeability remained to be defined. The endothelial glycocalyx layer (EGL) is a network of membrane-bound proteoglycans and glycoproteins lining the vascular endothelium that plays a key role in regulating endothelial barrier function. Here, we demonstrate that DENV NS1 disrupts the EGL on human pulmonary microvascular endothelial cells, inducing degradation of sialic acid and shedding of heparan sulfate proteoglycans. This effect is mediated by NS1-induced expression of sialidases and heparanase, respectively. NS1 also activates cathepsin L, a lysosomal cysteine proteinase, in endothelial cells, which activates heparanase via enzymatic cleavage. Specific inhibitors of sialidases, heparanase, and cathepsin L prevent DENV NS1-induced EGL disruption and endothelial hyperpermeability. All of these effects are specific to NS1 from DENV1-4 and are not induced by NS1 from West Nile virus, a related flavivirus. Together, our data suggest an important role for EGL disruption in DENV NS1-mediated endothelial dysfunction during severe dengue disease. PMID:27416066

  11. A potent oral P-selectin blocking agent improves microcirculatory blood flow and a marker of endothelial cell injury in patients with sickle cell disease.

    Science.gov (United States)

    Kutlar, Abdullah; Ataga, Kenneth I; McMahon, Lillian; Howard, Joanna; Galacteros, Frederic; Hagar, Ward; Vichinsky, Elliott; Cheung, Anthony T W; Matsui, Neil; Embury, Stephen H

    2012-05-01

    Abnormal blood flow accounts for most of the clinical morbidity of sickle cell disease (SCD) [1,2]. Most notably, occlusion of flow in the microvasculature causes the acute pain crises [3] that are the commonest cause for patients with SCD to seek medical attention [4] and major determinants of their quality of life [5]. Based on evidence that endothelial P-selectin is central to the abnormal blood flow in SCD we provide results from four of our studies that are germane to microvascular blood flow in SCD. A proof-of-principle study established that doses of heparin lower than what are used for anticoagulation but sufficient to block P-selectin improved microvascular blood flow inpatients with SCD. An in vitro study showed that Pentosan Polysulfate Sodium (PPS) had greater P-selectin blocking activity than heparin. A Phase I clinical study demonstrated that a single oral dose of PPS increased microvascular blood flow in patients with SCD. A Phase II clinical study that was not completed documented that daily oral doses of PPS administered for 8 weeks lowered plasma levels of sVCAM-1 and tended to improve microvascular blood flow in patients with SCD. These data support the concept that P-selectin on the microvascular endothelium is critical to both acute vascular occlusion and chronically impaired microvascular blood flow in SCD. They also demonstrate that oral PPS is beneficial to microvascular sickle cell blood flow and has potential as an efficacious agent for long-term prophylactic therapy of SCD.

  12. Human Cytomegalovirus and Human Umbilical Vein Endothelial Cells: Restriction of Primary Isolation to Blood Samples and Susceptibilities of Clinical Isolates from Other Sources to Adaptation

    OpenAIRE

    2002-01-01

    In immunocompromised patients with disseminated infection, human cytomegalovirus (HCMV) is widespread in the microvascular endothelium of multiple organs. Human umbilical vein endothelial cells (HUVEC) were used in parallel to human embryonic lung fibroblasts (HELF) to recover HCMV from blood samples of immunocompromised patients. Using the shell vial technique, comparable median numbers of p72-positive HUVEC and HELF cells were found with the 26 HCMV-positive buffy coat samples out of 150 ex...

  13. Toll-like receptor 4 regulates early endothelial activation during ischemic acute kidney injury

    Science.gov (United States)

    Chen, Jianlin; John, Reji; Richardson, James A.; Shelton, John M.; Zhou, Xin J.; Wang, Yanxia; Wu, Qing Qing; Hartono, John R.; Winterberg, Pamela D.; Lu, Christopher Y.

    2012-01-01

    Ischemic acute kidney injury (AKI) triggers an inflammatory response which exacerbates injury that requires increased expression of endothelial adhesion molecules. To study this further, we used in situ hybridization, immunohistology, and isolated endothelial cells, and found increased Toll-like receptor 4 (TLR4) expression on endothelial cells of the vasa rectae of the inner stripe of the outer medulla of the kidney 4 h after reperfusion. This increase was probably due to reactive oxygen species, known to be generated early during ischemic AKI, because the addition of hydrogen peroxide increased TLR4 expression in MS1 microvascular endothelial cells in vitro. Endothelial TLR4 may regulate adhesion molecule (CD54 and CD62E) expression as they were increased on endothelia of wild-type but not TLR4 knockout mice in vivo. Further, the addition of high-mobility group protein B1, a TLR4 ligand released by injured cells, increased adhesion molecule expression on endothelia isolated from wild-type but not TLR4 knockout mice. TLR4 was localized to proximal tubules in the cortex and outer medulla after 24 h of reperfusion. Thus, at least two different cell types express TLR4, each of which contributes to renal injury by temporally different mechanisms during ischemic AKI. PMID:20927041

  14. Glioblastoma cell-secreted interleukin-8 induces brain endothelial cell permeability via CXCR2.

    Directory of Open Access Journals (Sweden)

    Julie Dwyer

    Full Text Available Glioblastoma constitutes the most aggressive and deadly of brain tumors. As yet, both conventional and molecular-based therapies have met with limited success in treatment of this cancer. Among other explanations, the heterogeneity of glioblastoma and the associated microenvironment contribute to its development, as well as resistance and recurrence in response to treatments. Increased vascularity suggests that tumor angiogenesis plays an important role in glioblastoma progression. However, the molecular crosstalk between endothelial and glioblastoma cells requires further investigation. To examine the effects of glioblastoma-derived signals on endothelial homeostasis, glioblastoma cell secretions were collected and used to treat brain endothelial cells. Here, we present evidence that the glioblastoma secretome provides pro-angiogenic signals sufficient to disrupt VE-cadherin-mediated cell-cell junctions and promote endothelial permeability in brain microvascular endothelial cells. An unbiased angiogenesis-specific antibody array screen identified the chemokine, interleukin-8, which was further demonstrated to function as a key factor involved in glioblastoma-induced permeability, mediated through its receptor CXCR2 on brain endothelia. This underappreciated interface between glioblastoma cells and associated endothelium may inspire the development of novel therapeutic strategies to induce tumor regression by preventing vascular permeability and inhibiting angiogenesis.

  15. Mononuclear Phagocyte-Derived Microparticulate Caspase-1 Induces Pulmonary Vascular Endothelial Cell Injury.

    Directory of Open Access Journals (Sweden)

    Srabani Mitra

    Full Text Available Lung endothelial cell apoptosis and injury occurs throughout all stages of acute lung injury (ALI/ARDS and impacts disease progression. Lung endothelial injury has traditionally been focused on the role of neutrophil trafficking to lung vascular integrin receptors induced by proinflammatory cytokine expression. Although much is known about the pathogenesis of cell injury and death in ALI/ARDS, gaps remain in our knowledge; as a result of which there is currently no effective pharmacologic therapy. Enzymes known as caspases are essential for completion of the apoptotic program and secretion of pro-inflammatory cytokines. We hypothesized that caspase-1 may serve as a key regulator of human pulmonary microvascular endothelial cell (HPMVEC apoptosis in ALI/ARDS. Our recent experiments confirm that microparticles released from stimulated monocytic cells (THP1 induce lung endothelial cell apoptosis. Microparticles pretreated with the caspase-1 inhibitor, YVAD, or pan-caspase inhibitor, ZVAD, were unable to induce cell death of HPMVEC, suggesting the role of caspase-1 or its substrate in the induction of HPMVEC cell death. Neither un-induced microparticles (control nor direct treatment with LPS induced apoptosis of HPMVEC. Further experiments showed that caspase-1 uptake into HPMVEC and the induction of HPMVEC apoptosis was facilitated by caspase-1 interactions with microparticulate vesicles. Altering vesicle integrity completely abrogated apoptosis of HPMVEC suggesting an encapsulation requirement for target cell uptake of active caspase-1. Taken together, we confirm that microparticle centered caspase-1 can play a regulator role in endothelial cell injury.

  16. Aberrant production of extracellular matrix proteins and dysfunction in kidney endothelial cells with a short duration of diabetes.

    Science.gov (United States)

    Grutzmacher, Cathy; Park, SunYoung; Zhao, Yun; Morrison, Margaret E; Sheibani, Nader; Sorenson, Christine M

    2013-01-01

    Diabetic nephropathy is the most common cause of end-stage renal disease and is a major risk factor for cardiovascular disease. In the United States, microvascular complications during diabetic nephropathy contribute to high morbidity and mortality rates. However, the cell-autonomous impact of diabetes on kidney endothelial cell function requires further investigation. Male Akita/+ [autosomal dominant mutation in the insulin II gene (Ins2)] mice reproducibly develop diabetes by 4 wk of age. Here, we examined the impact a short duration of diabetes had on kidney endothelial cell function. Kidney endothelial cells were prepared from nondiabetic and diabetic mice (4 wk of diabetes) to delineate the early changes in endothelial cell function. Kidney endothelial cells from Akita/+ mice following 4 wk of diabetes demonstrated aberrant expression of extracellular matrix proteins including decreased osteopontin and increased fibronectin expression which correlated with increased α5-integrin expression. These changes were associated with the attenuation of migration and capillary morphogenesis. Kidney endothelial cells from Akita/+ mice had decreased VEGF levels but increased levels of endothelial nitric oxide synthase(eNOS) and NO, suggesting uncoupling of VEGF-mediated NO production. Knocking down eNOS expression in Akita/+ kidney endothelial cells increased VEGF expression, endothelial cell migration, and capillary morphogenesis. Furthermore, attenuation of sprouting angiogenesis of aortas from Akita/+ mice with 8 wk of diabetes was restored in the presence of the antioxidant N-acetylcysteine. These studies demonstrate that aberrant endothelial cell function with a short duration of diabetes may set the stage for vascular dysfunction and rarefaction at later stages of diabetes.

  17. Method for Dual Viral Vector Mediated CRISPR-Cas9 Gene Disruption in Primary Human Endothelial Cells

    Science.gov (United States)

    Gong, Haixia; Liu, Menglin; Klomp, Jeff; Merrill, Bradley J.; Rehman, Jalees; Malik, Asrar B.

