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Sample records for endothelial marker podoplanin

  1. Podoplanin is a novel myoepithelial cell marker in pleomorphic adenoma and other salivary gland tumors with myoepithelial differentiation.

    PubMed

    Tsuneki, Masayuki; Maruyama, Satoshi; Yamazaki, Manabu; Essa, Ahmed; Abé, Tatsuya; Babkair, Hamzah Ali; Ahsan, Md Shahidul; Cheng, Jun; Saku, Takashi

    2013-03-01

    The expression of podoplanin, one of the representative immunohistochemical markers for lymphatic endothelium, is upregulated in various kinds of cancers. Based on our previous studies, we have developed a hypothesis that podoplanin plays a role in cell adhesion via its association with extracellular matrix (ECM). Since salivary pleomorphic adenoma is histologically characterized by its ECM-enriched stroma, we firstly wanted to explore the expression modes of podoplanin in pleomorphic adenoma and related salivary tumors by immunohistochemistry. In normal salivary gland, podoplanin was specifically localized in myoepithelial cells, which were also positively labeled by antibodies against P63, of the intercalated duct as well as acini. In pleomorphic adenoma, podoplanin was colocalized with P63 and CD44 in basal cells of glandular structures as well as in stellate/spindle cells in myxochondroid matrices, where perlecan and hyaluronic acid were enriched. The expression of podoplanin was confirmed at both protein and mRNA levels in pleomorphic adenoma cell systems (SM-AP1 and SM-AP4) by using immunofluorescence, western blotting, and reverse transcription polymerase chain reaction. Podoplanin was localized on the cell border as well as in the external periphery of the cells. Moreover, podoplanin expression was also confirmed in tumor cells with myoepithelial differentiation in myoepithelioma and intraductal papilloma. The results indicate that podoplanin can be regarded as a novel myoepithelial marker in salivary gland tumors and suggest that podoplanin's communication with ECM molecules is essential to phenotypic differentiation to myoepithelial cells.

  2. The Spectrum of Podoplanin Expression in Encapsulating Peritoneal Sclerosis

    PubMed Central

    Braun, Niko; Alscher, M. Dominik; Fritz, Peter; Latus, Joerg; Edenhofer, Ilka; Reimold, Fabian; Alper, Seth L.; Kimmel, Martin; Biegger, Dagmar; Lindenmeyer, Maja; Cohen, Clemens D.; Wüthrich, Rudolf P.; Segerer, Stephan

    2012-01-01

    Encapsulating peritoneal sclerosis (EPS) is a life threatening complication of peritoneal dialysis (PD). Podoplanin is a glycoprotein expressed by mesothelial cells, lymphatic endothelial cells, and myofibroblasts in peritoneal biopsies from patients with EPS. To evaluate podoplanin as a marker of EPS we measured podoplanin mRNA and described the morphological patterns of podoplanin-positive cells in EPS. Included were 20 peritoneal biopsies from patients with the diagnosis of EPS (n = 5), patients on PD without signs of EPS (n = 5), and control patients (uremic patients not on PD, n = 5, non-uremic patients n = 5). EPS patient biopsies revealed significantly elevated levels of podoplanin mRNA (p<0.05). In 24 peritoneal biopsies from patients with EPS, podoplanin and smooth muscle actin (SMA) were localized by immunohistochemistry. Four patterns of podoplanin distribution were distinguishable. The most common pattern (8 of 24) consisted of organized, longitudinal layers of podoplanin-positive cells and vessels in the fibrotic zone (“organized” pattern). 7 of 24 biopsies demonstrated a diffuse distribution of podoplanin-positive cells, accompanied by occasional, dense clusters of podoplanin-positive cells. Five biopsies exhibited a mixed pattern, with some diffuse areas and some organized areas ("mixed"). These contained cuboidal podoplanin-positive cells within SMA-negative epithelial structures embedded in extracellular matrix. Less frequently observed was the complete absence of, or only focal accumulations of podoplanin-positive fibroblasts outside of lymphatic vessels (podoplanin “low”, 4 of 24 biopsies). Patients in this group exhibited a lower index of systemic inflammation and a longer symptomatic period than in EPS patients with biopsies of the "mixed" type (p<0.05). In summary we confirm the increased expression of podoplanin in EPS, and distinguish EPS biopsies according to different podoplanin expression patterns which are

  3. Syk and Src Family Kinases Regulate C-type Lectin Receptor 2 (CLEC-2)-mediated Clustering of Podoplanin and Platelet Adhesion to Lymphatic Endothelial Cells*

    PubMed Central

    Pollitt, Alice Y.; Poulter, Natalie S.; Gitz, Eelo; Navarro-Nuñez, Leyre; Wang, Ying-Jie; Hughes, Craig E.; Thomas, Steven G.; Nieswandt, Bernhard; Douglas, Michael R.; Owen, Dylan M.; Jackson, David G.; Dustin, Michael L.; Watson, Steve P.

    2014-01-01

    The interaction of C-type lectin receptor 2 (CLEC-2) on platelets with Podoplanin on lymphatic endothelial cells initiates platelet signaling events that are necessary for prevention of blood-lymph mixing during development. In the present study, we show that CLEC-2 signaling via Src family and Syk tyrosine kinases promotes platelet adhesion to primary mouse lymphatic endothelial cells at low shear. Using supported lipid bilayers containing mobile Podoplanin, we further show that activation of Src and Syk in platelets promotes clustering of CLEC-2 and Podoplanin. Clusters of CLEC-2-bound Podoplanin migrate rapidly to the center of the platelet to form a single structure. Fluorescence lifetime imaging demonstrates that molecules within these clusters are within 10 nm of one another and that the clusters are disrupted by inhibition of Src and Syk family kinases. CLEC-2 clusters are also seen in platelets adhered to immobilized Podoplanin using direct stochastic optical reconstruction microscopy. These findings provide mechanistic insight by which CLEC-2 signaling promotes adhesion to Podoplanin and regulation of Podoplanin signaling, thereby contributing to lymphatic vasculature development. PMID:25368330

  4. Platelets Regulate the Migration of Keratinocytes via Podoplanin/CLEC-2 Signaling during Cutaneous Wound Healing in Mice.

    PubMed

    Asai, Jun; Hirakawa, Satoshi; Sakabe, Jun-ichi; Kishida, Tsunao; Wada, Makoto; Nakamura, Naomi; Takenaka, Hideya; Mazda, Osam; Urano, Tetsumei; Suzuki-Inoue, Katsue; Tokura, Yoshiki; Katoh, Norito

    2016-01-01

    Podoplanin is an endogenous ligand for C-type lectin-like receptor 2 (CLEC-2), which is expressed on platelets. Recent evidence indicates that this specific marker of lymphatic endothelial cells is also expressed by keratinocytes at the edge of wounds. However, whether podoplanin or platelets play a role in keratinocyte activity during wound healing remains unknown. We evaluated the effect of podoplanin expression levels on keratinocyte motility using cultured primary normal human epidermal keratinocytes (NHEKs). Down-regulation of podoplanin in NHEKs via transfection with podoplanin siRNA inhibited their migration, indicating that podoplanin plays a mandatory role in this process. In addition, down-regulation of podoplanin was correlated with up-regulation of E-cadherin, suggesting that podoplanin-mediated stimulation of keratinocyte migration is associated with a loss of E-cadherin. Both the addition of platelets and treatment with CLEC-2 inhibited the migration of NHEKs. The down-regulation of RhoA activity and the up-regulation of E-cadherin in keratinocytes were also induced by CLEC-2. In conclusion, these results suggest that podoplanin/CLEC-2 signaling regulates keratinocyte migration via modulating E-cadherin expression through RhoA signaling. Altering the regulation of keratinocyte migration by podoplanin might be a novel therapeutic approach to improve wound healing.

  5. Human podoplanin-positive monocytes and platelets enhance lymphangiogenesis through the activation of the podoplanin/CLEC-2 axis.

    PubMed

    Hur, Jin; Jang, Jae Hee; Oh, Il-Young; Choi, Jae-Il; Yun, Ji-Yeon; Kim, Joonoh; Choi, Young-Eun; Ko, Seung-Bum; Kang, Jin-A; Kang, Jeehoon; Lee, Sang Eun; Lee, Hwan; Park, Young-Bae; Kim, Hyo-Soo

    2014-08-01

    Emerging studies suggested that murine podoplanin-positive monocytes (PPMs) are involved in lymphangiogenesis. The goal of this study was to demonstrate the therapeutic lymphangiogenesis of human PPMs by the interaction with platelets. Aggregation culture of human peripheral blood mononuclear cells (PBMCs) resulted in cellular aggregates termed hematospheres. During 5-day culture, PPMs expanded exponentially and expressed several lymphatic endothelial cell-specific markers including vascular endothelial growth factor receptor (VEGFR)-3 and well-established lymphangiogenic transcription factors. Next, we investigated the potential interaction of PPMs with platelets that had C-type lectin-like receptor-2 (CLEC-2), a receptor of podoplanin. In vitro coculture of PPMs and platelets stimulated PPMs to strongly express lymphatic endothelial markers and upregulate lymphangiogenic cytokines. Recombinant human CLEC-2 also stimulated PPMs through Akt and Erk signaling. Likewise, platelets in coculture with PPMs augmented secretion of a lymphangiogenic cytokine, interleukin (IL)-1-β, via podoplanin/CLEC-2 axis. The supernatant obtained from coculture was able to enhance the migration, viability, and proliferation of lymphatic endothelial cell. Local injection of hematospheres with platelets significantly increased lymphatic neovascularization and facilitated wound healing in the full-thickness skin wounds of nude mice. Cotreatment with PPMs and platelets augments lymphangiogenesis through podoplanin/CLEC-2 axis, which thus would be a promising novel strategy of cell therapy to treat human lymphatic vessel disease.

  6. D2-40/podoplanin expression in the human placenta.

    PubMed

    Wang, Y; Sun, J; Gu, Y; Zhao, S; Groome, L J; Alexander, J S

    2011-01-01

    Placental tissue expresses many lymphatic markers. The current study was undertaken to examine if D2-40/podoplanin, a lymphatic endothelial marker, was expressed in the human placenta, and how it is altered developmentally and pathologically. We examined D2-40/podoplanin and VEGFR-3 expressions in placentas from normotensive pregnancies at different gestational ages and in placentas from women with clinically defined preeclampsia. D2-40 expression in systemic lymphatic vessel endothelium served as a positive control. Protein expression for D2-40, VEGFR-3, and β-actin was determined by Western blot in placentas from normotensive (n = 6) and preeclamptic (n = 5) pregnancies. Our results show that D2-40/podoplanin was strongly expressed in the placenta, mainly as a network plexus pattern in the villous stroma throughout gestation. CD31 was limited to villous core fetal vessel endothelium and VEGFR-3 was found in both villous core fetal vessel endothelium and trophoblasts. D2-40/podoplanin expression was significantly decreased, and VEGFR-3 significantly increased in preeclamptic placental tissues compared to normotensive placental controls. Placental villous stroma is a reticular-like structure, and the localization of D2-40 to the stroma suggests that a lymphatic-like conductive network may exist in the human placenta. D2-40/podoplanin is an O-linked sialoglycoprotein. Although little is known regarding biological functions of sialylated glycoproteins within the placenta, placental D2-40/podoplanin may support fetal vessel angiogenesis during placenta development and reduced D2-40/podoplanin expression in preeclamptic placenta may contribute to altered interstitial fluid homeostasis and impaired angiogenesis in this pregnancy disorder.