    2017-01-01

    Human endothelial cells (ECs) are widely used to study mechanisms of angiogenesis, inflammation, and endothelial permeability. Targeted gene disruption induced by Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-CRISPR-Associated Protein 9 (Cas9) nuclease gene editing is potentially an important tool for definitively establishing the functional roles of individual genes in ECs. We showed that co-delivery of adenovirus encoding EGFP-tagged Cas9 and lentivirus encoding a single guide RNA (sgRNA) in primary human lung microvascular ECs (HLMVECs) disrupted the expression of the Tie2 gene and protein. Tie2 disruption increased basal endothelial permeability and prevented permeability recovery following injury induced by the inflammatory stimulus thrombin. Thus, gene deletion via viral co-delivery of CRISPR-Cas9 in primary human ECs provides a novel platform to investigate signaling mechanisms of normal and perturbed EC function without the need for clonal expansion. PMID:28198371

  18. Adapting coculture in vitro models of the blood-brain barrier for use in cancer research: maintaining an appropriate endothelial monolayer for the assessment of transendothelial migration.

    Science.gov (United States)

    Vandenhaute, Elodie; Drolez, Aurore; Sevin, Emmanuel; Gosselet, Fabien; Mysiorek, Caroline; Dehouck, Marie-Pierre

    2016-05-01

    Although brain metastases are the most common brain tumors in adults, there are few treatment options in this setting. To colonize the brain, circulating tumor cells must cross the blood-brain barrier (BBB), which is situated within specialized, restrictive microvascular endothelium. Understanding how cancer cells manage to transmigrate through the BBB might enable this process to be prevented. In vitro models are dedicated tools for characterizing the cellular and molecular mechanisms that underlie transendothelial migration process, as long as they accurately mimic the brain endothelium's in vivo characteristics. The objective of the present study was to adapt an existing in vitro model of the human BBB for use in studying cancer cell transmigration. The model is based on the coculture of endothelial cells (ECs, derived from cord blood hematopoietic stem cells) and brain pericytes. To allow the migration of cancer cells into the lower compartment, our model had to be transposed onto inserts with a larger pore size. However, we encountered a problem when culturing ECs on large (3-μm)-pore inserts: the cells crossed the membrane and formed a non-physiological second layer on the lower face of the insert. Using 3-μm-pore inserts (in a 12-well plate format), we report here on a method that enables the maintenance of a single monolayer of ECs on the insert's upper face only. Under these chosen conditions, the ECs exhibited typical BBB properties found in the original model (including restricted paracellular permeability and the expression of continuous tight junctions). This modified in vitro model of the human BBB enabled us to investigate the migratory potential of the MDA-MB-231 cell line (derived from highly metastatic human breast cancer cells). Last, the results obtained were compared with the rate of transmigration through endothelia with no BBB features.

  19. Leishmaniosis--a report about the microvascular and cellular architecture of the infected spleen in Canis familiaris.

    Science.gov (United States)

    Alexandre-Pires, G; Pais, D; Correia, M; Pina, J A Esperança

    2006-04-01

    Leishmaniosis is an anthropozoonosis caused by an intracellular protozoan parasite that causes a wide spectrum of diseases in humans and dogs worldwide. In the Mediterranean basin, Portugal, Central and South America, and in the Middle East, visceral leishmaniosis is caused by Leishmania infantum. In these areas, dogs are believed to be the natural reservoirs of this parasite. In the case of visceral leishmaniosis, the spleen is one of the several hematopoietic and immunocompetent organs involved. Since this viscera is a blood filter, the authors investigated the expression of the morphological and microvascular environment and modifications of the spleen cell population related to immunological responses to this parasitic condition. The tools used to perform this study were scanning electronic microscopy of intact tissue and corrosion casts, transmission electronic microscopy, histology and immunohistochemistry. The results reveal three important modifications concerning the spleen's microvascular architecture when compared with its normal pattern, independently of the serological titer obtained with indirect immunofluorescence. (1) A marked scarcity of the sinusoidal system sheet that surrounds the central artery/arteriole of the white pulp; (2) A huge development of pulp venules and veins; (3) The presence of a surprising development of reticular fibers. The authors postulate that independent of the virulence of the parasite involved and the type of immunity prevalent in a particular host, the spleen develops blood dynamic conditions that permit reduction in the speed of blood flow so that cells involved in immunological processes can proliferate and differentiate, and also contributes to trapping lymphocytes within the area through the differentiation of characteristics that resemble those of HEV endothelial cells.

  20. Complementary populations of human adipose CD34+ progenitor cells promote growth, angiogenesis, and metastasis of breast cancer.

    Science.gov (United States)

    Orecchioni, Stefania; Gregato, Giuliana; Martin-Padura, Ines; Reggiani, Francesca; Braidotti, Paola; Mancuso, Patrizia; Calleri, Angelica; Quarna, Jessica; Marighetti, Paola; Aldeni, Chiara; Pruneri, Giancarlo; Martella, Stefano; Manconi, Andrea; Petit, Jean-Yves; Rietjens, Mario; Bertolini, Francesco

    2013-10-01

    Obesity is associated with an increased frequency, morbidity, and mortality of several types of neoplastic diseases, including postmenopausal breast cancer. We found that human adipose tissue contains two populations of progenitors with cooperative roles in breast cancer. CD45(-)CD34(+)CD31(+)CD13(-)CCRL2(+) endothelial cells can generate mature endothelial cells and capillaries. Their cancer-promoting effect in the breast was limited in the absence of CD45(-)CD34(+)CD31(-)CD13(+)CD140b(+) mesenchymal progenitors/adipose stromal cells (ASC), which generated pericytes and were more efficient than endothelial cells in promoting local tumor growth. Both endothelial cells and ASCs induced epithelial-to-mesenchymal transition (EMT) gene expression in luminal breast cancer cells. Endothelial cells (but not ASCs) migrated to lymph nodes and to contralateral nascent breast cancer lesions where they generated new vessels. In vitro and in vivo, endothelial cells were more efficient than ASCs in promoting tumor migration and in inducing metastases. Granulocyte colony-stimulating factor (G-CSF) effectively mobilized endothelial cells (but not ASCs), and the addition of chemotherapy and/or of CXCR4 inhibitors did not increase endothelial cell or ASC blood mobilization. Our findings suggest that adipose tissue progenitor cells cooperate in driving progression and metastatic spread of breast cancer.

  1. Breast Diseases

    Science.gov (United States)

    ... bumps, and discharges (fluids that are not breast milk). If you have a breast lump, pain, discharge or skin irritation, see your health care provider. Minor and serious breast problems have similar symptoms. Although many women fear cancer, most breast problems are not cancer. Some common ...

  2. Brain microvascular pericytes are immunoactive in culture: cytokine, chemokine, nitric oxide, and LRP-1 expression in response to lipopolysaccharide

    Directory of Open Access Journals (Sweden)

    Erickson Michelle A

    2011-10-01

    Full Text Available Abstract Background Brain microvascular pericytes are important constituents of the neurovascular unit. These cells are physically the closest cells to the microvascular endothelial cells in brain capillaries. They significantly contribute to the induction and maintenance of the barrier functions of the blood-brain barrier. However, very little is known about their immune activities or their roles in neuroinflammation. Here, we focused on the immunological profile of brain pericytes in culture in the quiescent and immune-challenged state by studying their production of immune mediators such as nitric oxide (NO, cytokines, and chemokines. We also examined the effects of immune challenge on pericyte expression of low density lipoprotein receptor-related protein-1 (LRP-1, a protein involved in the processing of amyloid precursor protein and the brain-to-blood efflux of amyloid-β peptide. Methods Supernatants were collected from primary cultures of mouse brain pericytes. Release of nitric oxide (NO was measured by the Griess reaction and the level of S-nitrosylation of pericyte proteins measured with a modified "biotin-switch" method. Specific mitogen-activated protein kinase (MAPK pathway inhibitors were used to determine involvement of these pathways on NO production. Cytokines and chemokines were analyzed by multianalyte technology. The expression of both subunits of LRP-1 was analyzed by western blot. Results Lipopolysaccharide (LPS induced release of NO by pericytes in a dose-dependent manner that was mediated through MAPK pathways. Nitrative stress resulted in S-nitrosylation of cellular proteins. Eighteen of twenty-three cytokines measured were released constitutively by pericytes or with stimulation by LPS, including interleukin (IL-12, IL-13, IL-9, IL-10, granulocyte-colony stimulating factor, granulocyte macrophage-colony stimulating factor, eotaxin, chemokine (C-C motif ligand (CCL-3, and CCL-4. Pericyte expressions of both subunits of

  3. Insights into the molecular mechanisms of diabetes-induced endothelial dysfunction

    DEFF Research Database (Denmark)

    Saad, Mohamed I.; Abdelkhalek, Taha M.; Saleh, Moustafa M.;

    2015-01-01

    to be 600 million people by 2035. Diabetes is associated with microvascular and macrovascular complications resulting in accelerated endothelial dysfunction (ED), atherosclerosis, and cardiovascular disease (CVD). Unfortunately, the complex pathophysiology of diabetic cardiovascular damage is not fully......Diabetes mellitus is a heterogeneous, multifactorial, chronic disease characterized by hyperglycemia owing to insulin insufficiency and insulin resistance (IR). Recent epidemiological studies showed that the diabetes epidemic affects 382 million people worldwide in 2013, and this figure is expected......-associated metabolic disturbances (IR, subclinical inflammation, dyslipidemia, hyperglycemia, dysregulated production of adipokines, defective incretin and gut hormones production/action, and oxidative stress) and ED, focusing on oxidative stress and endothelial progenitor cells (EPCs). In addition, we re...