  7. Phenotypically heterogeneous podoplanin-expressing cell populations are associated with the lymphatic vessel growth and fibrogenic responses in the acutely and chronically infarcted myocardium

    PubMed Central

    Cimini, Maria; Cannatá, Antonio; Pasquinelli, Gianandrea; Rota, Marcello

    2017-01-01

    Cardiac lymphatic vasculature undergoes substantial expansion in response to myocardial infarction (MI). However, there is limited information on the cellular mechanisms mediating post-MI lymphangiogenesis and accompanying fibrosis in the infarcted adult heart. Using a mouse model of permanent coronary artery ligation, we examined spatiotemporal changes in the expression of lymphendothelial and mesenchymal markers in the acutely and chronically infarcted myocardium. We found that at the time of wound granulation, a three-fold increase in the frequency of podoplanin-labeled cells occurred in the infarcted hearts compared to non-operated and sham-operated counterparts. Podoplanin immunoreactivity detected LYVE-1-positive lymphatic vessels, as well as masses of LYVE-1-negative cells dispersed between myocytes, predominantly in the vicinity of the infarcted region. Podoplanin-carrying populations displayed a mesenchymal progenitor marker PDGFRα, and intermittently expressed Prox-1, a master regulator of the lymphatic endothelial fate. At the stages of scar formation and maturation, concomitantly with the enlargement of lymphatic network in the injured myocardium, the podoplanin-rich LYVE-1-negative multicellular assemblies were apparent in the fibrotic area, aligned with extracellular matrix deposits, or located in immediate proximity to activated blood vessels with high VEGFR-2 content. Of note, these podoplanin-containing cells acquired the expression of PDGFRβ or a hematoendothelial epitope CD34. Although Prox-1 labeling was abundant in the area affected by MI, the podoplanin-presenting cells were not consistently Prox-1-positive. The concordance of podoplanin with VEGFR-3 similarly varied. Thus, our data reveal previously unknown phenotypic and structural heterogeneity within the podoplanin-positive cell compartment in the infarcted heart, and suggest an alternate ability of podoplanin-presenting cardiac cells to generate lymphatic endothelium and pro

  8. Morphological study of tooth development in podoplanin-deficient mice.

    PubMed

    Takara, Kenyo; Maruo, Naoki; Oka, Kyoko; Kaji, Chiaki; Hatakeyama, Yuji; Sawa, Naruhiko; Kato, Yukinari; Yamashita, Junro; Kojima, Hiroshi; Sawa, Yoshihiko

    2017-01-01

    Podoplanin is a mucin-type highly O-glycosylated glycoprotein identified in several somatyic cells: podocytes, alveolar epithelial cells, lymphatic endothelial cells, lymph node stromal fibroblastic reticular cells, osteocytes, odontoblasts, mesothelial cells, glia cells, and others. It has been reported that podoplanin-RhoA interaction induces cytoskeleton relaxation and cell process stretching in fibroblastic cells and osteocytes, and that podoplanin plays a critical role in type I alveolar cell differentiation. It appears that podoplanin plays a number of different roles in contributing to cell functioning and growth by signaling. However, little is known about the functions of podoplanin in the somatic cells of the adult organism because an absence of podoplanin is lethal at birth by the respiratory failure. In this report, we investigated the tooth germ development in podoplanin-knockout mice, and the dentin formation in podoplanin-conditional knockout mice having neural crest-derived cells with deficiency in podoplanin by the Wnt1 promoter and enhancer-driven Cre recombinase: Wnt1-Cre;PdpnΔ/Δmice. In the Wnt1-Cre;PdpnΔ/Δmice, the tooth and alveolar bone showed no morphological abnormalities and grow normally, indicating that podoplanin is not critical in the development of the tooth and bone.

  9. Morphological study of tooth development in podoplanin-deficient mice

    PubMed Central

    Takara, Kenyo; Maruo, Naoki; Oka, Kyoko; Kaji, Chiaki; Hatakeyama, Yuji; Sawa, Naruhiko; Kato, Yukinari; Yamashita, Junro; Kojima, Hiroshi; Sawa, Yoshihiko

    2017-01-01

    Podoplanin is a mucin-type highly O-glycosylated glycoprotein identified in several somatyic cells: podocytes, alveolar epithelial cells, lymphatic endothelial cells, lymph node stromal fibroblastic reticular cells, osteocytes, odontoblasts, mesothelial cells, glia cells, and others. It has been reported that podoplanin-RhoA interaction induces cytoskeleton relaxation and cell process stretching in fibroblastic cells and osteocytes, and that podoplanin plays a critical role in type I alveolar cell differentiation. It appears that podoplanin plays a number of different roles in contributing to cell functioning and growth by signaling. However, little is known about the functions of podoplanin in the somatic cells of the adult organism because an absence of podoplanin is lethal at birth by the respiratory failure. In this report, we investigated the tooth germ development in podoplanin-knockout mice, and the dentin formation in podoplanin-conditional knockout mice having neural crest-derived cells with deficiency in podoplanin by the Wnt1 promoter and enhancer-driven Cre recombinase: Wnt1-Cre;PdpnΔ/Δmice. In the Wnt1-Cre;PdpnΔ/Δmice, the tooth and alveolar bone showed no morphological abnormalities and grow normally, indicating that podoplanin is not critical in the development of the tooth and bone. PMID:28222099

  10. Podoplanin Expression in Canine Melanoma.

    PubMed

    Ogasawara, Satoshi; Honma, Ryusuke; Kaneko, Mika K; Fujii, Yuki; Kagawa, Yumiko; Konnai, Satoru; Kato, Yukinari

    2016-12-01

    A type I transmembrane protein, podoplanin (PDPN), is expressed in several normal cells such as lymphatic endothelial cells or pulmonary type I alveolar cells. We recently demonstrated that anticanine PDPN monoclonal antibody (mAb), PMab-38, recognizes canine PDPN of squamous cell carcinomas, but does not react with lymphatic endothelial cells. Herein, we investigated whether PMab-38 reacts with canine melanoma. PMab-38 reacted with 90% of melanoma cells (9/10 cases) using immunohistochemistry. Of interest, PMab-38 stained the lymphatic endothelial cells and cancer-associated fibroblasts in melanoma tissues, although it did not stain any lymphatic endothelial cells in normal tissues. PMab-38 could be useful for uncovering the function of PDPN in canine melanomas.

  11. Podoplanin Expression in Canine Melanoma

    PubMed Central

    Ogasawara, Satoshi; Honma, Ryusuke; Kaneko, Mika K.; Fujii, Yuki; Kagawa, Yumiko; Konnai, Satoru

    2016-01-01

    A type I transmembrane protein, podoplanin (PDPN), is expressed in several normal cells such as lymphatic endothelial cells or pulmonary type I alveolar cells. We recently demonstrated that anticanine PDPN monoclonal antibody (mAb), PMab-38, recognizes canine PDPN of squamous cell carcinomas, but does not react with lymphatic endothelial cells. Herein, we investigated whether PMab-38 reacts with canine melanoma. PMab-38 reacted with 90% of melanoma cells (9/10 cases) using immunohistochemistry. Of interest, PMab-38 stained the lymphatic endothelial cells and cancer-associated fibroblasts in melanoma tissues, although it did not stain any lymphatic endothelial cells in normal tissues. PMab-38 could be useful for uncovering the function of PDPN in canine melanomas. PMID:27918691

  12. Characterization of Monoclonal Antibody LpMab-3 Recognizing Sialylated Glycopeptide of Podoplanin

    PubMed Central

    Oki, Hiroharu; Ogasawara, Satoshi; Kaneko, Mika Kato; Takagi, Michiaki; Yamauchi, Masanori

    2015-01-01

    Podoplanin (PDPN/Aggrus/T1α/gp36/OTS-8), a type I transmembrane sialoglycoprotein, is involved in platelet aggregation, cell invasion, and cancer metastasis. Podoplanin expression in cancer cells or cancer-associated fibroblasts was reported to be involved in poor prognosis of several cancers. Furthermore, podoplanin is expressed in lymphatic endothelial cells or lung type I alveolar cells. Although many anti-podoplanin monoclonal antibodies (MAbs), such as NZ-1 and D2–40, have been established, almost all anti-podoplanin MAbs are produced against a platelet aggregation-inducing (PLAG) domain. In this study, we produced and characterized a novel anti-podoplanin monoclonal antibody, LpMab-3, the epitope of which is a sialylated glycopeptide of podoplanin. We identified the minimum epitope of LpMab-3 as Thr76–Glu81 of human podoplanin, which is different from PLAG domain, using Western blot analysis and flow cytometry. Immunohistochemical analysis showed that LpMab-3 is useful for detecting lung type I alveolar cells and lymphatic endothelial cells. Because LpMab-3 detects only sialylated podoplanin, it could be useful for uncovering the physiological function of sialylated human podoplanin. PMID:25723283

  13. A Cancer-specific Monoclonal Antibody Recognizes the Aberrantly Glycosylated Podoplanin

    PubMed Central

    Kato, Yukinari; Kaneko, Mika Kato

    2014-01-01

    Podoplanin (PDPN/Aggrus/T1α), a platelet aggregation-inducing mucin-like sialoglycoprotein, is highly expressed in many cancers and normal tissues. A neutralizing monoclonal antibody (mAb; NZ-1) can block the association between podoplanin and C-type lectin-like receptor-2 (CLEC-2) and inhibit podoplanin-induced cancer metastasis, but NZ-1 reacts with podoplanin-expressing normal cells such as lymphatic endothelial cells. In this study, we established a cancer-specific mAb (CasMab) against human podoplanin. Aberrantly glycosylated podoplanin including keratan sulfate or aberrant sialylation, which was expressed in LN229 glioblastoma cells, was used as an immunogen. The newly established LpMab-2 mAb recognized both an aberrant O-glycosylation and a Thr55-Leu64 peptide from human podoplanin. Because LpMab-2 reacted with podoplanin-expressing cancer cells but not with normal cells, as shown by flow cytometry and immunohistochemistry, it is an anti-podoplanin CasMab that is expected to be useful for molecular targeting therapy against podoplanin-expressing cancers. PMID:25080943

  14. Podoplanin Expression Correlates with Disease Progression in Mycosis Fungoides.

    PubMed

    Jankowska-Konsur, Alina; Kobierzycki, Christopher; Grzegrzółka, Jędrzej; Piotrowska, Aleksandra; Gomulkiewicz, Agnieszka; Glatzel-Plucinska, Natalia; Reich, Adam; Podhorska-Okołów, Marzenna; Dzięgiel, Piotr; Szepietowski, Jacek C

    2017-02-08

    The aim of this study was to investigate the role of lymphangiogenesis in the clinical progression and outcome of mycosis fungoides. Immunohistochemistry and Western blot techniques were used to assess the expression of podoplanin and vascular endothelial growth factor C in mycosis fungoides. Expression of vascular endothelial growth factor C measured by immunohistochemistry was significantly higher in mycosis fungoides samples in comparison with control cases (chronic benign dermatoses) (p = 0.0012). Increased expression of podoplanin was found in advanced vs. early mycosis fungoides (p < 0.0001), and was positively correlated with cutaneous and nodal involvement (p < 0.001, p < 0.0001; respectively). Higher podoplanin expression was also significantly associated with shorter survival (p < 0.001). Strong positive correlation was observed between expression of podoplanin analysed by immunohistochemistry and Western blot (r = 0.75, p < 0.0001). A similar association was shown regarding expression of vascular endothelial growth factor C (r = 0.68, p = 0.0007). In conclusion, these results suggest that increased expression of podoplanin is associated with poor clinical course, as well as shorter survival, of patients with mycosis fungoides.

  15. Podoplanin expression in adamantinoma of long bones and osteofibrous dysplasia.

    PubMed

    Kashima, Takeshi G; Dongre, Arunthati; Flanagan, Adrienne M; Hogendoorn, Pancras C W; Taylor, Richard; Athanasou, Nicholas A

    2011-07-01

    Adamantinoma of long bones (ALB) and osteofibrous dysplasia (OFD) are rare osteolytic bone tumours that principally arise in the tibia. Both ALB and OFD contain epithelial and stromal elements, as well as areas of fibro-osseous proliferation. We assessed expression of podoplanin, a glycoprotein found in osteocytes, in OFD and ALB as well as in fibrous dysplasia and metastatic cancer. Forty-two cases of ALB and OFD, 20 cases of fibrous dysplasia and 20 cases of metastatic carcinoma to bone were stained by immunohistochemistry for expression of podoplanin, epithelial (cytokeratin, epithelial membrane antigen) and vascular (CD34, LYVE-1) markers. Podoplanin was expressed in epithelial cells and tumour glands in ALB as well as in scattered intertrabecular stromal cells in both ALB and OFD. Podoplanin was not expressed by intertrabecular stromal cells in fibrous dysplasia or in metastatic adenocarcinoma. Podoplanin was expressed by osteocytes but not osteoblasts of woven and lamellar bone trabeculae in ALB, OFD, fibrous dysplasia and skeletal metastases. The finding of a common osteocyte marker in OFD/ALB stromal cells is in keeping with a close histogenetic relationship between OFD and ALB; this may reflect the prominence of fibro-osseous proliferation in these tumours. The expression of podoplanin in an osteolytic tumour of the tibia may be useful as a diagnostic discriminant in distinguishing OFD from fibrous dysplasia and ALB from metastatic adenocarcinoma.

  16. Podoplanin expression in the development and progression of laryngeal squamous cell carcinomas

    PubMed Central

    2010-01-01

    Background Podoplanin expression is attracting interest as a marker for cancer diagnosis and prognosis. We therefore investigated the expression pattern and clinical significance of podoplanin during the development and progression of laryngeal carcinomas. Results Podoplanin expression was determined by immunohistochemistry in paraffin-embedded tissue specimens from 84 patients with laryngeal premalignancies and 53 patients with laryngeal squamous cell carcinomas. We found podoplanin expression extending from the basal to the suprabasal layer of the epithelium in 37 (44%) of 84 dysplastic lesions, whereas normal epithelium showed negligible expression. Patients carrying podoplanin-positive lesions had a higher laryngeal cancer incidence than those with negative expression reaching borderline statistical significance (51% versus 30%, P = 0.071). Podoplanin expression in laryngeal carcinomas exhibited two distinct patterns. 20 (38%) cases showed diffuse expression in most tumour cells and 33 (62%) focal expression at the proliferating periphery of tumour nests. High podoplanin expression was inversely correlated with T classification (P = 0.033), disease stage (P = 0.006), and pathological grade (P = 0.04). There was a trend, although not significant, towards reduced disease-specific survival for patients with low podoplanin levels (P = 0.31) and diffuse expression pattern (P = 0.08). Conclusions Podoplanin expression increases in the early stages of laryngeal tumourigenesis and it seems to be associated with a higher laryngeal cancer risk. Podoplanin expression in laryngeal squamous cell carcinomas, however, diminishes during tumour progression. Taken together, these data support a role for podoplanin expression in the initiation but not in the progression of laryngeal cancers. PMID:20196862

  17. Single cell time-lapse analysis reveals that podoplanin enhances cell survival and colony formation capacity of squamous cell carcinoma cells

    PubMed Central

    Miyashita, Tomoyuki; Higuchi, Youichi; Kojima, Motohiro; Ochiai, Atsushi; Ishii, Genichiro

    2017-01-01

    Tumor initiating cells (TICs) are characterized by high clonal expansion capacity. We previously reported that podoplanin is a TIC-specific marker for the human squamous cell carcinoma cell line A431. The aim of this study is to explore the molecular mechanism underlying the high clonal expansion potential of podoplanin-positive A431cells using Fucci imaging. Single podoplanin-positive cells created large colonies at a significantly higher frequency than single podoplanin-negative cells, whereas no difference was observed between the two types of cells with respect to cell cycle status. Conversely, the cell death ratio of progenies derived from podoplanin-positive single cell was significantly lower than that of cells derived from podoplanin-negative cells. Single A431 cells, whose podoplanin expression was suppressed by RNA interference, exhibited increased cell death ratios and decreased frequency of large colony forming. Moreover, the frequency of large colony forming decreased significantly when podoplanin-positive single cells was treated with a ROCK (Rho-associated coiled-coil kinase) inhibitor, whereas no difference was observed in single podoplanin-negative cells. Our current study cleared that high clonal expansion capacity of podoplanin-positive TICs populations was the result of reduced cell death by podoplanin-mediated signaling. Therefore, podoplanin activity may be a therapeutic target in the treatment of squamous cell carcinomas. PMID:28059107

  18. Novel Identity and Functional Markers for Human Corneal Endothelial Cells

    PubMed Central

    Bartakova, Alena; Alvarez-Delfin, Karen; Weisman, Alejandra D.; Salero, Enrique; Raffa, Gabriella A.; Merkhofer, Richard M.; Kunzevitzky, Noelia J.; Goldberg, Jeffrey L.

    2016-01-01

    Purpose Human corneal endothelial cell (HCEC) density decreases with age, surgical complications, or disease, leading to vision impairment. Such endothelial dysfunction is an indication for corneal transplantation, although there is a worldwide shortage of transplant-grade tissue. To overcome the current poor donor availability, here we isolate, expand, and characterize HCECs in vitro as a step toward cell therapy. Methods Human corneal endothelial cells were isolated from cadaveric corneas and expanded in vitro. Cell identity was evaluated based on morphology and immunocytochemistry, and gene expression analysis and flow cytometry were used to identify novel HCEC-specific markers. The functional ability of HCEC to form barriers was assessed by transendothelial electrical resistance (TEER) assays. Results Cultured HCECs demonstrated canonical morphology for up to four passages and later underwent endothelial-to-mesenchymal transition (EnMT). Quality of donor tissue influenced cell measures in culture including proliferation rate. Cultured HCECs expressed identity markers, and microarray analysis revealed novel endothelial-specific markers that were validated by flow cytometry. Finally, canonical HCECs expressed higher levels of CD56, which correlated with higher TEER than fibroblastic HCECs. Conclusions In vitro expansion of HCECs from cadaveric donor corneas yields functional cells identifiable by morphology and a panel of novel markers. Markers described correlated with function in culture, suggesting a basis for cell therapy for corneal endothelial dysfunction. PMID:27196322

  19. Aberrant Lymphatic Endothelial Progenitors in Lymphatic Malformation Development

    PubMed Central

    Wu, June K.; Kitajewski, Christopher; Reiley, Maia; Keung, Connie H.; Monteagudo, Julie; Andrews, John P.; Liou, Peter; Thirumoorthi, Arul; Wong, Alvin

    2015-01-01

    Lymphatic malformations (LMs) are vascular anomalies thought to arise from dysregulated lymphangiogenesis. These lesions impose a significant burden of disease on affected individuals. LM pathobiology is poorly understood, hindering the development of effective treatments. In the present studies, immunostaining of LM tissues revealed that endothelial cells lining aberrant lymphatic vessels and cells in the surrounding stroma expressed the stem cell marker, CD133, and the lymphatic endothelial protein, podoplanin. Isolated patient-derived CD133+ LM cells expressed stem cell genes (NANOG, Oct4), circulating endothelial cell precursor proteins (CD90, CD146, c-Kit, VEGFR-2), and lymphatic endothelial proteins (podoplanin, VEGFR-3). Consistent with a progenitor cell identity, CD133+ LM cells were multipotent and could be differentiated into fat, bone, smooth muscle, and lymphatic endothelial cells in vitro. CD133+ cells were compared to CD133− cells isolated from LM fluids. CD133− LM cells had lower expression of stem cell genes, but expressed circulating endothelial precursor proteins and high levels of lymphatic endothelial proteins, VE-cadherin, CD31, podoplanin, VEGFR-3 and Prox1. CD133− LM cells were not multipotent, consistent with a differentiated lymphatic endothelial cell phenotype. In a mouse xenograft model, CD133+ LM cells differentiated into lymphatic endothelial cells that formed irregularly dilated lymphatic channels, phenocopying human LMs. In vivo, CD133+ LM cells acquired expression of differentiated lymphatic endothelial cell proteins, podoplanin, LYVE1, Prox1, and VEGFR-3, comparable to expression found in LM patient tissues. Taken together, these data identify a novel LM progenitor cell population that differentiates to form the abnormal lymphatic structures characteristic of these lesions, recapitulating the human LM phenotype. This LM progenitor cell population may contribute to the clinically refractory behavior of LMs. PMID:25719418

  20. Novel function for blood platelets and podoplanin in developmental separation of blood and lymphatic circulation.

    PubMed

    Uhrin, Pavel; Zaujec, Jan; Breuss, Johannes M; Olcaydu, Damla; Chrenek, Peter; Stockinger, Hannes; Fuertbauer, Elke; Moser, Markus; Haiko, Paula; Fässler, Reinhard; Alitalo, Kari; Binder, Bernd R; Kerjaschki, Dontscho

    2010-05-13

    During embryonic development, lymph sacs form from the cardinal vein, and sprout centrifugally to form mature lymphatic networks. Separation of the lymphatic from the blood circulation by a hitherto unknown mechanism is essential for the homeostatic function of the lymphatic system. O-glycans on the lymphatic endothelium have recently been suggested to be required for establishment and maintenance of distinct blood and lymphatic systems, primarily by mediating proper function of podoplanin. Here, we show that this separation process critically involves platelet activation by podoplanin. We found that platelet aggregates build up in wild-type embryos at the separation zone of podoplanin(+) lymph sacs and cardinal veins, but not in podoplanin(-/-) embryos. Thus, podoplanin(-/-) mice develop a "nonseparation" phenotype, characterized by a blood-filled lymphatic network after approximately embryonic day 13.5, which, however, partially resolves in postnatal mice. The same embryonic phenotype is also induced by treatment of pregnant mice with acetyl salicylic acid, podoplanin-blocking antibodies, or by inactivation of the kindlin-3 gene required for platelet aggregation. Therefore, interaction of endothelial podoplanin of the developing lymph sac with circulating platelets from the cardinal vein is critical for separating the lymphatic from the blood vascular system.

  1. The origin of endothelial cells in novel structures, Bonghan ducts and Bonghan corpuscles determined using immunofluorescence.