  4. EFFECT OF APLYSIN ON PROLIFERATION AND VASCULAR ENDOTHELIAL GROWTH FACTOR EXPRESSION IN HUMAN BREAST CANCER CELL LINE%海兔素对人乳腺癌细胞增殖及血管内皮生长因子表达的影响

    Institute of Scientific and Technical Information of China (English)

    贺娟; 梁惠; 韩磊; 马爱国; 孙永叶

    2009-01-01

    目的 观察海兔素(aplysin)对人乳腺癌细胞系MCF-7细胞增殖和血管内皮生长因子(VEGF)表达的抑制作用,以及对二甲基苯蒽(DMBA)诱导乳腺癌大鼠肿瘤组织中VEGF表达的抑制作用.方法 用MTT法和ELISA法分别检测海兔素对MCF-7细胞增殖及VEGF表达的影响.用免疫组织化学法检测海兔素对DMBA诱导的大鼠乳腺癌组织中VEGF的影响.结果 海兔素体外对MCF-7细胞有明显的增殖抑制作用,并伴随VEGF表达水平下降.体内可有效抑制大鼠乳腺癌组织中VEGF的表达,且与海兔素浓度有关.结论 海兔素体外对MCF-7细胞具有明显的细胞毒作用并能抑制其VEGF的表达,体内可有效抑制乳腺癌组织中VEGF的表达.%Objective To assess the effects of aplysin on proliferation inhibition and vascular endothelial growth factor (VEGF) level in human breast carcinoma (MCF-7) cells in vitro and to investigate the VEGF level of the brest cancer tissue induced by DMBA in rats. Method After MCF-7 cells were treated with aplysin from 10 to 40μg/ml for 48h and 96h, cell viability of proliferation was assessed by MTT assay and VEGF level was assessed by ELISA. Seventy-five female Wistar rats were randomly divided into five groups and were daily given 20mg/kg-bw(A group),40mg/kg-bw (B group), 80mg/kg-bw (C group) aplysin, and salad oil for both D and E groups. The rats in A, B, C and D groups were induced breast cancer by DMBA, a carcinogen, and those in E group were free from DMBA during 16 w treatment. VEGF level of the brest cancer tissue was assessed by immunohistochemistry. Results Within the concentration of 10-40 μg/ml, aplysin strongly inhibited proliferation of MCF-7 cells, accompanied by decrease in VEGF level. Aplysin decreased expression of VEGF in DMBA-induced breast cancer tissue of rats. Conclusion Aplysin could inhibit breast cancer cells with cytotoxicity and VEGF expression. Aplysin could decrease the expression of VEGF in breast cancer tissue

  5. Vitamin D levels and microvascular complications in type 2 diabetes

    Directory of Open Access Journals (Sweden)

    Sarita Bajaj

    2014-01-01

    Full Text Available Background: Vitamin D has important actions on glucose metabolism. These include improved insulin exocytosis, direct stimulation of insulin receptor, improved uptake of glucose by peripheral tissues, improving insulin resistance. It has got various pleiotropic effects like suppression of cell mediated immunity, regulation of cell proliferation, stimulation of neurotropic factors such as nerve growth factor, Glial cell line-derived neurotrophic factor, neurotropin, suppression of RAAS, reduction of albuminuria, immunomodulatory effects, and anti-inflammatory effects. Thus, vitamin D is implicated in many ways in the pathogenesis of retinopathy, neuropathy and nephropathy. Objectives: To study the correlation of vitamin D levels with microvascular complications in type 2 diabetes. Materials and Methods: Cross-sectional case-control study of 18 patients (18-70 years, who met the American Diabetes Association 2011 criteria for type 2 diabetes, was conducted. Age and sex matched healthy controls were taken. Subjects were evaluated for the presence of microvascular complications by clinical evaluation, urine examination, fundus examination, nerve conduction studies, and various biochemical tests. 25-OH cholecalciferol levels were done for each. Cut off level for vitamin D deficiency was 20 ng/ml. Results: Mean vitamin D was lower in type 2 diabetics than healthy subjects (19.046 vs. 27.186 ng/ml. Prevalence of vitamin D deficiency and insufficiency was found to significantly higher in diabetics when compared to healthy subjects (P = 0.0001. Vitamin D deficiency was found to be significantly associated with neuropathy (χ2 = 5.39, df = 1, P = 0.020, retinopathy, (χ2 = 6.6, df = 1, P = 0.010 and nephropathy (χ2 = 10. 52, df = 1, P = 0.001. Lower levels of vitamin D were found to be associated with increasing prevalence of combinations of microvascular complications namely neuropathy with retinopathy (P = 0.036, neuropathy with nephropathy (P = 0

  6. Skeletal muscle microvascular function in girls with Turner syndrome

    Science.gov (United States)

    West, Sarah L.; O'Gorman, Clodagh S.; Elzibak, Alyaa H.; Caterini, Jessica; Noseworthy, Michael D.; Rayner, Tammy; Hamilton, Jill; Wells, Greg D.

    2014-01-01

    Background Exercise intolerance is prevalent in individuals with Turner Syndrome (TS). We recently demonstrated that girls with TS have normal aerobic but altered skeletal muscle anaerobic metabolism compared to healthy controls (HC). The purpose of this study was to compare peripheral skeletal muscle microvascular function in girls with TS to HC after exercise. We hypothesized that girls with TS would have similar muscle blood-oxygen level-dependent (BOLD) magnetic resonance imaging (MRI) signal responses during recovery from exercise compared to HC. Methods Thirteen TS participants and 8 HC completed testing. BOLD MRI was used to measure skeletal muscle microvascular response during 60 second recovery, following 60 s of exercise at 65% of maximal workload. Exercise and recovery were repeated four times, and the BOLD signal time course was fit to a four-parameter sigmoid function. Results Participants were 13.7 ± 3.1 years old and weighed 47.9 ± 14.6 kg. The mean change in BOLD signal intensity following exercise at the end of recovery, the mean response time of the function/the washout of deoxyhemoglobin, and the mean half-time of recovery were similar between the TS and HC groups. Conclusions Our results demonstrate that compared to HC, peripheral skeletal muscle microvascular function following exercise in girls with TS is not impaired. General significance This study supports the idea that the aerobic energy pathway is not impaired in children with TS in response to submaximal exercise. Other mechanisms are likely responsible for exercise intolerance in TS; this needs to be further investigated. PMID:26676172

  7. Endothelial Barrier Protection by Local Anesthetics: Ropivacaine and Lidocaine Block Tumor Necrosis Factor-α–induced Endothelial Cell Src Activation

    Science.gov (United States)

    Piegeler, Tobias; Votta-Velis, E. Gina; Bakhshi, Farnaz R.; Mao, Mao; Carnegie, Graeme; Bonini, Marcelo G.; Schwartz, David E.; Borgeat, Alain; Beck-Schimmer, Beatrice; Minshall, Richard D.

    2014-01-01

    Background Pulmonary endothelial barrier dysfunction mediated in part by Src-kinase activation plays a crucial role in acute inflammatory disease. Proinflammatory cytokines, such as tumor necrosis factor-α (TNFα), activate Src via phosphatidylinositide 3-kinase/Akt-dependent nitric oxide generation, a process initiated by recruitment of phosphatidylinositide 3-kinase regulatory subunit p85 to TNF-receptor-1. Because amide-linked local anesthetics have well-established anti-inflammatory effects, the authors hypothesized that ropivacaine and lidocaine attenuate inflammatory Src signaling by disrupting the phosphatidylinositide 3-kinase–Akt–nitric oxide pathway, thus blocking Src-dependent neutrophil adhesion and endothelial hyperpermeability. Methods Human lung microvascular endothelial cells, incubated with TNFα in the absence or presence of clinically relevant concentrations of ropivacaine and lidocaine, were analyzed by Western blot, probing for phosphorylated/activated Src, endothelial nitric oxide synthase, Akt, intercellular adhesion molecule-1, and caveolin-1. The effect of ropivacaine on TNFα-induced nitric oxide generation, co-immunoprecipitation of TNF-receptor-1 with p85, neutrophil adhesion, and endothelial barrier disruption were assessed. Results Ropivacaine and lidocaine attenuated TNFα-induced Src activation (half-maximal inhibitory concentration [IC50] = 8.611 × 10−10 M for ropivacaine; IC50 = 5.864 × 10−10 M for lidocaine) and endothelial nitric oxide synthase phosphorylation (IC50 = 7.572 × 10−10 M for ropivacaine; IC50 = 6.377 × 10−10 M for lidocaine). Akt activation (n = 7; P = 0.006) and stimulus-dependent binding of TNF-receptor-1 and p85 (n = 6; P = 0.043) were blocked by 1 nM of ropivacaine. TNFα-induced neutrophil adhesion and disruption of endothelial monolayers via Src-dependent intercellular adhesion molecule-1- and caveolin-1-phosphorylation, respectively, were also attenuated. Conclusions Ropivacaine and lidocaine

  8. Sleep quality and duration are related to microvascular function: the Amsterdam Growth and Health Longitudinal Study.

    Science.gov (United States)

    Bonsen, Thomas; Wijnstok, Nienke J; Hoekstra, Trynke; Eringa, Etto C; Serné, Erik H; Smulders, Yvo M; Twisk, Jos W R

    2015-04-01

    Sleep and sleep disorders are related to cardiovascular disease, and microvascular function is an early cardiovascular disease marker. Therefore, the relationship of sleep (measured in sleep quality and duration) with microvascular function was examined in healthy adults. Sleep quality was assessed with the validated Sleep Wake Experience List (SWEL) questionnaire. Duration of sleep was self-reported in an additional question. Microvascular function was measured using nailfold capillaroscopy. Linear regression analyses were used to examine the relationship between sleep and microvascular function. Potential confounders included physical activity, smoking, blood pressure, body mass index and several biochemical parameters. Analyses were performed in 259 participants (116 men). For women reporting insufficient (sleep duration, microvascular function (post-ischaemic capillary recruitment) was significantly lower (b = -11.17; P = 0.04) compared to women reporting sufficient sleep duration. There was no relationship between sleep quality and microvascular function in females. In males, a trend towards lower capillary recruitment was found in those reporting a combination of poor sleep quality and insufficient duration (b = -7.54; P = 0.09), compared to those reporting good sleep quality as well as sufficient duration. This study suggests an association between sleep and microvascular function. Which aspects of sleep exactly affect microvascular function, and if indeed the association is different between males and females in other samples, needs further research.