    PubMed

    Yi, Sun-Shin; Hwang, In-Koo; Kim, Min-Su; Soh, Kwang-Sup; Yoon, Yeo-Sung

    2009-09-01

    Bonghan ducts (BHDs), and their associated Bonghan corpuscles (BHCs), which are novel threadlike structures, were recently observed in rats and rabbits by using various methods. As further support for the putative circulatory function of the novel threadlike structures (NTS), we investigated the presence and the origin of the endothelial cells within these structures. We immunostained the NTS with anti-CD146, an endothelial cell marker, and with anti-podoplanin, a lymphatic cell marker. Positive expression of CD146 in the BHDs was obtained, and the distribution of endothelial cells showed that the inner boundaries of the channels in the subducts branched from the BHDs and curled around, in a complicated manner, inside a BHCs. The negative expression of podoplanin implies that the endothelial cells in the BHDs are likely to be of vascular and not of lymphatic origin.

  2. Kaposi's sarcoma-associated herpesvirus infection of blood endothelial cells induces lymphatic differentiation.

    PubMed

    Carroll, Patrick A; Brazeau, Elizabeth; Lagunoff, Michael

    2004-10-10

    Kaposi's sarcoma-associated herpesvirus (KSHV) is necessary for KS, a highly vascularized tumor predominated by endothelial-derived spindle cells that express markers of lymphatic endothelium. Following KSHV infection of TIME cells, an immortalized human dermal microvascular endothelial cell (DMVEC) line, expression of many genes specific to lymphatic endothelium, including VEGFR3, podoplanin, LYVE-1, and Prox-1, is significantly increased. Increases in VEGFR3 and podoplanin protein are also demonstrated following latent infection. Examination of cytokine secretion showed that KSHV infection significantly induces hIL-6 while strongly inhibiting secretion of IL-8, a gene product that is decreased by differentiation of blood to lymphatic endothelial cells. These studies support the hypotheses that latent KSHV infection of blood endothelial cells drives their differentiation to lymphatic endothelial cells.

  3. Podoplanin: emerging functions in development, the immune system, and cancer.

    PubMed

    Astarita, Jillian L; Acton, Sophie E; Turley, Shannon J

    2012-01-01

    Podoplanin (PDPN) is a well-conserved, mucin-type transmembrane protein expressed in multiple tissues during ontogeny and in adult animals, including the brain, heart, kidney, lungs, osteoblasts, and lymphoid organs. Studies of PDPN-deficient mice have demonstrated that this molecule plays a critical role in development of the heart, lungs, and lymphatic system. PDPN is widely used as a marker for lymphatic endothelial cells and fibroblastic reticular cells of lymphoid organs and for lymphatics in the skin and tumor microenvironment. Much of the mechanistic insight into PDPN biology has been gleaned from studies of tumor cells; tumor cells often upregulate PDPN as they undergo epithelial-mesenchymal transition and this upregulation is correlated with increased motility and metastasis. The physiological role of PDPN that has been most studied is its ability to aggregate and activate CLEC-2-expressing platelets, as PDPN is the only known endogenous ligand for CLEC-2. However, more recent studies have revealed that PDPN also plays crucial roles in the biology of immune cells, including T cells and dendritic cells. This review will provide a comprehensive overview of the diverse roles of PDPN in development, immunology, and cancer.

  4. Obstructive Sleep Apnoea Syndrome, Endothelial Function and Markers of Endothelialization. Changes after CPAP

    PubMed Central

    Sanchez Armengol, Angeles; Moreno-Luna, Rafael; Caballero-Eraso, Candela; Macher, Hada C.; Villar, Jose; Merino, Ana M; Castell, Javier; Capote, Francisco; Stiefel, Pablo

    2015-01-01

    Study objectives This study tries to assess the endothelial function in vivo using flow-mediated dilatation (FMD) and several biomarkers of endothelium formation/restoration and damage in patients with obstructive sleep apnoea (OSA) syndrome at baseline and after three months with CPAP therapy. Design Observational study, before and after CPAP therapy. Setting and Patients We studied 30 patients with apnoea/hypopnoea index (AHI) >15/h that were compared with themselves after three months of CPAP therapy. FMD was assessed non-invasively in vivo using the Laser-Doppler flowmetry. Circulating cell-free DNA (cf-DNA) and microparticles (MPs) were measured as markers of endothelial damage and the vascular endothelial growth factor (VEGF) was determined as a marker of endothelial restoration process. Measurements and results After three month with CPAP, FMD significantly increased (1072.26 ± 483.21 vs. 1604.38 ± 915.69 PU, p< 0.005) cf-DNA and MPs significantly decreased (187.93 ± 115.81 vs. 121.28 ± 78.98 pg/ml, p<0.01, and 69.60 ± 62.60 vs. 39.82 ± 22.14 U/μL, p<0.05, respectively) and VEGF levels increased (585.02 ± 246.06 vs. 641.11 ± 212.69 pg/ml, p<0.05). These changes were higher in patients with more severe disease. There was a relationship between markers of damage (r = -0.53, p<0.005) but not between markers of damage and restoration, thus suggesting that both types of markers should be measured together. Conclusions CPAP therapy improves FMD. This improvement may be related to an increase of endothelial restoration process and a decrease of endothelial damage. PMID:25815511

  5. Immunohistochemical Examination of Novel Rat Monoclonal Antibodies against Mouse and Human Podoplanin

    PubMed Central

    Kaji, Chiaki; Tsujimoto, Yuta; Kato Kaneko, Mika; Kato, Yukinari; Sawa, Yoshihiko

    2012-01-01

    This study aims to develop new monoclonal antibodies (mAbs) against mouse and human podoplanin. Rats were immunized with synthetic peptides, corresponding to amino acids 38–51 of mouse podoplanin or human podoplanin which is 100% homologous to the same site of monkey podoplanin; anti-mouse podoplanin mAb PMab-1 (IgG2a) and anti-human mAb NZ-1.2 (IgG2a) were established. In immunocytochemistry, the mouse melanoma B16-F10 and mouse podoplanin (mPDPN)-expressed CHO transfectant were stained by PMab-1; human lymphatic endothelial cells (LEC) and human podoplanin (hPDPN)-expressed squamous cell carcinoma HSC3 transfectant, were stained by NZ-1.2. Western-blot analysis detected an about 40-kDa protein in CHO-mPDPN and B16-F10 by PMab-1, and in HSC3-hPDPN and LEC by NZ-1.2. In frozen sections, PMab-1 reacted with mouse kidney, pulmonary alveoli, pulmonary pleura, and salivary gland myoepithelial cells while NZ-1.2 reacted to the human salivary gland myoepithelial cells. The immunostaining of paraffin-embedded sections also showed the reaction of PMab-1 or NZ-1.2 to the mouse or monkey kidney glomerulus, pulmonary alveoli, and lung lymphatic vessels. These results indicate that the two novel rat mAbs to the mouse and human/monkey podoplanin are useful for Western-blot and immunostaining of somatic tissues on paraffin-embedded sections as well as frozen sections. PMID:23012488

  6. High expression of podoplanin in squamous cell carcinoma of the tongue occurs predominantly in patients ≤40 years but does not correlate with tumour spread

    PubMed Central

    Lindell Jonsson, Eva; Boldrup, Linda; Califano, Luigi; Coates, Philip J; Tartaro, Gianpaolo; Lo Muzio, Lorenzo; Fåhraeus, Robin; Colella, Giuseppe; Dell'Aversana Orabona, Giovanni; Loljung, Lotta; Santagata, Mario; Rossiello, Riccardo; Wilms, Torben; Danielsson, Karin; Laurell, Göran

    2015-01-01

    Abstract More than 30% of patients with squamous cell carcinoma (SCC) of the mobile tongue have clinically undetectable lymph node metastasis. Tumour cells can spread as single cells or collectively. A protein known to play a role in both processes is podoplanin, which is expressed in endothelial cells not only in lymph vessels but also in some aggressive tumours with high invasive and metastatic potential. Here we studied samples from 129 patients with primary SCC of the tongue for expression of podoplanin using immunohistochemistry. mRNA levels were analysed in another 27 cases of tongue SCC with adjacent clinically tumour‐free tongue tissue and 14 tongue samples from healthy donors. Higher levels of podoplanin were seen in tumours compared to both normal tongue and clinically normal tongue in the tumour vicinity. No association was found between levels of podoplanin, presence of lymph node metastases or other clinical factors. Patients aged 40 or less were more likely to express high levels of podoplanin protein compared to older patients (p = 0.027). We conclude that levels of podoplanin in primary tongue SCCs are not associated with lymph node metastases. However, tongue SCCs arising in young patients (≤40 years of age) are more likely to express high levels of podoplanin than tongue SCCs that arise in the more elderly. The data suggest that podoplanin has a distinctive role in young patients, who are known to have a poor prognosis: these patients may, therefore, benefit from podoplanin inhibitory therapies. PMID:27499910

  7. High expression of podoplanin in squamous cell carcinoma of the tongue occurs predominantly in patients ≤40 years but does not correlate with tumour spread.

    PubMed

    Sgaramella, Nicola; Lindell Jonsson, Eva; Boldrup, Linda; Califano, Luigi; Coates, Philip J; Tartaro, Gianpaolo; Lo Muzio, Lorenzo; Fåhraeus, Robin; Colella, Giuseppe; Dell'Aversana Orabona, Giovanni; Loljung, Lotta; Santagata, Mario; Rossiello, Riccardo; Wilms, Torben; Danielsson, Karin; Laurell, Göran; Nylander, Karin

    2016-01-01

    More than 30% of patients with squamous cell carcinoma (SCC) of the mobile tongue have clinically undetectable lymph node metastasis. Tumour cells can spread as single cells or collectively. A protein known to play a role in both processes is podoplanin, which is expressed in endothelial cells not only in lymph vessels but also in some aggressive tumours with high invasive and metastatic potential. Here we studied samples from 129 patients with primary SCC of the tongue for expression of podoplanin using immunohistochemistry. mRNA levels were analysed in another 27 cases of tongue SCC with adjacent clinically tumour-free tongue tissue and 14 tongue samples from healthy donors. Higher levels of podoplanin were seen in tumours compared to both normal tongue and clinically normal tongue in the tumour vicinity. No association was found between levels of podoplanin, presence of lymph node metastases or other clinical factors. Patients aged 40 or less were more likely to express high levels of podoplanin protein compared to older patients (p = 0.027). We conclude that levels of podoplanin in primary tongue SCCs are not associated with lymph node metastases. However, tongue SCCs arising in young patients (≤40 years of age) are more likely to express high levels of podoplanin than tongue SCCs that arise in the more elderly. The data suggest that podoplanin has a distinctive role in young patients, who are known to have a poor prognosis: these patients may, therefore, benefit from podoplanin inhibitory therapies.

  8. Endothelial function markers in parkinsonian patients with hyperhomocysteinemia.

    PubMed

    Bostantjopoulou, Sevasti; Katsarou, Zoe; Frangia, Theodora; Hatzizisi, Olga; Papazisis, Kostas; Kyriazis, George; Kiosseoglou, Gregory; Kazis, Aristidis

    2005-08-01

    Hyperhomocysteinemia is considered a risk factor for vascular disease causing endothelial damage and consequently atherogenesis. The purpose of this study was to investigate the effect of elevated homocysteine on certain biochemical markers of endothelial function in patients with idiopathic Parkinson's disease (PD). Blood homocysteine levels were assessed in 57 PD patients and 40 matched normal controls. Investigation of the C677T 5,10 methylenetetrahydrofolate reductase (MTHFR) genotype was also performed in 43 PD patients. The following markers of endothelial function were assessed: superoxide dismutase (SOD), nitric oxide (NO), sICAM-1 and sE-selectin. Homocysteine levels were found mildly elevated in PD patients particularly in those treated with L-Dopa. MTHFR genotype did not influence significantly this finding. SOD activity was found reduced but it was not correlated to homocysteine levels. All other parameters measured were normal and were not related to hyperhomocysteinemia. Our findings indicate that mild hyperhomocysteinemia in PD patients was not associated with endothelial dysfunction.

  9. Suprabasin as a novel tumor endothelial cell marker

    PubMed Central

    Alam, Mohammad T; Nagao-Kitamoto, Hiroko; Ohga, Noritaka; Akiyama, Kosuke; Maishi, Nako; Kawamoto, Taisuke; Shinohara, Nobuo; Taketomi, Akinobu; Shindoh, Masanobu; Hida, Yasuhiro; Hida, Kyoko

    2014-01-01

    Recent studies have reported that stromal cells contribute to tumor progression. We previously demonstrated that tumor endothelial cells (TEC) characteristics were different from those of normal endothelial cells (NEC). Furthermore, we performed gene profile analysis in TEC and NEC, revealing that suprabasin (SBSN) was upregulated in TEC compared with NEC. However, its role in TEC is still unknown. Here we showed that SBSN expression was higher in isolated human and mouse TEC than in NEC. SBSN knockdown inhibited the migration and tube formation ability of TEC. We also showed that the AKT pathway was a downstream factor of SBSN. These findings suggest that SBSN is involved in the angiogenic potential of TEC and may be a novel TEC marker. PMID:25283635

  10. Emerging roles of podoplanin in vascular development and homeostasis.

    PubMed

    Pan, Yanfang; Xia, Lijun

    2015-12-01

    Podoplanin (PDPN) is a mucin-type O-glycoprotein expressed in diverse cell types, such as lymphatic endothelial cells (LECs) in the vascular system and fibroblastic reticular cells (FRCs) in lymph nodes. PDPN on LECs or FRCs activates CLEC-2 in platelets, triggering platelet activation and/or aggregation through downstream signaling events, including activation of Syk kinase. This mechanism is required to initiate and maintain separation of blood and lymphatic vessels and to stabilize high endothelial venule integrity within lymphnodes. In the vascular system, normal expression of PDPN at the LEC surface requires transcriptional activation of Pdpn by Prox1 and modification of PDPN with core 1-derived O-glycans. This review provides a comprehensive overview of the roles of PDPN in vascular development and lymphoid organ maintenance and discusses the mechanisms that regulate PDPN expression related to its function.

  11. Porcine EPCs downregulate stem cell markers and upregulate endothelial maturation markers during in vitro cultivation.

    PubMed

    Avci-Adali, Meltem; Nolte, Andrea; Simon, Perikles; Ziemer, Gerhard; Wendel, Hans P

    2009-10-01

    In recent years, interest in endothelial progenitor cells (EPCs) in the field of tissue engineering and regenerative medicine has increased tremendously. However, each clinical stem cell application requires prior validation through animal experiments. This study investigates the isolation and characterization of porcine EPCs from peripheral blood and the change of their cell surface marker expression during in vitro cultivation. RT-PCR demonstrated that the EPCs express stem cell markers CD34 and CD133, which decrease with in vitro cultivation time. Throughout the cultivation process EPCs did not express monocytic (CD14) or haematopoietic marker (CD45). Surprisingly, the CD31 and VE-cadherin expression in EPCs was significantly higher than in endothelial cells (ECs). In contrast, the VEGFR2 and E-selectin expression was significantly lower than in ECs, but increased during the expansion process. This study clarifies the characteristic properties of porcine EPCs during cell culture and may help to improve the impact of EPC-based therapies in porcine animal studies.

  12. Ulex europaeus I lectin as a marker for tumors derived from endothelial cells.

    PubMed

    Miettinen, M; Holthofer, H; Lehto, V P; Miettinen, A; Virtanen, I

    1983-01-01

    Some skin and soft tumors, which generally are assumed to be derived from endothelial cells or blood vessels, were characterized with fluorochrome-labeled Ulex europaeus I agglutinin (UEA I), recently shown to bind specifically to endothelial cells in various normal human tissues. The staining pattern was compared with that obtained with immunostaining using antibodies against factor-VIII-related antigen (FVIII-RAG), a known marker for endothelial cells. The results showed that UEA-I is a specific and a more sensitive marker for the endothelial cells in benign vascular lesions as compared with anti-FVIII-RAG. UEA-I also stained many neoplastic cells of endothelial sarcomas, which generally were negative for FVIII-RAG. Melanomas, anaplastic carcinomas, and other types of sarcomas were negative for both UEA-I and FVIII-RAG. The results suggest that UEA-I lectin is a specific and sensitive adjunct tool in demonstrating endothelial cells and endothelial derivation of human tumors.

  13. Magic roundabout, a tumor endothelial marker: expression and signaling.

    PubMed

    Seth, Pankaj; Lin, Yanfeng; Hanai, Jun-ichi; Shivalingappa, Venkatesha; Duyao, Mabel P; Sukhatme, Vikas P

    2005-07-01

    Molecular signals that guide blood vessels to specific paths are not fully deciphered, but are thought to be similar to signals that mediate neuronal guidance. These cues are not only critical for normal blood vessel development, but may also play a major role in tumor angiogenesis. In this study, we have demonstrated the tumor endothelial specific expression of a Robo family member, magic roundabout (MRB), functionally characterized its role in endothelial cell migration and defined a signaling pathway that might mediate this function. We show that MRB is differentially over-expressed in tumor endothelial cells versus normal adult endothelial cells in numerous solid tumors. Moreover, over-expression of MRB in endothelial cells activates MRB in a ligand-independent fashion, and activation of MRB via Slit2, a putative ligand, results in inhibition of VEGF and FGF induced migration. We also demonstrate that MRB induced inhibition of endothelial migration is partially mediated by the Ras-Raf-Mek-Erk signaling pathway. We therefore hypothesize that expression of MRB is involved in regulating the migration of endothelial cells during tumor angiogenesis.

  14. Tumor endothelial markers define novel subsets of cancer-specific circulating endothelial cells associated with antitumor efficacy

    PubMed Central

    Mehran, Reza; Nilsson, Monique; Khajavi, Mehrdad; Du, Zhiqiang; Cascone, Tina; Wu, Hua Kang; Cortes, Andrea; Xu, Li; Zurita, Amado; Schier, Robert; Riedel, Bernhard; El-Zein, Randa; Heymach, John V.

    2014-01-01

    Circulating endothelial cells (CEC) are derived from multiple sources including bone marrow (circulating endothelial progenitors [CEP]) and established vasculature (mature CEC). Although CEC have shown promise as a biomarker for cancer patients, their utility has been limited in part by the lack of specificity for tumor vasculature and the different non-malignant causes that can impact CEC. Tumor endothelial markers (TEM) are antigens enriched in tumor vs non-malignant endothelia. We hypothesized that TEMs may be detectable on CEC and that these circulating TEM+ endothelial cells (CTEC) may be a more specific marker for cancer and tumor response than standard CEC. We found that tumor-bearing mice had a relative increase in numbers of circulating CTEC, specifically with increased levels of TEM7 and TEM8 expression. Following treatment with various vascular targeting agents, we observed a decrease in CTEC that correlated with the reductions in tumor growth. We extended these findings to human clinical samples and observed that CTEC were present in esophageal cancer and non-small cell lung cancer (NSCLC) patients (N=40) and their levels decreased after surgical resection. These results demonstrate that CTEC are detectable in preclinical cancer models and cancer patients. Further, they suggest that CTEC offer a novel cancer-associated marker that may be useful as a blood-based surrogate for assessing the presence of tumor vasculature and antiangiogenic drug activity. PMID:24626092

  15. High Throughput Gene Expression Analysis Identifies Reliable Expression Markers of Human Corneal Endothelial Cells

    PubMed Central

    Chng, Zhenzhi; Peh, Gary S. L.; Herath, Wishva B.; Cheng, Terence Y. D.; Ang, Heng-Pei; Toh, Kah-Peng; Robson, Paul; Mehta, Jodhbir S.; Colman, Alan

    2013-01-01

    Considerable interest has been generated for the development of suitable corneal endothelial graft alternatives through cell-tissue engineering, which can potentially alleviate the shortage of corneal transplant material. The advent of less invasive suture-less key-hole surgery options such as Descemet’s Stripping Endothelial Keratoplasty (DSEK) and Descemet’s Membrane Endothelial Keratoplasty (DMEK), which involve transplantation of solely the endothelial layer instead of full thickness cornea, provide further impetus for the development of alternative endothelial grafts for clinical applications. A major challenge for this endeavor is the lack of specific markers for this cell type. To identify genes that reliably mark corneal endothelial cells (CECs) in vivo and in vitro, we performed RNA-sequencing on freshly isolated human CECs (from both young and old donors), CEC cultures, and corneal stroma. Gene expression of these corneal cell types was also compared to that of other human tissue types. Based on high throughput comparative gene expression analysis, we identified a panel of markers that are: i) highly expressed in CECs from both young donors and old donors; ii) expressed in CECs in vivo and in vitro; and iii) not expressed in corneal stroma keratocytes and the activated corneal stroma fibroblasts. These were SLC4A11, COL8A2 and CYYR1. The use of this panel of genes in combination reliably ascertains the identity of the CEC cell type. PMID:23844023

  16. Urine albumin to creatinine ratio: A marker of early endothelial dysfunction in youth

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The urine albumin-to-creatinine ratio (UACR) is a useful predictor of cardiovascular (CV) events in adults. Its relationship to vascular function in children is not clear. We investigated whether UACR was related to insulin resistance and endothelial function, a marker of subclinical atherosclerosis...

  17. Characterization and microarray analysis of genes in human lymphatic endothelial cells from patients with breast cancer.

    PubMed

    Kawai, Yoshiko; Minami, Takashi; Fujimori, Minoru; Hosaka, Kayoko; Mizuno, Risuke; Ikomi, Fumitaka; Kodama, Tatsuhiko; Ohhashi, Toshio

    2007-01-01

    We successfully isolated human lymphatic endothelial cells from afferent lymph vessels (HALEC) of sentinel lymph nodes in patients with breast cancer by using trypsin digestion. The cells were cultured in EGM-2 medium with 10% FBS under the condition of 5% O2, 5% CO2, and 90% N2 at 37 degrees C. The cultured cells exhibited a monolayer with cobblestone appearance and a marked phagocytosis of Dil-Ac-LDL. Immunohistochemical lymphatic vessel markers were also found, such as podoplanin, LYVE-1, VEGF receptor 3, and Prox-1. Quantitative RT-PCR analysis also showed that podoplanin, VEGF R3, and Prox-1 mRNA were expressed more selectively in the cultured cells. The cells had marked immunoreactivity to antisera of ecNOS, iNOS, COX1, and weak reactivity of COX2. Constitutively expressed cell-type specific genes of the cultured cells were also analyzed by oligonucleotide microarray methods. Compared with human umbilical vein endothelial cells (HUVEC), the cells selectively expressed 88 known genes such as angiopoietin-like 4, oxygen radicals-related enzymes, and adhesion molecules and the related proteoglycans. The findings suggest that the cultured cells seem to be human lymphatic endothelial cells. In conclusion, the isolated, cannulated and enzymatic digested method we adopted for culture of human lymphatic endothelial cells may be easy and useful for investigating cellular, molecular biological, and genomic properties of the cells.