  9. Correlation of ultrasound features with the expression of vascular endothelial growth factor, cyclooxygenase-2, metalloprotease-9 in breast cancer%血管内皮生长因子、环氧化酶-2和基质金属蛋白酶-9在乳腺癌中的表达与超声征象的相关性研究

    Institute of Scientific and Technical Information of China (English)

    冉张申; 王克强; 李湘奇

    2015-01-01

    Objective By comparing with the relationship between important signs on ultrasound in breast cancer and the protein expression of vascular endothelial growth factor (VEGF),cyclooxygenase-2 (COX-2),metalloprotease-9 (MMP-9) biology,the relationship between molecular pathology and ultrasound features were studied,which provided a certain theoretical basis for the early diagnosis,treatment and prognosis of breast cancer patients.Methods 89 cases of breast lesions were confirmed by ultrasonic examination,operation and pathology.By using double-blind method,ultrasound features were grouped into burr,calcification,abnormal vascular,lymph node by physicians with rich breast diagnosis experience based on BI-RADS classification method.In each case,the expression of VEGF,COX-2,MMP-9 were detected by immunohistochemical SP method,then the relationship between the expression of VEGF,COX-2,MMP-9 and breast ultrasound features were analyzed.Results positive rate of protein expression of VEGF,COX-2,MMP-9 in burr syndrome group were higher than that of no-burr syndrome group (P < 0.05),no obvious difference(P > 0.05) between the calcification group and no calcification group,higher in abnormal breast cancer group than that of no-abnormal blood vessels (P < 0.05),higher in lymph node metastasis group than that of no-lymph node metastasis (P < 0.05).Conclusions The high expression of VEGF,COX-2,MMP-9 could have an influence on burr syndrome,abnormal blood vessels,lymph node metastasis in breast cancer ultrasound features,which could be used as a biological basis of malignant signs in ultrasound of breast cancer.And at the same time,it's prompted that the breast cancer patients with ultrasound features is high in malignant degree and with poor prognosis.%目的 通过比较乳腺癌组织中血管内皮生长因子(VEGF)、环氧化酶-2(COX-2)、基质金属蛋白酶-9(MMP-9)三种蛋白的表达与乳腺癌超声重要征象的关系,初步探讨乳腺癌超声征象与分

  10. Atrial natriuretic peptide prevents cancer metastasis through vascular endothelial cells.

    Science.gov (United States)

    Nojiri, Takashi; Hosoda, Hiroshi; Tokudome, Takeshi; Miura, Koichi; Ishikane, Shin; Otani, Kentaro; Kishimoto, Ichiro; Shintani, Yasushi; Inoue, Masayoshi; Kimura, Toru; Sawabata, Noriyoshi; Minami, Masato; Nakagiri, Tomoyuki; Funaki, Soichiro; Takeuchi, Yukiyasu; Maeda, Hajime; Kidoya, Hiroyasu; Kiyonari, Hiroshi; Shioi, Go; Arai, Yuji; Hasegawa, Takeshi; Takakura, Nobuyuki; Hori, Megumi; Ohno, Yuko; Miyazato, Mikiya; Mochizuki, Naoki; Okumura, Meinoshin; Kangawa, Kenji

    2015-03-31

    Most patients suffering from cancer die of metastatic disease. Surgical removal of solid tumors is performed as an initial attempt to cure patients; however, surgery is often accompanied with trauma, which can promote early recurrence by provoking detachment of tumor cells into the blood stream or inducing systemic inflammation or both. We have previously reported that administration of atrial natriuretic peptide (ANP) during the perioperative period reduces inflammatory response and has a prophylactic effect on postoperative cardiopulmonary complications in lung cancer surgery. Here we demonstrate that cancer recurrence after curative surgery was significantly lower in ANP-treated patients than in control patients (surgery alone). ANP is known to bind specifically to NPR1 [also called guanylyl cyclase-A (GC-A) receptor]. In mouse models, we found that metastasis of GC-A-nonexpressing tumor cells (i.e., B16 mouse melanoma cells) to the lung was increased in vascular endothelium-specific GC-A knockout mice and decreased in vascular endothelium-specific GC-A transgenic mice compared with control mice. We examined the effect of ANP on tumor metastasis in mice treated with lipopolysaccharide, which mimics systemic inflammation induced by surgical stress. ANP inhibited the adhesion of cancer cells to pulmonary arterial and micro-vascular endothelial cells by suppressing the E-selectin expression that is promoted by inflammation. These results suggest that ANP prevents cancer metastasis by inhibiting the adhesion of tumor cells to inflamed endothelial cells.

  11. Cryptogenic stroke following abdominal free flap breast reconstruction surgery

    Science.gov (United States)

    Xie, Huizhuang; Malata, Charles M.

    2014-01-01

    INTRODUCTION Abdominal free flap breast reconstruction is regarded as the gold standard method of post-mastectomy breast reconstruction by many. It is a major surgery which can be associated with varied systemic complications. To date, there have been no reports of cerebrovascular complications in the literature which examine the possible relation between thromboembolism and patent foramen ovale (PFO) in patients undergoing microvascular breast reconstruction. PRESENTATION OF CASE A 54-year old female with a pre-existing PFO developed a stroke following bilateral mastectomies and immediate free flap breast reconstruction on postoperative day 5. This was attributed to an air embolus caused by central venous pressure line removal. After uneventful intra and early postoperative periods, the patient had collapsed suddenly on day 5 and become unresponsive immediately following the removal of a central venous line. Brain magnetic resonance imaging confirmed a cerebrovascular accident. This resolved within 48 h following therapeutic heparinisation. A clinical diagnosis of paradoxical embolism was made and she was subsequently referred to the cardiologists for angiographic closure of the PFO. DISCUSSION The case study herein reported gives an account that PFO can have considerable health implications in the early postoperative period and conceivably intraoperatively in patients undergoing major reconstructive surgeries. CONCLUSION Surgeons and cardiologists should be aware of this cerebrovascular complication secondary to PFO following major reconstructive surgery such as microvascular breast reconstruction. It also serves to challenge microvascular surgeons to reconsider routine use of central venous pressure lines in free flap patients who might otherwise have good peripheral vessels for postoperative fluid and antibiotic administration. PMID:25437687

  12. Delayed increases in microvascular pathology after experimental traumatic brain injury are associated with prolonged inflammation, blood-brain barrier disruption, and progressive white matter damage.

    Science.gov (United States)

    Glushakova, Olena Y; Johnson, Danny; Hayes, Ronald L

    2014-07-01

    Traumatic brain injury (TBI) is a significant risk factor for chronic traumatic encephalopathy (CTE), Alzheimer's disease (AD), and Parkinson's disease (PD). Cerebral microbleeds, focal inflammation, and white matter damage are associated with many neurological and neurodegenerative disorders including CTE, AD, PD, vascular dementia, stroke, and TBI. This study evaluates microvascular abnormalities observed at acute and chronic stages following TBI in rats, and examines pathological processes associated with these abnormalities. TBI in adult rats was induced by controlled cortical impact (CCI) of two magnitudes. Brain pathology was assessed in white matter of the corpus callosum for 24 h to 3 months following injury using immunohistochemistry (IHC). TBI resulted in focal microbleeds that were related to the magnitude of injury. At the lower magnitude of injury, microbleeds gradually increased over the 3 month duration of the study. IHC revealed TBI-induced focal abnormalities including blood-brain barrier (BBB) damage (IgG), endothelial damage (intercellular adhesion molecule 1 [ICAM-1]), activation of reactive microglia (ionized calcium binding adaptor molecule 1 [Iba1]), gliosis (glial fibrillary acidic protein [GFAP]) and macrophage-mediated inflammation (cluster of differentiation 68 [CD68]), all showing different temporal profiles. At chronic stages (up to 3 months), apparent myelin loss (Luxol fast blue) and scattered deposition of microbleeds were observed. Microbleeds were surrounded by glial scars and co-localized with CD68 and IgG puncta stainings, suggesting that localized BBB breakdown and inflammation were associated with vascular damage. Our results indicate that evolving white matter degeneration following experimental TBI is associated with significantly delayed microvascular damage and focal microbleeds that are temporally and regionally associated with development of punctate BBB breakdown and progressive inflammatory responses. Increased

  13. Cross-Talk between CLL Cells and Bone Marrow Endothelial Cells: Role of Signal Transducer and Activator of Transcription-3

    Science.gov (United States)

    Badoux, Xavier; Bueso-Ramos, Carlos; Harris, David; Li, Ping; Liu, Zhiming; Burger, Jan; O’Brien, Susan; Ferrajoli, Alessandra; Keating, Michael J.; Estrov, Zeev

    2014-01-01

    Summary Chronic lymphocytic leukemia (CLL) bone marrow is characterized by increased angiogenesis. However, the molecular mediators of neovascularization and the biological significance of increased endothelial cell proliferation in CLL require further investigation. Because signal transducer and activator of transcription (STAT)-3 is constitutively activated in CLL we studied the role of STAT3 in modulating vascular endothelial growth factor (VEGF) expression and the effect of vascular endothelial cells on CLL cells. Using chromatin immunoprecipitation (ChIP) we found that anti-STAT3 antibodies immunoprecipitated DNA of STAT3, VEGF and other STAT3-regulated genes. In addition, STAT3-short interfering RNA significantly reduced mRNA levels of VEGF in CLL cells suggesting that STAT3 induces VEGF expression in CLL. Remarkably, bone marrow CLL cells expressed high levels of VEGF and high VEGF levels were detected in the plasma of patients with untreated CLL and correlated with white blood cell count. CLL bone marrow biopsies revealed increased microvascular density and attachment of CLL cells to endothelial cells. Co-culture of CLL and human umbilical vein endothelial cells (HUVEC) cells showed a similar attachment. Furthermore, co-culture studies with HUVEC showed that HUVEC protected CLL cells from spontaneous apoptosis by direct cell-to-cell contact as assessed by flow cytometry using Annexin V. Our data suggest that constitutively activated STAT3 induces VEGF production by CLL cells and CLL cells derive a survival advantage from endothelial cells via cell-to cell contact. PMID:21733558

  14. Dose dependent effects of platelet derived chondroitinsulfate A on the binding of CCL5 to endothelial cells

    Directory of Open Access Journals (Sweden)

    Krämer Bernhard K

    2008-12-01

    Full Text Available Abstract Background Chemokines immobilized on endothelial cells play a central role in the induced firm adhesion and transendothelial migration of leukocytes. Activation of platelets at sites of vascular injury is considered to support leukocyte adhesion and extravasation. However, activated platelets also secrete soluble glycosaminoglycans that can interfere with immobilization of chemokines. We therefore analyzed the impact of platelet derived glycosaminoglycans on the immobilization of the chemokine CCL5 (RANTES on human microvascular endothelial cells and their influence on CCL5-CCR5 interactions. Results We confirm that undiluted serum in contrast to plasma decreases binding of CCL5 to endothelial cells. However, when lower concentrations of serum were used, CCL5-presentation on endothelial cells was markedly enhanced. This enhancement was neutralized if serum was digested with chondroinitase ABC. Using different chondroitinsulfate-subtypes we demonstrate that chondroitinsulfate A mediates the enhanced presentation of CCL5 on endothelial cells, whereas chondroitinsulfate B/C even at low concentrations block CCL5 binding. CCR5 downregulation on CCR5-transfected CHO cells or human monocytes is increased by preincubation of CCL5 with serum or chondroitinsulfate A. Conclusion We show that chondroitinsulfate A released from platelets increases the binding of chemokines to endothelial cells and supports receptor internalization in a dose dependent manner. These data help to understand the proinflammatory effects of activated platelets.