  18. The use of novel lymphatic endothelial cell-specific immunohistochemical markers to differentiate cutaneous angiosarcomas in dogs.

    PubMed

    Halsey, C H C; Worley, D R; Curran, K; Charles, J B; Ehrhart, E J

    2016-09-01

    Lymphangiosarcomas are uncommon vascular neoplasms that arise from lymphatic endothelial cells (LECs). They efface and replace normal subcutaneous tissue and are characterised by arborising, vascular channels lined by a single layer of pleomorphic endothelial cells and a paucity of erythrocytes. Lymphangiosarcomas are architecturally similar to hemangiosarcomas, a common malignancy of vascular origin arising from blood vascular endothelial cells. Common immunohistochemical markers for vascular endothelium, such as Factor VIII-related antigen (F8RA) and CD31, have traditionally been used to confirm the diagnosis of tumours of vascular origin. However, these markers fail to differentiate between lymphangiosarcoma and hemangiosarcoma, which often show overlapping morphologic features, disparate clinical behaviour and require different treatment modalities. Here we describe the use of two novel LEC-specific markers, lymphatic vessel endothelial receptor-1 (LYVE-1) and prospero-related homeobox gene-1 (PROX-1), to further differentiate between vascular tumours of lymphatic (lymphangiosarcoma) and blood (hemangiosarcoma) endothelial cell origin in the dog.

  19. Selective targeting of liver cancer with the endothelial marker CD146

    PubMed Central

    Thomann, Stefan; Longerich, Thomas; Bazhin, Alexandr V.; Mier, Walter; Schemmer, Peter; Ryschich, Eduard

    2014-01-01

    Hepatocellular carcinomas are well-vascularized tumors; the endothelial cells in these tumors have a specific phenotype. Our aim was to develop a new approach for tumor-specific drug delivery with monoclonal antibody targeting of endothelial ligands. CD146, a molecule expressed on the endothelial surface of hepatocellular carcinoma, was identified as a promising candidate for targeting. In the present study, endothelial cells immediately captured circulating anti-CD146 (ME-9F1) antibody, while antibody binding in tumors was significantly higher than in hepatic endothelium. Macroscopically, after intravenous injection, there were no differences in the mean accumulation of anti-CD146 antibody in tumor compared to liver tissue, due to a compensating higher blood vessel density in the liver tissue. Additional blockade of nontumoral epitopes and intra-arterial administration, improved selective antibody capture in the tumor microvasculature and largely prevented antibody distribution in the lung and liver. The potential practical use of this approach was demonstrated by imaging of radionuclide-labeled ME-9F1 antibody, which showed excellent tumor-selective uptake. Our results provide a promising principle for the use of endothelial markers for intratumoral drug delivery. Tumor endothelium–based access might offer new opportunities for the imaging and therapy of hepatocellular carcinoma and other liver malignancies. PMID:25238265

  20. [Effects of mexidol and sulodexide on the level of specific markers of endothelial dysfunction in animals with experimental diabetes mellitus].

    PubMed

    Tiurenkov, I N; Voronkov, A V; Slietsans, A A; Snigur, G L

    2012-01-01

    Streptozotocin-induced diabetes leads to the development of endothelial dysfunction, as evidenced by decreased expression of endothelial nitric oxide synthase (eNOS) and increased expression of endothelin-1 as specific markers of endothelial disorders. All test substances showed endotelioprotective activity by increasing the concentration of eNOS and reducing the level of endothelin-1. With respect to the degree of impact on the eNOS and endothelin-1 levels, the compounds studied can be rated as follows: sulodexide > meksidol.

  1. The Multifaceted Role of Podoplanin Expression in Hepatocellular Carcinoma

    PubMed Central

    Cioca, Andreea; Ceausu, Amalia R.; Marin, Irina; Raica, Marius; Cimpean, Anca Maria

    2017-01-01

    The role of podoplanin in hepatocellular carcinoma (HCC) is not clear yet. The aim of our study was to evaluate the expression of podoplanin in HCC and to determine its role in hepatocarcinogenesis. We performed immunohistochemistry with monoclonal D2-40 antibody, on paraffin-embedded tissue sections of 72 patients diagnosed with HCC. Lymphatic vessels density (LVD) was increased in patients who had vascular invasion at the time of diagnosis (P=0.018) and in those with associated cirrhosis (P=0.006). Tumor cells showing podoplanin expression were correlated with histological grade (P=0.040). Podoplanin-expressing cancer associated fibroblasts (CAFs) were correlated with both LVD (P=0.019) and tumor cells (P=0.015). Our results sustain the dual role of podoplanin in HCC by its involvement in both HCC tumorigenesis, lymphatic neovascularization and tumor invasion invasiveness. A possible crosstalk between epithelial and stromal tumor cells in HCC tumor microenvironment may be mediated by podoplanin, but this hypothesis needs further studies to elucidate this interrelation. PMID:28348421

  2. ERG is a novel and reliable marker for endothelial cells in central nervous system tumors.

    PubMed

    Haber, Matthew A; Iranmahboob, Amir; Thomas, Cheddhi; Liu, Mengling; Najjar, Amanda; Zagzag, David

    2015-01-01

    ETS-related gene (ERG) is a transcription factor that has been linked to angiogenesis. Very little research has been done to assess ERG expression in central nervous system (CNS) tumors. We evaluated 57 CNS tumors, including glioblastomas (GBMs) and hemangioblastomas (HBs), as well as two arteriovenous malformations and four samples of normal brain tissue with immunohistochemistry using a specific ERG rabbit monoclonal antibody. In addition, immunostains for CD31, CD34, and α-smooth muscle actin (α-SMA) were performed on all samples. CD31 demonstrated variable and sometimes weak immunoreactivity for endothelial cells. Furthermore, in 1 case of a GBM, CD34 stained not only endothelial cells, but also tumor cells. In contrast, we observed that ERG was only expressed in the nuclei of endothelial cells, for example, in the hyperplastic vascular complexes that comprise the glomeruloid microvascular proliferation seen in GBMs. Conversely, α-SMA immunoreactivity was identified in the abluminal cells of these hyperplastic vessels. Quantitative evaluation with automated methodology and custom Matlab 2008b software was used to calculate percent staining of ERG in each case. We observed significantly higher quantitative expression of ERG in HBs than in other CNS tumors. Our results show that ERG is a novel, reliable, and specific marker for endothelial cells within CNS tumors that can be used to better study the process of neovascularization.

  3. Discovery of molecular markers to discriminate corneal endothelial cells in the human body.

    PubMed

    Yoshihara, Masahito; Ohmiya, Hiroko; Hara, Susumu; Kawasaki, Satoshi; Hayashizaki, Yoshihide; Itoh, Masayoshi; Kawaji, Hideya; Tsujikawa, Motokazu; Nishida, Kohji

    2015-01-01

    The corneal endothelium is a monolayer of hexagonal corneal endothelial cells (CECs) on the inner surface of the cornea. CECs are critical in maintaining corneal transparency through their barrier and pump functions. CECs in vivo have a limited capacity in proliferation, and loss of a significant number of CECs results in corneal edema called bullous keratopathy which can lead to severe visual loss. Corneal transplantation is the most effective method to treat corneal endothelial dysfunction, where it suffers from donor shortage. Therefore, regeneration of CECs from other cell types attracts increasing interests, and specific markers of CECs are crucial to identify actual CECs. However, the currently used markers are far from satisfactory because of their non-specific expression in other cell types. Here, we explored molecular markers to discriminate CECs from other cell types in the human body by integrating the published RNA-seq data of CECs and the FANTOM5 atlas representing diverse range of cell types based on expression patterns. We identified five genes, CLRN1, MRGPRX3, HTR1D, GRIP1 and ZP4 as novel markers of CECs, and the specificities of these genes were successfully confirmed by independent experiments at both the RNA and protein levels. Notably none of them have been documented in the context of CEC function. These markers could be useful for the purification of actual CECs, and also available for the evaluation of the products derived from other cell types. Our results demonstrate an effective approach to identify molecular markers for CECs and open the door for the regeneration of CECs in vitro.

  4. Flow cytometry-based isolation of dermal lymphatic endothelial cells from newborn rats.

    PubMed

    Thiele, W; Rothley, M; Schmaus, A; Plaumann, D; Sleeman, J

    2014-12-01

    The lymphatic system plays a key role in tissue homeostasis, fatty acid transport, and immune surveillance. Pathologically, dysfunction of the lymphatic system results in edema, and increased lymphangiogenesis can contribute to tumor metastasis. Lymphatic vessels are composed of lymphatic endothelial cells (LECs) that can be identified by distinct marker molecules such as Prox-1, podoplanin, VEGFR-3 and LYVE-1. Primary LECs represent a valuable tool for the study of basic functions of the lymphatic system. However, their isolation remains a challenge, particularly if rodent tissues are used as a source. We developed a method for the isolation of rat dermal LECs from the skin of newborn rats based on sequential enzymatic digestion with trypsin and Liberase followed by flow cytometric sorting using LYVE-1 specific antibodies. Cells isolated according to this protocol expressed the lymphatic markers Prox-1, podoplanin, LYVE-1 and VEGFR-3, and displayed an endothelial-like morphology when taken into culture. These primary cells can be used for studying lymphatic biology in rat models, and the protocol we describe here therefore represents an important extension of the experimental repertoire available for rats and for modeling the human lymphatic system.

  5. Superior sensitivity of novel molecular imaging probe: simultaneously targeting two types of endothelial injury markers

    PubMed Central

    Sun, Dawei; Nakao, Shintaro; Xie, Fang; Zandi, Souska; Schering, Alexander; Hafezi-Moghadam, Ali

    2010-01-01

    The need remains great for early diagnosis of diseases. The special structure of the eye provides a unique opportunity for noninvasive light-based imaging of fundus vasculature. To detect endothelial injury at the early and reversible stage of adhesion molecule up-regulation, we generated novel imaging agents that target two distinct types of endothelial molecules, a mediator of rolling, P-selectin, and one that mediates firm adhesion, ICAM-1. Interactions of these double-conjugated fluorescent microspheres (MSs) in retinal or choroidal microvasculature were visualized in live animals by scanning laser ophthalmoscopy. The new imaging agents showed significantly higher sensitivity for detection of endothelial injury than singly conjugated MSs (rPSGL-1- or α-ICAM-1-conjugated), both in terms of rolling (P<0.01) and firm adhesion (P<0.01). The rolling flux of α-ICAM-1-conjugated MSs did not differ in EIU animals, whereas double-conjugated MSs showed significantly higher rolling flux (P<0.01), revealing that ICAM-1 in vivo supports rolling, once MS interaction with the endothelium is initiated. Double-conjugated MSs specifically detected firmly adhering leukocytes (P<0.01), allowing in vivo quantification of immune response. Antiinflammatory treatment with dexamethasone led to reduced leukocyte accumulation (P<0.01) as well as MS interaction (P<0.01), which suggests that treatment success and resolution of inflammation is quantitatively reflected with this molecular imaging approach. This work introduces novel imaging agents for noninvasive detection of endothelial injury in vivo. Our approach may be developed further to diagnose human disease at a much earlier stage than currently possible.—Sun, D., Nakao, S., Xie, F., Zandi, S., Schering, A., Hafezi-Moghadam, A. Superior sensitivity of novel molecular imaging probe: simultaneously targeting two types of endothelial injury markers. PMID:20103715

  6. Tumor endothelial marker 5 expression in endothelial cells during capillary morphogenesis is induced by the small GTPase Rac and mediates contact inhibition of cell proliferation

    SciTech Connect

    Vallon, Mario; Rohde, Franziska; Janssen, Klaus-Peter; Essler, Markus

    2010-02-01

    Tumor endothelial marker (TEM) 5 is an adhesion G-protein-coupled receptor upregulated in endothelial cells during tumor and physiologic angiogenesis. So far, the mechanisms leading to upregulation of TEM5 and its function during angiogenesis have not been identified. Here, we report that TEM5 expression in endothelial cells is induced during capillary-like network formation on Matrigel, during capillary morphogenesis in a three-dimensional collagen I matrix, and upon confluence on a two-dimensional matrix. TEM5 expression was not induced by a variety of soluble angiogenic factors, including VEGF and bFGF, in subconfluent endothelial cells. TEM5 upregulation was blocked by toxin B from Clostridium difficile, an inhibitor of the small GTPases Rho, Rac, and Cdc42. The Rho inhibitor C3 transferase from Clostridium botulinum did not affect TEM5 expression, whereas the Rac inhibitor NSC23766 suppressed TEM5 upregulation. An excess of the soluble TEM5 extracellular domain or an inhibitory monoclonal TEM5 antibody blocked contact inhibition of endothelial cell proliferation resulting in multilayered islands within the endothelial monolayer and increased vessel density during capillary formation. Based on our results we conclude that TEM5 expression during capillary morphogenesis is induced by the small GTPase Rac and mediates contact inhibition of proliferation in endothelial cells.

  7. Chimeric Anti-Human Podoplanin Antibody NZ-12 of Lambda Light Chain Exerts Higher Antibody-Dependent Cellular Cytotoxicity and Complement-Dependent Cytotoxicity Compared with NZ-8 of Kappa Light Chain.

    PubMed

    Kaneko, Mika K; Abe, Shinji; Ogasawara, Satoshi; Fujii, Yuki; Yamada, Shinji; Murata, Takeshi; Uchida, Hiroaki; Tahara, Hideaki; Nishioka, Yasuhiko; Kato, Yukinari

    2017-02-01

    Podoplanin (PDPN), a type I transmembrane 36-kDa glycoprotein, is expressed not only in normal cells, such as renal epithelial cells (podocytes), lymphatic endothelial cells, and pulmonary type I alveolar cells, but also in cancer cells, including brain tumors and lung squamous cell carcinomas. Podoplanin activates platelet aggregation by binding to C-type lectin-like receptor-2 (CLEC-2) on platelets, and the podoplanin/CLEC-2 interaction facilitates blood/lymphatic vessel separation. We previously produced neutralizing anti-human podoplanin monoclonal antibody (mAb), clone NZ-1 (rat IgG2a, lambda), which neutralizes the podoplanin/CLEC-2 interaction and inhibits platelet aggregation and cancer metastasis. Human-rat chimeric antibody, NZ-8, was previously developed using variable regions of NZ-1 and human constant regions of heavy chain (IgG1) and light chain (kappa chain). Although NZ-8 showed high antibody-dependent cellular cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC) against human podoplanin-expressing cancer cells, the binding affinity of NZ-8 was lower than that of NZ-1. Herein, we produced a novel human-rat chimeric antibody, NZ-12, the constant regions of which consist of IgG1 heavy chain and lambda light chain. Using flow cytometry, we demonstrated that the binding affinity of NZ-12 was much higher than that of NZ-8. Furthermore, ADCC and CDC activities of NZ-12 were significantly increased against glioblastoma cell lines (LN319 and D397) and lung cancer cell line (PC-10). These results suggested that NZ-12 could become a promising therapeutic antibody against podoplanin-expressing brain tumors and lung cancers.

  8. Endothelial Dysfunction: An Early Cardiovascular Risk Marker in Asymptomatic Obese Individuals with Prediabetes

    PubMed Central

    Gupta, Alok K.; Ravussin, Eric; Johannsen, Darcy L.; Stull, April J.; Cefalu, William T.; Johnson, William D.

    2012-01-01

    Aims To elucidate if endothelial dysfunction is an early CV risk marker in obese men and women with prediabetes. Study Design Cross-sectional study. Place and Duration of Study Clinical Research Unit, Pennington Biomedical Research Center, Baton Rouge, LA. United States. Methodology Overweight and obese status denotes an increasing adipose tissue burden which spills over into ectopic locations, including the visceral compartment, muscle and liver. Associated co-morbidities enhance cardiovascular (CV) risk. Endothelium which is the largest receptor-effector end-organ in our bodies, while responding to numerous physical and chemical stimuli maintains vascular homeostasis. Endothelial dysfunction (ED) is the initial perturbation, which precedes fatty streak known to initiate atherosclerosis: insidious process which often culminates as sudden catastrophic CV adverse event. Asymptomatic men and women; [n=42] coming in after an overnight fast had demographic, anthropometric, clinical chemistry and resting endothelial function [EF: increased test finger peripheral arterial tone (PAT) relative to control; expressed as relative hyperemia index (RHI)] assessments. Results Adults with desirable weight [n=12] and overweight [n=8] state, had normal fasting plasma glucose [Mean(SD)]: FPG [91.1(4.5), 94.8(5.8) mg/dL], insulin [INS, 2.3(4.4), 3.1(4.8) μU/ml], insulin sensitivity by homeostasis model assessment [HOMA-IR, 0.62(1.2), 0.80(1.2)] and desirable resting clinic blood pressure [SBP/DBP, 118(12)/74(5), 118(13)/76(8) mmHg]. Obese adults [n=22] had prediabetes [FPG, 106.5(3.5) mg/dL], hyperinsulinemia [INS 18.0(5.2) μU/ml], insulin resistance [HOMA-IR 4.59(2.3)], prehypertension [PreHTN; SBP/DBP 127(13)/81(7) mmHg] and endothelial dysfunction [ED; reduced RHI 1.7(0.3) vs. 2.4(0.3); all p<0.05]. Age-adjusted RHI correlated with BMI [r=−0.53; p<0.001]; however, BMI-adjusted RHI was not correlated with age [r=−0.01; p=0.89]. Conclusion Endothelial dysfunction reflective of

  9. Biomarkers Discovery for Colorectal Cancer: A Review on Tumor Endothelial Markers as Perspective Candidates

    PubMed Central

    2016-01-01

    Colorectal cancer (CRC) is the third most common cancer in the world. The early detection of CRC, during the promotion/progression stages, is an enormous challenge for a successful outcome and remains a fundamental problem in clinical approach. Despite the continuous advancement in diagnostic and therapeutic methods, there is a need for discovery of sensitive and specific, noninvasive biomarkers. Tumor endothelial markers (TEMs) are associated with tumor-specific angiogenesis and are potentially useful to discriminate between tumor and normal endothelium. The most promising TEMs for oncogenic signaling in CRC appeared to be the TEM1, TEM5, TEM7, and TEM8. Overexpression of TEMs especially TEM1, TEM7, and TEM8 in colorectal tumor tissue compared to healthy tissue suggests their role in tumor blood vessels formation. Thus TEMs appear to be perspective candidates for early detection, monitoring, and treatment of CRC patients. This review provides an update on recent data on tumor endothelial markers and their possible use as biomarkers for screening, diagnosis, and therapy of colorectal cancer patients. PMID:27965519

  10. Coagulopathy in patients with acute pulmonary embolism: a pilot study of whole blood coagulation and markers of endothelial damage.

    PubMed

    Lehnert, Per; Johansson, Pär I; Ostrowski, Sisse R; Møller, Christian H; Bang, Lia E; Olsen, Peter Skov; Carlsen, Jørn

    2017-02-01

    Whole blood coagulation and markers of endothelial damage were studied in patients with acute pulmonary embolism (PE), and evaluated in relation to PE severity. Twenty-five patients were enrolled prospectively each having viscoelastical analysis of whole blood done using thrombelastography (TEG) and Multiplate aggregometry. Fourteen of these patients were investigated for endothelial damage by ELISA measurements of Syndecan-1 (endothelial glycocalyx degradation), soluble endothelial Selectin (endothelial cell activation), soluble Thrombomodulin (endothelial cell injury) and Histone Complexed DNA fragments (endothelial cytotoxic histones). The mean values of TEG and Multiplate parameters were all within the reference levels, but a significant difference between patients with high and intermediate risk PE was observed for Ly30 (lytic activity) 1.5% [0-10] vs. 0.2% [0-2.2] p = .04, and ADP (platelet reactivity) 92 U [20-145] vs. 59 U [20-111] p = .03. A similar difference was indicated for functional fibrinogen 21 mm [17-29] vs. 18 mm [3-23] p = .05. Analysis of endothelial markers identified a significant difference in circulating levels between high and intermediate risk PE patients for Syndecan-1 118.6 ng/mL [76-133] vs. 36.3 ng/mL [11.8-102.9] p = .008. In conclusion, patients with acute PE had normal whole blood coagulation, but high risk PE patients had signs of increased activity of the haemostatic system and significantly increased level of endothelial glycocalyx degradation.

  11. Differential Expression of Stem Cell Markers and Vascular Endothelial Growth Factor in Human Retinoblastoma Tissue

    PubMed Central

    Kim, Martha; Kim, Jeong Hun; Kim, Jin Hyoung; Kim, Dong Hun

    2010-01-01

    Purpose To investigate the relationship between vascular endothelial growth factor (VEGF) and the cancer stem cell-vascular niche complex in human retinoblastoma tissue. Methods Six human retinoblastoma specimens primarily enucleated for Reese-Ellsworth classification stage 5a were stained to detect cancer stem cell markers, including ABCG2 for the stem cell marker and MCM2 for the neural stem cell marker, as well as to detect VEGF for the angiogenic cytokine. Using immunofluorescence, the expression of these proteins was analyzed, and their relative locations noted. Results In non-neoplastic retina of tumor-bearing eyes, ABCG2 and MCM2 were sporadically expressed in the ganglion cell layer and the inner nuclear layer, whereas VEGF was sporadically expressed in inner retina where retinal vessels are abundantly distributed. In the tumor, ABCG2 was strongly expressed out of Wintersteiner rosettes, whereas MCM2 and VEGF were strongly stained in the rosettes. Interestingly, the outer portion of the rosettes was positive for MCM2, and the inner portion of the rosettes was positive for VEGF. Conclusions Our data demonstrated that MCM2 and VEGF are strongly expressed in the rosettes of the tumor, which were far from the area of ABCG2-positive cells. Although VEGF might not directly contribute to the cancer stem cell-vascular niche complex, it could play some role in the differentiation of tumor cells to build up the rosettes. PMID:20157412

  12. Immunohistochemical Expression of VEGF and Podoplanin in Uterine Cervical Squamous Intraepithelial Lesions

    PubMed Central

    Belfort-Mattos, Patrícia Napoli; Focchi, Gustavo Rubino de Azevedo; Ribalta, Julisa Chamorro Lascasas; Megale De Lima, Tatiana; Nogueira Carvalho, Carmen Regina; Kesselring Tso, Fernanda; De Góis Speck, Neila Maria

    2016-01-01

    VEGF and podoplanin (PDPN) have been identified as angiogenesis and/or lymphangiogenesis regulators and might be essential to restrict tumor growth, progression, and metastasis. In the present study, we evaluate the association between the expression of these markers and CIN grade. Immunohistochemistry was performed in 234 uterine cervical samples using conventional histologic sections or TMA with the monoclonal antibodies to VEGF (C-1 clone) and podoplanin (D2-40 clone). Positive-staining rates of VEGF in 191 CIN specimens were significantly associated with histological grade (P < 0.001). Negative and/or focal immunostaining for PDPN were more frequent in CIN 3 (P = 0.016). We found that patients with CIN 3 more frequently had strong and more diffuse staining for VEGF and diminished staining for PDPN (P = 0.018). Strong and more diffuse VEGF immunoexpressions in CIN 2 and CIN 3 were detected when compared to CIN 1. Negative and/or focal PDPN immunoexpression appear to be more frequent in CIN 3. Moderate to strong VEGF expression may be a tendency among patients with high-grade lesions and diminished PDPN expression. PMID:27313335

  13. Endothelial cell labeling with indium-111-oxine as a marker of cell attachment to bioprosthetic surfaces

    SciTech Connect

    Sharefkin, J.B.; Lather, C.; Smith, M.; Rich, N.M.