  15. Mecanotransduction and Endothelial Cells

    Institute of Scientific and Technical Information of China (English)

    S.MULLER; JF.; STOLTZ2

    2005-01-01

    1 IntroductionAtherosclerosis preferentially occurs in areas of complex blood flow where there are disturbed flow and low fluid shear stress, whereas laminar blood flow and high shear stress are atheroprotective~([1]). Reports of others and our studies suggest a steady laminar flow decreases some molecules and genes expression of vascular endothelial cells (EC) that may promote atherosclerosis, as well as it can differentially regulate production of many vasoactive factors at the level of gene expression an...

  16. Recovering endothelial function

    OpenAIRE

    Bryce Moncloa, Alfonso; Médico Cardiólogo, Académico Asociado de la Academia Nacional de Medicina, Presidente del Colegio Panamericano del Endotelio, Miembro del Comité Institucional de Investigaciones del Comité Ejecutivo de la Sociedad Latino Americana de Hipertensión Arterial y del Colegio Americano de Cardiología (ACC).; Morales Villegas, Enrique C.; Médico Internista y Cardiólogo, Fundador y Director del Centro de Investigación Cardiometabólica de Aguascalientes. México; Urquiaga Calderón, Juan; Médico Internista y Cardiólogo, Fundador y Director del Centro de Investigación Cardiometabólica de Aguascalientes. México; Larrauri Vigna, Cesar; Médico Cardiólogo, Miembro de la Sociedad Peruana de Cardiología, Sociedad Peruana de Hipertensión, Sociedad Peruana de Medicina Interna, Colegio Panamericano del Endotelio.

    2014-01-01

    Atherosclerosis starts early in life. The presence of risk factors like hypertension, smoking, dyslipidemia and diabetes as well as obesity and metabolic syndrome accelerates its progress. These factors generate endothelial dysfunction, oxidative stress and inflammation, with early appearance of foamy cells, fatty streaks and atheromatous plaques. These plaques are vulnerable to erosion and rupture, the so called atherothrombotic phenomenon, leading to acute vascular events like acute coronar...

  17. Microvascular flow estimation by microbubble-assisted Nakagami imaging.

    Science.gov (United States)

    Tsui, Po-Hsiang; Yeh, Chih-Kuang; Chang, Chien-Cheng

    2009-04-01

    The destruction and replenishment of microbubbles has been previously applied to estimating blood flow in the microcirculation. The rate of increase of the time-intensity curve (TIC) due to microbubbles flowing into the region-of-interest (ROI) as measured from the conventional B-mode images reflects the flow velocity. In this study, we monitored microbubble replenishment using a new proposed approach called the time-Nakagami-parameter curve (TNC) obtained from the parametric image based on the Nakagami statistical parameter for quantifying the microvascular flow velocity. The Nakagami parameter is estimated from signal envelope to reflect the backscattered statistics. The feasibility of using the TNC to estimate the microvascular flow was explored by carrying out phantom measurements and in vivo animal experiments. The rates of increase of the TIC and TNC were quantified as the rate constants beta(I) and beta(N) of monoexponential fitted curves, respectively. The experimental results showed that beta(N) behaves similarly to the conventional beta(I) in quantifying the flow velocity. Moreover, the tolerance to the effects of clutter is greater for the TNC than for the TIC, which makes it possible to use beta(N) to differentiate various flow velocities even when the ROI contains nonperfused areas. This finding suggests that the TNC-based technique can be used as a complementary tool for the conventional TIC to improve measurement of blood flow in the microcirculation.

  18. The role of the microvascular tortuosity in tumor transport phenomena.

    Science.gov (United States)

    Penta, R; Ambrosi, D

    2015-01-07

    The role of the microvascular network geometry in transport phenomena in solid tumors and its interplay with the leakage and pressure drop across the vessels is qualitatively and quantitatively discussed. Our starting point is a multiscale homogenization, suggested by the sharp length scale separation that exists between the characteristic vessels and the tumor tissue spatial scales, referred to as the microscale and the macroscale, respectively. The coupling between interstitial and capillary compartment is described by a double Darcy model on the macroscale, whereas the geometric information on the microvascular structure is encoded in the effective hydraulic conductivities, which are numerically computed by solving classical differential problems on the microscale representative cell. Then, microscale information is injected into the macroscopic model, which is analytically solved in a prototypical geometry and compared with previous experimentally validated, phenomenological models. In this way, we are able to capture the role of the standard blood flow determinants in the tumor, such as tumor radius, tissue hydraulic conductivity and vessels permeability, as well as influence of the vascular tortuosity on fluid convection. The results quantitatively confirm that transport of blood (and, as a consequence, of any advected anti-cancer drug) can be dramatically impaired by increasing the geometrical complexity of the microvasculature. Hence, our quantitative analysis supports the argument that geometric regularization of the capillary network improves blood transport and drug delivery in the tumor mass.

  19. Angina pectoris in women: focus on microvascular disease.

    Science.gov (United States)

    Zuchi, Cinzia; Tritto, Isabella; Ambrosio, Giuseppe

    2013-02-20

    Ischemic heart disease (IHD) is the leading cause of death among women in Western countries, and it is associated with higher morbidity and mortality than in men. Nevertheless, IHD in women remains underdiagnosed and undertreated, and the misperception that females are "protected" against cardiovascular disease leads to underestimation of their cardiovascular risk; instead, women with chest pain have a high risk of cardiovascular events. Women suffering from angina pectoris tend to have different characteristics compared to men, with a high prevalence of non-significant coronary artery disease. Angina in women is more commonly microvascular in origin than in men, and therefore standard diagnostic algorithms may be suboptimal for women. This different pathophysiology impacts clinical management of IHD in women. While response to medical therapy may differ in women, they are scarcely represented in clinical trials. Therefore, solid data in terms of gender efficacy of antianginal drugs are lacking, and particularly when angina is microvascular in origin women often continue to be symptomatic despite maximal therapy with classical antianginal drugs. Recently, new molecules have shown promising results in women. In conclusion, women with angina are a group of patients in whom it seems appropriate to concentrate efforts aimed at reducing morbidity and improving quality of life.

  20. Bosentan fosters microvascular de-remodelling in systemic sclerosis.

    Science.gov (United States)

    Guiducci, S; Bellando Randone, S; Bruni, C; Carnesecchi, G; Maresta, A; Iannone, F; Lapadula, G; Matucci Cerinic, M

    2012-12-01

    Bosentan, a dual endothelin receptor antagonist, may reduce blood pressure by blocking the vasoconstrictor effect of endothelin-1. In systemic sclerosis (SSc) nailfold videocapillaroscopy (NVC); allows diagnostic and follow-up of microvascular damage. Distinct NVC patterns have been identified for the evaluation of severity of SSc microvascular damage. The objective of this study is to evaluate the modification of the microvasculature under Bosentan therapy in SSc patients with pulmonary arterial hypertension (PAH). Nine patients with PAH related to SSc in New York Heart Association classes III-IV were treated with Bosentan 125 mg twice a day. NVC optical probe videocapillaroscopy equipped with 100× and 200× contact lenses and connected to image analyse software was performed before and after 12 months of Bosentan therapy to evaluate the modification of microvasculature. Nine PAH SSc patients treated with Iloprost were used as controls. Before Bosentan therapy, seven patients showed at NVC severe loss of capillaries with large avascular areas and vascular architectural disorganisation which are typically "late" SSc pattern. After 12 months of Bosentan, NVC pattern changed in seven patients from "late" into "active" SSc pattern. The disappearance of avascular areas and capillary haemorrhages was the most striking result. Two patients had an "active" SSc pattern, not modified by Bosentan treatment. These data show that Bosentan may improve NVC pattern in SSC and the presence of new capillaries suggests that it may favour angiogenesis. Bosentan may improve and stabilise the microvasculature in long-term treatment modulating the structural modifications detected by NVC.

  1. Monitoring microvascular free flaps with tissue oxygen measurement and PET.

    Science.gov (United States)

    Schrey, Aleksi R; Kinnunen, Ilpo A J; Grénman, Reidar A; Minn, Heikki R I; Aitasalo, Kalle M J

    2008-07-01

    Tissue oxygen measurement and positron emission tomography (PET) were evaluated as methods for predicting ischemia in microvascular free flaps of the head and neck. Ten patients with head and neck squamous cell cancer underwent resection of the tumour followed by microvascular reconstruction with a free flap. Tissue oxygenation of the flap (P(ti)O(2)) was continuously monitored for three postoperative (POP) days and the blood flow of the flap was assessed using oxygen-15 labelled water and PET. In three free flaps a perfusion problem was suspected due to a remarkable drop in P(ti)O(2)-values, due to two anastomosis problems and due to POP turgor. No flap losses occurred. During the blood flow measurements with PET [mean 8.5 mL 100 g(-1) min(-1 )(SD 2.5)], the mean P(ti)O(2) of the flaps [46.8 mmHg (SD 17.0)] appeared to correlate with each other in each patient (pmonitoring system of free flaps. The perfusion-study with PET correlates with P(ti)O(2)-measurement.

  2. EFFECT OF GENISTEIN ON EXPRESSION OF VASCULAR ENDOTHELIAL GROWTH FACTOR IN MDA-MB-453 BREAST CANCER CELLS%染料木黄酮对MDA-MB-453乳腺癌细胞VEGF表达的影响

    Institute of Scientific and Technical Information of China (English)

    余小平; 糜漫天; 朱俊东

    2005-01-01

    目的:观察染料木黄酮(genistein,Gen)对MDA-MB-453乳腺癌细胞血管内皮生长因子(vascular endothelial growth factor,VEGF)表达的影响,探讨Gen抗HER-2/neu高表达乳腺癌血管生成的分子机制.方法:5×10-5mol/L Gen处理MDA-MB-453细胞24、48、72 h后,应用免疫组织化学、Western blot及RT-PCR法检测MDA-MB-453乳腺癌细胞VEGF的表达变化.结果:Gen处理MDA-MB-453细胞24、48、72 h后,VEGF的mRNA和蛋白表达量随着处理时间的延长逐渐下降.结论:Gen能在转录和翻译水平下调HER-2/neu高表达乳腺癌细胞VEGF的表达,这可能是Gen抑制HER-2/neu高表达乳腺癌血管生成的机制之一.

  3. Carotid Artery Intima-Media Thickness and Cutaneous Microvascular Function are Associated With Vitamin C Levels in Young Patients With Type 1 Diabetes

    DEFF Research Database (Denmark)

    Odermarsky, Michal; Lykkesfeldt, Jens; Liuba, Petru

    2008-01-01

    Background: Vascular endothelial dysfunction and accelerated thickening of arterial intima contribute to increased cardiovascular morbidity in type 1 diabetes. Although vitamin C has important antioxidant functions, and increased oxidative stress is a central mechanism of vascular abnormalities......, and the measured skin perfusion was expressed as % maximal increase from baseline. Markers of systemic inflammation (C-reactive protein, fibrinogen and orosomucoid), lipids (total, LDL and HDL cholesterol) and glycosylated hemoglobin (HbA1c) were measured in EDTA plasma. Ascorbate was analyzed in meta......-phosphoric acid-stabilized EDTA-plasma by high-performance liquid chromatography. Results: In univariate regression analysis, both cIMT and skin microvascular response to ACH correlated with ascorbate levels (r=-0.29; p=0.04, and r=0.43; p=0.004 respectively). The relationships between these vascular indexes...

  4. 乳腺癌组织中 VEGF-D、MMP-2及 MMP-9的表达及其临床意义%The Expressions and Clinical Significance of Matrix Metalloproteinases 2(MMP-2),Ma-trix Metalloproteinases 9 (MMP-9) and Vascular Endothelial Growth Factor-D (VEGF-D) in Breast Cancer

    Institute of Scientific and Technical Information of China (English)

    张阿娜

    2016-01-01

    目的:探讨乳腺癌组织中基质金属蛋白酶-2(MMP-2)、基质金属蛋白酶-9(MMP-9)和血管内皮生长因子D( VEGF-D)表达的临床意义。方法采用免疫组化法,对60例乳腺癌组织及60例正常组织中 MMP-2、MMP-9及VEGF-D表达情况进行检测。结果60例乳腺癌组织中,MMP-2、MMP-9阳性表达率分别为61.67%、56.67%,VEGF-D阳性表达率为61.67%。 MMP-2、MMP-9及VEGF-D阳性表达率Ⅲ~Ⅳ期者显著高于Ⅰ~Ⅱ期者;低分化者显著高于高、中分化者;有淋巴结转移者显著高于无淋巴结转移者;且MMP-2与VEGF-D表达,MMP-9与VEGF-D表达均呈正相关性。结论低分化、Ⅲ~Ⅳ期乳腺癌患者高表达MMP-2、MMP-9与VEGF-D,MMPs与VEGF-D表达呈正相关性,可通过阻断VEGF-D及MMPs活性控制、治疗乳腺癌。%Objective To explore the expressions of matrix metalloproteinases 2 ( MMP-2 ) ,matrix metalloproteinases 9 (MMP-9) and vascular endothelial growth factor-D(VEGF-D)and their significance in breast cancer tissues.Methods The ex-pressions of MMP-2,MMP-9 and VEGF-D in 60 cases of breast cancer tissues and 60 cases of normal esophageal tissues were de-tected by immunohistochemistry.Results The expression rates of MMP-2,MMP-9 and VEGF-D in breast cancer tissues were 61.67%,56.67%and 61.67%.The expression rates of MMP-2,MMP-9 and VEGF-D in grade Ⅲ-Ⅳ were significantly higher than that of gradeⅠ-Ⅱ.The expression rates of MMP-2,MMP-9 and VEGF-D in patients with poor differentiation were signifi-cantly higher than that with well-mediate differentiation.The expression rates of MMP-2,MMP-9 and VEGF-D in patients with lymph node metastasis were significantly higher than that without lymph node metastasis.MMP-2 and VEGF-D expression were positively correlated.MMP-9 and VEGF-D expression were also positively correlated.Conclusion The expression rates of MMP-2,MMP-9 and VEGF-D are closely related to poor differentiation

  5. Twist和Snail在乳腺癌中的表达及其与血管生成的关系%Relationship of expression of Twist and Snail with angiogenesis in breast cancer

    Institute of Scientific and Technical Information of China (English)

    蔡凤林; 章佳新; 符德元; 何春兰; 邵稳喜

    2013-01-01

    Objective:To study the expression of Twist and Snail in breast cancer and their relationship with angiogenesis. Methods: Expression of Twist and Snail was detected in 80 cases of breast cancer tissue by immunohisto-chemistry. CD34 marking vascular endothelial cell was used to measure the microvascular desity( MVD). Results:The positive Twist expression was 67. 5% in breast cancer,and the positive Snail was 72. 5% . The positive group of Twist and Snail protein had higher MVD than that in negative group(P <0.05). There was a significant correlation between the expressions of Twist and Snail in breast cancer , r = 0. 529, P < 0.01. Conclusion: The overexpression of Twist and Snail protein were found in breast cancer. Their co - expression could contribute to the invasion and metastasis in human breast cancer. They were valuable molecular markers to be used for monitoring the disease.%目的:探讨Twist和Snail蛋白在乳腺癌组织中的表达及其与微血管密度的关系.方法:应用免疫组织化学法检测80例乳腺癌组织Twist、Snail蛋白的表达情况.CD34抗体标记乳腺癌血管内皮细胞,计算微血管密度(MVD).结果:乳腺癌组织中Twist的阳性表达率为67.5% (54/80),Snail的阳性表达率为72.5%(58/80),Twist、Snail阳性组MVD显著高于Twist、Snail阴性组,P<0.05.在乳腺癌组织中Twist、Snail蛋白的表达之间存在显著正相关,r =0.529,P<0.01.结论:乳腺癌组织中存在Twist、Snail蛋白的高表达,两者的联合表达能增加肿瘤细胞侵袭力,促进乳腺癌的浸润、转移,联合检测有助于判断患者的预后.

  6. Breast Gangrene

    Directory of Open Access Journals (Sweden)

    Husasin Irfan

    2011-08-01

    Full Text Available Abstract Background Breast gangrene is rare in surgical practice. Gangrene of breast can be idiopathic or secondary to some causative factor. Antibiotics and debridement are used for management. Acute inflammatory infiltrate, severe necrosis of breast tissue, necrotizing arteritis, and venous thrombosis is observed on histopathology. The aim of was to study patients who had breast gangrene. Methods A prospective study of 10 patients who had breast gangrene over a period of 6 years were analyzed Results All the patients in the study group were female. Total of 10 patients were encountered who had breast gangrene. Six patients presented with breast gangrene on the right breast whereas four had on left breast. Out of 10 patients, three had breast abscess after teeth bite followed by gangrene, one had iatrogenic trauma by needle aspiration of erythematous area of breast under septic conditions. Four had history of application of belladonna on cutaneous breast abscess and had then gangrene. All were lactating female. Amongst the rest two were elderly, one of which was a diabetic who had gangrene of breast and had no application of belladonna. All except one had debridement under cover of broad spectrum antibiotics. Three patients had grafting to cover the raw area. Conclusion Breast gangrene occurs rarely. Etiology is variable and mutifactorial. Teeth bite while lactation and the iatrogenic trauma by needle aspiration of breast abscess under unsterlised conditions could be causative. Uncontrolled diabetes can be one more causative factor for the breast gangrene. Belladonna application as a topical agent could be inciting factor. Sometimes gangrene of breast can be idiopathic. Treatment is antibiotics and debridement.

  7. Thalidomide effect in endothelial cell of acute radiation proctitis

    Institute of Scientific and Technical Information of China (English)

    Ki-Tae Kim; Hiun-Suk Chae; Jin-Soo Kim; Hyung-Keun Kim; Young-Seok Cho; Whang Choi; Kyu-Yong Choi; Sang-Young Rho; Suk-Jin Kang

    2008-01-01

    AIM: To determine whether thalidomide prevents microvascular injury in acute radiation proctitis in white rats. METHODS: Fourteen female Wistar rats were used:six in the radiation group,six in the thalidomide group,and two in normal controls.The radiation and thalidomide groups were irradiated at the pelvic area using a single 30 Gy exposure.The thalidomide (150 mg/kg) was injected into the peritoneum for 7 d from the day of irradiation.All animals were sacrificed and the rectums were removed on day 8 after irradiation.The microvessels of resected specimens were immunohistochemically stained with thrombomodulin (TM),yon Willebrand Factor (vWF),and vascular endothelial growth factor (VEGF).RESULTS: The microscopic scores did not differ significantly between the radiation and thalidomide groups,but both were higher than in the control group.Expression of TM was significantly lower in the endothelial cells (EC) of the radiation group than in the control and thalidomide groups (P < 0.001).The number of capillaries expressing vWF in the EC was higher in the radiation group (15.3 ± 6.8) than in the control group (3.7 ± 1.7),and the number of capillaries expressing vWF was attenuated by thalidomide (10.8 ± 3.5,P < 0.001).The intensity of VEGF expression in capillaries was greater in the radiation group than in the control group and was also attenuated by thalidomide (P = 0.003).CONCLUSION: The mechanisms of acute radiationinduced proctitis in the rats are related to endothelial cell injury of microvessel,which may be attenuated with thalidomide.

  8. Fate of cerium dioxide nanoparticles in endothelial cells: exocytosis

    Energy Technology Data Exchange (ETDEWEB)

    Strobel, Claudia, E-mail: Claudia.Strobel@med.uni-jena.de [Jena University Hospital – Friedrich Schiller University Jena, Department of Experimental Radiology, Institute of Diagnostic and Interventional Radiology (Germany); Oehring, Hartmut [Jena University Hospital – Friedrich Schiller University Jena, Institute of Anatomy II (Germany); Herrmann, Rudolf [University of Augsburg, Department of Physics (Germany); Förster, Martin [Jena University Hospital – Friedrich Schiller University Jena, Department of Internal Medicine I, Division of Pulmonary Medicine and Allergy/Immunology (Germany); Reller, Armin [University of Augsburg, Department of Physics (Germany); Hilger, Ingrid, E-mail: ingrid.hilger@med.uni-jena.de [Jena University Hospital – Friedrich Schiller University Jena, Department of Experimental Radiology, Institute of Diagnostic and Interventional Radiology (Germany)

    2015-05-15

    Although cytotoxicity and endocytosis of nanoparticles have been the subject of numerous studies, investigations regarding exocytosis as an important mechanism to reduce intracellular nanoparticle accumulation are rather rare and there is a distinct lack of knowledge. The current study investigated the behavior of human microvascular endothelial cells to exocytose cerium dioxide (CeO{sub 2}) nanoparticles (18.8 nm) by utilization of specific inhibitors [brefeldin A; nocodazole; methyl-β-cyclodextrin (MβcD)] and different analytical methods (flow cytometry, transmission electron microscopy, inductively coupled plasma mass spectrometry). Overall, it was found that endothelial cells were able to release CeO{sub 2} nanoparticles via exocytosis after the migration of nanoparticle containing endosomes toward the plasma membrane. The exocytosis process occurred mainly by fusion of vesicular membranes with plasma membrane resulting in the discharge of vesicular content to extracellular environment. Nevertheless, it seems to be likely that nanoparticles present in the cytosol could leave the cells in a direct manner. MβcD treatment led to the strongest inhibition of the nanoparticle exocytosis indicating a significant role of the plasma membrane cholesterol content in the exocytosis process. Brefeldin A (inhibitor of Golgi-to-cell-surface-transport) caused a higher inhibitory effect on exocytosis than nocodazole (inhibitor of microtubules). Thus, the transfer from distal Golgi compartments to the cell surface influenced the exocytosis process of the CeO{sub 2} nanoparticles more than the microtubule-associated transport. In conclusion, endothelial cells, which came in contact with nanoparticles, e.g., after intravenously applied nano-based drugs, can regulate their intracellular nanoparticle amount, which is necessary to avoid adverse nanoparticle effects on cells.

  9. Levamisole induced apoptosis in cultured vascular endothelial cells

    Science.gov (United States)

    Artwohl, Michaela; Hölzenbein, Thomas; Wagner, Ludwig; Freudenthaler, Angelika; Waldhäusl, Werner; Baumgartner-Parzer, Sabina M

    2000-01-01

    To better understand the anticancer activity of Levamisole (LMS), which serves as an adjuvant in colon cancer therapy in combination with 5-Fluorouracil, this study analyses LMS' ability to induce apoptosis and growth arrest in cultured human micro- and macrovascular endothelial cells (ECs) and fibroblasts. Cells exposed (24 h) to Levamisole (range: 0.5–2 mmol l−1) alone or in combination with antioxidants (10 mmol l−1 glutathione or 5 mmol l−1 N-Acetylcysteine or 0.1 mmol l−1 Tocopherol) were evaluated for apoptosis (3H-thymidine assays, in situ staining), mRNA/protein expression (Northern/Western blot), and proliferation (3H-thymidine incorporation). Levamisole dose-dependently increased apoptosis in ECs to 230% (HUVECs-human umbilical vein ECs), 525% (adult human venous ECs) and 600% (human uterine microvascular ECs) but not in fibroblasts compared to control cells (set as 100%). Levamisole increased in ECs integrin-dependent matrix adhesion, inhibited proliferation (−70%), reduced expression of survival factors such as clusterin (−30%), endothelin-1 (−43%), bcl-2 (−34%), endothelial NO-synthase (−32%) and pRb (Retinoblastoma protein: −89%), and increased that of growth arrest/death signals such as p21 (+73%) and bak (+50%). LMS (2 mmol l−1)-induced apoptosis was inhibited by glutathione (−50%) and N-Acetylcysteine (−36%), which also counteracted reduction by Levamisole of pRb expression, suggesting reactive oxygen species and pRb play a role in these processes. The ability of LMS to selectively induce apoptosis and growth arrest in endothelial cells potentially hints at vascular targeting to contribute to Levamisole's anticancer activity. PMID:11139434

  10. Endothelial RIG-I activation impairs endothelial function

    Energy Technology Data Exchange (ETDEWEB)

    Asdonk, Tobias, E-mail: tobias.asdonk@ukb.uni-bonn.de [Department of Medicine/Cardiology, University of Bonn, Sigmund-Freud-Str. 25, 53105 Bonn (Germany); Motz, Inga; Werner, Nikos [Department of Medicine/Cardiology, University of Bonn, Sigmund-Freud-Str. 25, 53105 Bonn (Germany); Coch, Christoph; Barchet, Winfried; Hartmann, Gunther [Institute for Clinical Chemistry and Clinical Pharmacology, University of Bonn, Sigmund-Freud-Str. 25, 53105 Bonn (Germany); Nickenig, Georg; Zimmer, Sebastian [Department of Medicine/Cardiology, University of Bonn, Sigmund-Freud-Str. 25, 53105 Bonn (Germany)

    2012-03-30

    Highlights: Black-Right-Pointing-Pointer RIG-I activation impairs endothelial function in vivo. Black-Right-Pointing-Pointer RIG-I activation alters HCAEC biology in vitro. Black-Right-Pointing-Pointer EPC function is affected by RIG-I stimulation in vitro. -- Abstract: Background: Endothelial dysfunction is a crucial part of the chronic inflammatory atherosclerotic process and is mediated by innate and acquired immune mechanisms. Recent studies suggest that pattern recognition receptors (PRR) specialized in immunorecognition of nucleic acids may play an important role in endothelial biology in a proatherogenic manner. Here, we analyzed the impact of endothelial retinoic acid inducible gene I (RIG-I) activation upon vascular endothelial biology. Methods and results: Wild type mice were injected intravenously with 32.5 {mu}g of the RIG-ligand 3pRNA (RNA with triphosphate at the 5 Prime end) or polyA control every other day for 7 days. In 3pRNA-treated mice, endothelium-depended vasodilation was significantly impaired, vascular oxidative stress significantly increased and circulating endothelial microparticle (EMP) numbers significantly elevated compared to controls. To gain further insight in RIG-I dependent endothelial biology, cultured human coronary endothelial cells (HCAEC) and endothelial progenitor cells (EPC) were stimulated in vitro with 3pRNA. Both cells types express RIG-I and react with receptor upregulation upon stimulation. Reactive oxygen species (ROS) formation is enhanced in both cell types, whereas apoptosis and proliferation is not significantly affected in HCAEC. Importantly, HCAEC release significant amounts of proinflammatory cytokines in response to RIG-I stimulation. Conclusion: This study shows that activation of the cytoplasmatic nucleic acid receptor RIG-I leads to endothelial dysfunction. RIG-I induced endothelial damage could therefore be an important pathway in atherogenesis.

  11. Angiogenesis and collagen type IV expression in different endothelial cell culture systems.

    Science.gov (United States)

    Bahramsoltani, M; Slosarek, I; De Spiegelaere, W; Plendl, J

    2014-04-01

    In vitro angiogenesis assays constitute an important tool for studying the mechanisms of angiogenesis and for identification of pro- and anti-angiogenic substances. Therefore, endothelial cell and media systems used for in vitro angiogenesis assays are required to mimic the angiogenic process in vivo including endothelial capability to express collagen type IV as a component of the basement membrane. In this study, the expression of collagen type IV and its α chains (α1-6) was investigated in different endothelial cell culture systems in vitro qualitatively and quantitatively. These systems included four different batches of microvascular endothelial cells derived from the human skin, heart and lung, from which only two batches were found to be angiogenic and two batches were classified as non-angiogenic. Distribution of the transcripts of the α chains of collagen type IV was similar in all cell and media systems investigated. However, secretion and deposition of a stable extracellular network of collagen type IV could only be observed in the angiogenic cultures. In conclusion, the consecutive steps of the angiogenic cascade in vivo as well as in vitro depend on an increasing secretion and subsequent extracellular deposition of collagen type IV.

  12. IL-6 acts on endothelial cells to preferentially increase their adherence for lymphocytes.

    Science.gov (United States)

    Watson, C; Whittaker, S; Smith, N; Vora, A J; Dumonde, D C; Brown, K A

    1996-07-01

    Using a quantitative monolayer adhesion assay, the current report shows that treatment of human umbilical vein endothelial cells (HUVEC) with IL-6 increases their adhesiveness for blood lymphocytes, particularly CD4+ cells, but not for polymorphonuclear cells and monocytes. This effect, which was most pronounced when using low concentrations of the cytokine (0.1-1.0 U/ml) and a short incubation period (4h), was also apparent with microvascular endothelial cells and a hybrid endothelial cell line. Skin lesions from patients with mycosis fungoides contain high levels of IL-6, and blood lymphocytes from patients with this disorder also exhibited an enhanced adhesion to IL-6-treated HUVEC. The cytokine enhanced intercellular adhesion molecule-1 (ICAM-1) expression and induced the expression of vascular cell adhesion molecule-1 (VCAM-1) and E-selectin on endothelial cells. Antibody blocking studies demonstrated that the vascular adhesion molecules ICAM-1, VCAM-1 and E-selectin and the leucocyte integrin LFA-1 all contributed to lymphocyte binding to endothelium activated by IL-6. It is proposed that IL-6 may be involved in the recruitment of lymphocytes into non-lymphoid tissue.

  13. Analysis of proteome response to the mobile phone radiation in two types of human primary endothelial cells

    Directory of Open Access Journals (Sweden)

    Kuster Niels

    2010-10-01

    Full Text Available Abstract Background Use of mobile phones has widely increased over the past decade. However, in spite of the extensive research, the question of potential health effects of the mobile phone radiation remains unanswered. We have earlier proposed, and applied, proteomics as a tool to study biological effects of the mobile phone radiation, using as a model human endothelial cell line EA.hy926. Exposure of EA.hy926 cells to 900 MHz GSM radiation has caused statistically significant changes in expression of numerous proteins. However, exposure of EA.hy926 cells to 1800 MHz GSM signal had only very small effect on cell proteome, as compared with 900 MHz GSM exposure. In the present study, using as model human primary endothelial cells, we have examined whether exposure to 1800 MHz GSM mobile phone radiation can affect cell proteome. Results Primary human umbilical vein endothelial cells and primary human brain microvascular endothelial cells were exposed for 1 hour to 1800 MHz GSM mobile phone radiation at an average specific absorption rate of 2.0 W/kg. The cells were harvested immediately after the exposure and the protein expression patterns of the sham-exposed and radiation-exposed cells were examined using two dimensional difference gel electrophoresis-based proteomics (2DE-DIGE. There were observed numerous differences between the proteomes of human umbilical vein endothelial cells and human brain microvascular endothelial cells (both sham-exposed. These differences are most likely representing physiological differences between endothelia in different vascular beds. However, the exposure of both types of primary endothelial cells to mobile phone radiation did not cause any statistically significant changes in protein expression. Conclusions Exposure of primary human endothelial cells to the mobile phone radiation, 1800 MHz GSM signal for 1 hour at an average specific absorption rate of 2.0 W/kg, does not affect protein expression, when the

  14. Breast lift

    Science.gov (United States)

    ... Planning to have more children Talk with a plastic surgeon if you are considering cosmetic breast surgery. Discuss ... before surgery: You may need a mammogram . Your plastic surgeon will do a routine breast exam. You may ...

  15. Breast; Sein

    Energy Technology Data Exchange (ETDEWEB)

    Bourgier, C.; Garbay, J.R.; Pichenot, C.; Uzan, C.; Delaloge, S.; Andre, F.; Spielmann, M.; Arriagada, R.; Lefkopoulos, D.; Marsigli, H.; Bondiau, P.Y.; Courdi, A.; Lallemand, M.; Peyrotte, I.; Chapellier, C.; Ferrero, J.M.; Chiovati, P.; Baldissera, A.; Frezza, G.; Vicenzi, L.; Palombarini, M.; Martelli, O.; Degli Esposti, C.; Donini, E.; Romagna CDR, E.; Romagna CDF, E.; Benmensour, M.; Bouchbika, Z.; Benchakroun, N.; Jouhadi, H.; Tawfiq, N.; Sahraoui, S.; Benider, A.; Gilliot, O.; Achard, J.L.; Auvray, H.; Toledano, I.; Bourry, N.; Kwiatkowski, F.; Verrelle, P.; Lapeyre, M.; Tebra Mrad, S.; Braham, I.; Chaouache, K.; Bouaouin, N.; Ghorbel, L.; Siala, W.; Sallemi, T.; Guermazi, M.; Frikha, M.; Daou, J.; El Omrani, A.; Chekrine, T.; Mangoni, M.; Castaing, M.; Folino, E.; Livi, L.; Dunant, A.; Mathieu, M.C.; Bitib, G.P.; Arriagada, R.; Marsigli, H

    2007-11-15

    Nine articles treat the question of breast cancer. Three-dimensional conformal accelerated partial breast irradiation: dosimetric feasibility study; test of dose escalation neo-adjuvant radiotherapy focused by Cyberknife in breast cancer; Three dimensional conformal partial irradiation with the technique by the Irma protocol ( dummy run multi centers of the Emilie Romagne area Italy); Contribution of the neo-adjuvant chemotherapy in the treatment of locally evolved cancers of the uterine cervix; Post operative radiotherapy of breast cancers (N0, pN) after neo-adjuvant chemotherapy. Radiotherapy of one or two mammary glands and ganglions areas,The breast cancer at man; breast conservative treatment; breast cancers without histological ganglions invasion; the breast cancer at 70 years old and more women; borderline mammary phyllod tumors and malignant. (N.C.)

  16. Directed assembly of three-dimensional microvascular networks

    Science.gov (United States)

    Therriault, Daniel

    Three-dimensional (3-D) microvascular networks with pervasive, interconnected channels less than 300 mum in diameter may find widespread application in microfluidic devices, biotechnology, sensors, and autonomic healing materials. Although microchannel arrays are readily constructed in two-dimensions by photolithographic or soft lithographic techniques, their construction in three-dimensions remains a challenging problem. The development of a microfabrication method to build 3-D microvascular networks based on direct-write assembly is described is this thesis. The method is based on the robotic deposition of a fugitive organic ink to form a free-standing scaffold structure. Secondary infiltration of a structural resin followed by setting of the matrix and removal of the scaffold yields an embedded pervasive network of smooth cylindrical channels (˜10--500 mum) with defined connectivity. Rheological and other material properties studies of fugitive organic ink were performed in order to identify the critical characteristics required for successful deposition of 3-D scaffolds by direct-write assembly. Guided by the results of these studies, several new ink formulations were screened for improved deposition performance. The most successful of these inks (40wt% microcrystalline wax, 60wt% petroleum jelly) showed excellent deposition and had an equilibrium modulus at room temperature (G 'eq ˜ 7.70 kPa 1 Hz) nearly two orders of magnitude higher than the original ink. The optimized ink was used to successfully build thick (i.e., ˜100 layers) scaffold structures at room temperature with negligible time-dependent deformation post-deposition. Secondary infiltration of the resin was accomplished at room temperature while maintaining the scaffold architecture. The optimized ink was also successfully extruded through small micronozzles (1 mum). The construction of 3-D microvascular networks enables microfluidic devices with unparallel geometric complexity. In one example, a

  17. Effects of Exenatide vs. Metformin on endothelial function in obese patients with pre-diabetes: a randomized trial

    Directory of Open Access Journals (Sweden)

    Kelly Aaron S

    2012-06-01

    Full Text Available Abstract Background Glucagon like peptide-1 (GLP-1 receptor agonist treatment may improve endothelial function via direct and indirect mechanisms. We compared the acute and chronic effects of the GLP-1 receptor agonist exenatide vs. metformin on endothelial function in patients with obesity and pre-diabetes. Methods We performed a randomized, open-label, clinical trial in 50 non-diabetic individuals (mean age 58.5 ± 10.0; 38 females with abdominal obesity and either impaired fasting glucose, elevated HbA1c, or impaired glucose tolerance (IGT who were randomized to receive 3-months of exenatide or metformin. Microvascular endothelial function, assessed by digital reactive hyperemia (reactive hyperemic index: RHI, C-reactive protein (CRP, circulating oxidized LDL (oxLDL, and vascular cell adhesion molecule-1 (VCAM-1 were measured at baseline and 3-months. Seven subjects with IGT participated in a sub-study comparing the effects of pre-administration of exenatide and metformin on postprandial endothelial function. Results There were no differences for the change in RHI (Δ exenatide: 0.01 ± 0.68 vs. Δ metformin: -0.17 ± 0.72, P = 0.348, CRP, oxLDL, or VCAM-1 between exenatide and metformin treatment. Triglycerides were reduced more with exenatide compared to metformin (Δ exenatide: -25.5 ± 45.7 mg/dL vs. Δ metformin: -2.9 ± 22.8 mg/dL, P = 0.032. In the sub-study, there was no difference in postprandial RHI between exenatide and metformin. Conclusions Three months of exenatide therapy had similar effects on microvascular endothelial function, markers of inflammation, oxidative stress, and vascular activation, as metformin, in patients with obesity and pre-diabetes. Clinical trials registration This study is registered on http://www.clinicaltrials.gov/: NCT00546728

  18. Relationship Between Serum Zinc Level and Microvascular Complications in Patients with Type 2 Diabetes

    Institute of Scientific and Technical Information of China (English)

    Ying-Ying Luo; Jie Zhao; Xue-Yao Han; Xiang-Hai Zhou; Jing Wu; Li-Nong Ji

    2015-01-01

    Background: Previous studies suggested that zinc level was related to a certain diabetic microvascular complication.However, the relationship between zinc level and all the microvascular complications in type 2 diabetic patients remains unknown.The purpose of this study was to analyze the relationship between zinc level and each diabetic microvascular complication and identify the features related to low serum zinc level.Methods: We included the hospitalized patients with type 2 diabetes (T2D) at our department from May 30, 2013 to March 31, 2014.We initially compared the serum zinc levels between patients with specific microvascular complications and those without.We then analyzed the association between zinc level and each microvascular complication.Furthermore, we identified the unique features of patients with high and low serum zinc levels and analyzed the risk factors related to low zinc level.Results: The 412 patients included 271 with microvascular complications and 141 without any microvascular complications.Serum zinc level was significantly lower in patients with diabetic retinopathy (P < 0.001), diabetic nephropathy (DN, P < 0.001), or diabetic peripheral neuropathy (P =0.002) compared with patients without that specific complication.Lower zinc level was an independent risk factor for DN (odds ratio =0.869, 95% confidence interval =0.765-0.987, P < 0.05).The subjects with lower serum zinc level had manifested a longer duration of diabetes, higher level of hemoglobin A 1 c, higher prevalence of hypertension and microvascular complications, and lower fasting and 2-h C-peptide levels.Conclusions: Lower serum zinc level in T2D patients was related to higher prevalence of diabetic microvascular complications, and represented as an independent risk factor for DN.Patients with lower zinc level were more likely to have a longer duration of diabetes, poorer glucose control, and worse β-cell function.

  19. Breast Cancer

    Science.gov (United States)

    Breast cancer affects one in eight women during their lives. No one knows why some women get breast cancer, but there are many risk factors. Risks that ... who have family members with breast or ovarian cancer may wish to be tested for the genes. ...

  20. Selenate enhances STAT3 transcriptional activity in endothelial cells: differential actions of selenate and selenite on LIF cytokine signaling and cell viability.

    Science.gov (United States)

    Alturkmani, Hani J; Zgheib, Carlos; Zouein, Fouad A; Alshaaer, Nour Eddin F; Kurdi, Mazen; Booz, George W

    2012-04-01

    Sodium selenate may have utility in treating Alzheimer's disease and diabetes; however, its impact on the associated proinflammatory cytokine signaling of endothelial cells has not been investigated. We report that treatment of human microvascular endothelial cells with sodium selenate at a pharmacological dose (100 μM) enhanced tyrosine phosphorylation of nuclear STAT3 on Y705 in response to IL-6-type cytokine, leukemia inhibitory factor (LIF), indicative of enhanced STAT3 activity. Accordingly, STAT3 nuclear binding to DNA was increased, as well as LIF-induced gene expression of chemokine (C-C motif) ligand 2 (CCL2). CCL2 plays a key role in inflammatory processes associated with neuronal degenerative and vascular diseases. The enhancing action of selenate on LIF-induced STAT3 Y705 phosphorylation was replicated by vanadate and a specific inhibitor of protein tyrosine phosphatase, non-receptor type 1 (PTP1B). Moreover, we observed that selenite, the cellular reduction bioproduct of selenate but not selenate itself, inhibited enzymatic activity of human recombinant PTP1B. Our findings support the conclusion that in human microvascular endothelial cells selenate has a vanadate-like effect in inhibiting PTP1B and enhancing proinflammatory STAT3 activation. These findings raise the possibility that beneficial actions of supranutritional levels of selenate for treating Alzheimer's and diabetes may be offset by a proinflammatory action on endothelial cells.

  1. Aligned-Braided Nanofibrillar Scaffold with Endothelial Cells Enhances Arteriogenesis.

    Science.gov (United States)

    Nakayama, Karina H; Hong, Guosong; Lee, Jerry C; Patel, Jay; Edwards, Bryan; Zaitseva, Tatiana S; Paukshto, Michael V; Dai, Hongjie; Cooke, John P; Woo, Y Joseph; Huang, Ngan F

    2015-07-28

    The objective of this study was to enhance the angiogenic capacity of endothelial cells (ECs) usin