    1983-03-01

    Canine vascular endothelium labeled with indium-111-oxine was used as a marker of cell attachment to vascular prosthetic surfaces with complex textures. Primarily cultured and freshly harvested endothelial cells both took up the label rapidly. An average of 72% of a 32 micro Ci labeling dose was taken up by 1.5 X 10(6) cells in 10 min in serum-free medium. Over 95% of freshly labeled cells were viable by trypan blue tests and only 5% of the label was released after 1 h incubations at 37 degrees C. Labeled and unlabeled cells had similar rates of attachment to plastic dishes. Scanning electron microscopic studies showed that labeled cells retained their ability to spread on tissue culture dishes even at low (1%) serum levels. Labeled endothelial cells seeded onto Dacron or expanded polytetrafluoroethylene vascular prostheses by methods used in current surgical models could be identified by autoradiography of microscopic sections of the prostheses, and the efficiency of cell attachment to the prosthesis could be measured by gamma counting. Indium-111 labeling affords a simple and rapid way to measure initial cell attachment to, and distribution on, vascular prosthetic materials. The method could also allow measurement of early cell loss from a flow surface in vivo by using external gamma imaging.

  14. Antibodies to endothelial cells in systemic lupus erythematosus: a potential marker for nephritis and vasculitis.

    PubMed

    D'Cruz, D P; Houssiau, F A; Ramirez, G; Baguley, E; McCutcheon, J; Vianna, J; Haga, H J; Swana, G T; Khamashta, M A; Taylor, J C

    1991-08-01

    Using an ELISA, anti-endothelial cell antibodies (AECA) have been found in sera obtained at the time of renal biopsy in 46 out of 57 patients (81%) with systemic lupus erythematosus (SLE) and nephritis (mean binding index (BI) = 84% +/- 52.8) compared with 22 out of 50 SLE patients (44%) without nephritis (mean BI = 45% +/- 35.9). Seventy normal human sera had a mean BI of 10% +/- 9.8. The highest levels were seen in patients with diffuse proliferative glomerulonephritis (WHO grade IV) and in patients with proteinuria and nephrotic syndrome. When the biopsies were assessed for activity and chronicity scores, AECA were associated with active renal lesions (P less than 0.001). AECA levels correlated with low complement levels but not with anti-DNA antibodies to extractable nuclear antigens (ENA), anti-cardiolipin or anti-neutrophil cytoplasmic antibodies. The presence of AECA conferred a positive predictive value of 0.68 for the presence of nephritis. Twenty-five patients had active vasculitis at the time of assay and the highest AECA values were seen in patients with both nephritis and vasculitis. No correlation was seen with serum immunoglobulin levels and immune complexes did not bind significantly to the endothelial surface. The possible role of these antibodies as a marker in lupus nephritis is discussed.

  15. Oral glucose tolerance test effects on endothelial inflammation markers in healthy subjects and diabetic patients.

    PubMed

    Derosa, G; D'Angelo, A; Salvadeo, S A T; Ferrari, I; Fogari, E; Gravina, A; Mereu, R; Palumbo, I; Maffioli, P; Randazzo, S; Cicero, A F G

    2010-01-01

    The aim of this study was to evaluate the effect of an oral glucose tolerance test (OGTT) on the level of endothelial dysfunction and vascular inflammation markers in healthy subjects (H) and diabetic overweight patients (D). We enrolled 256 healthy subjects and 274 type 2 diabetic patients. We evaluated blood glucose (BG), soluble intercellular adhesion molecule-1 (sICAM-1), interleukin-6 (IL-6), high-sensitivity C reactive protein (hsCRP), soluble vascular cell adhesion molecule-1 (sVCAM-1), soluble E-selectin (sE-selectin), and tumor necrosis factor-alpha (TNF-alpha) at baseline and after OGTT. We observed that BG, sICAM-1, IL-6, hs-CRP, sVCAM-1, sE-selectin, and TNF-alpha values were higher in D group than in H group. In a large sample of adult healthy subjects and type 2 diabetics we observed that both answer to an OGTT with a significant increase in biomarkers of systemic low-grade inflammation and endothelial dysfunction such as hsCRP, IL-6, TNF-alpha, sICAM-1, sVCAM-1, and sE-selectin. Type 2 diabetics experienced, however, a more significant increase in TNF-alpha, and sE-selectin.

  16. Endothelial dysfunction, inflammation, and oxidative stress in obese children and adolescents: markers and effect of lifestyle intervention.

    PubMed

    Montero, D; Walther, G; Perez-Martin, A; Roche, E; Vinet, A

    2012-05-01

    With an increasing prevalence, pediatric obesity is often a prelude to adulthood obesity, and represents a major public health issue. Comorbidities are very common and severe in obese adults, justifying the search for earlier markers or risk factors for cardiovascular diseases in obese children. Endothelial dysfunction has been found to be present in the early stages of atherosclerosis, and can be non-invasively assessed with widely accepted and well-standardized techniques at the macrocirculation level. Endothelial dysfunction at the microcirculation level is less documented in obese children. Obesity in children has been repeatedly and independently correlated to endothelial dysfunction, inflammation and oxidative stress markers, although the relationship between these factors remains to be investigated. However, this would not only allow substantial improvements in risk stratification, but also provide essential data regarding the evolution of endothelial dysfunction in childhood obesity, especially during puberty when pro-inflammatory and pro-oxidative changes, with relative insulin resistance, occur. Therapeutic strategies such as lifestyle interventions in early childhood obesity appear all the more necessary, optimally including both exercise and diet because of their known effects on inflammatory and oxidative stress markers, potentially reversing endothelial dysfunction.

  17. Relationship between Inflammatory Markers, Endothelial Activation Markers, and Carotid Intima-Media Thickness in HIV-Infected Patients Receiving Antiretroviral Therapy

    PubMed Central

    Ross, Allison C.; Rizk, Nesrine; O'Riordan, Mary Ann; Dogra, Vikram; El-Bejjani, Dalia; Storer, Norma; Harrill, Danielle; Tungsiripat, Marisa; Adell, Jerome; McComsey, Grace A.

    2014-01-01

    Background Human immunodeficiency virus (HIV)–infected patients are at increased risk of cardiovascular disease, which may be related to chronic inflammation and endothelial dysfunction despite virological control with antiretroviral therapy. The relationship between carotid intima-media thickness (IMT), a surrogate marker for cardiovascular disease, proinflammatory cytokines, and endothelial activation markers has not been fully explored in HIV-infected patients who are receiving antiretroviral therapy. Methods We conducted a prospective, cross-sectional, observational study of treated HIV-infected patients and healthy control subjects to evaluate the relationship between carotid IMT, proinflammatory cytokines, endothelial activation biomarkers, and metabolic parameters in treated HIV-infected patients, compared with healthy control subjects. Results We enrolled 73 HIV-infected patients and 21 control subjects. Common carotid artery and internal carotid artery IMT measurements, as well as tumor necrosis factor–α, high-sensitivity C-reactive protein, inter-leukin-6, myeloperoxidase, and soluble vascular cell adhesion molecule-1 levels were higher in the HIV-infected group. High-sensitivity C-reactive protein was the only biomarker that was positively correlated with carotid IMT in both groups. In the HIV-infected group, soluble vascular cell adhesion molecule–1 was positively correlated with all inflammatory cytokine levels. In multiple regression analysis, soluble vascular cell adhesion molecule–1, myeloperoxidase, and tumor necrosis factor–α levels were all associated with internal carotid artery IMT in the HIV-infected group, whereas age was associated with both common carotid artery and internal carotid artery IMT. Conclusions Enhanced endothelial activation, inflammation, and increased carotid IMT occur in HIV-infected patients despite antiretroviral therapy. Inflammatory markers are associated with endothelial activation, and both are associated

  18. Interactions between autophagic and endo-lysosomal markers in endothelial cells.

    PubMed

    Oeste, Clara L; Seco, Esther; Patton, Wayne F; Boya, Patricia; Pérez-Sala, Dolores

    2013-05-01

    Autophagic and endo-lysosomal degradative pathways are essential for cell homeostasis. Availability of reliable tools to interrogate these pathways is critical to unveil their involvement in physiology and pathophysiology. Although several probes have been recently developed to monitor autophagic or lysosomal compartments, their specificity has not been validated through co-localization studies with well-known markers. Here, we evaluate the selectivity and interactions between one lysosomal (Lyso-ID) and one autophagosomal (Cyto-ID) probe under conditions modulating autophagy and/or endo-lysosomal function in live cells. The probe for acidic compartments Lyso-ID was fully localized inside vesicles positive for markers of late endosome-lysosomes, including Lamp1-GFP and GFP-CINCCKVL. Induction of autophagy by amino acid deprivation in bovine aortic endothelial cells caused an early and potent increase in the fluorescence of the proposed autophagy dye Cyto-ID. Cyto-ID-positive compartments extensively co-localized with the autophagosomal fluorescent reporter RFP-LC3, although the time and/or threshold for organelle detection was different for each probe. Interestingly, use of Cyto-ID in combination with Lysotracker Red or Lyso-ID allowed the observation of structures labeled with either one or both probes, the extent of co-localization increasing upon treatment with protease inhibitors. Inhibition of the endo-lysosomal pathway with chloroquine or U18666A resulted in the formation of large Cyto-ID and Lyso-ID-positive compartments. These results constitute the first assessment of the selectivity of Cyto-ID and Lyso-ID as probes for the autophagic and lysosomal pathways, respectively. Our observations show that these probes can be used in combination with protein-based markers for monitoring the interactions of both pathways in live cells.

  19. Citrus Polyphenol Hesperidin Stimulates Production of Nitric Oxide in Endothelial Cells while Improving Endothelial Function and Reducing Inflammatory Markers in Patients with Metabolic Syndrome

    PubMed Central

    Rizza, Stefano; Muniyappa, Ranganath; Iantorno, Micaela; Kim, Jeong-a; Chen, Hui; Pullikotil, Philomena; Senese, Nicoletta; Tesauro, Manfredi; Lauro, Davide; Cardillo, Carmine

    2011-01-01

    Context: Hesperidin, a citrus flavonoid, and its metabolite hesperetin may have vascular actions relevant to their health benefits. Molecular and physiological mechanisms of hesperetin actions are unknown. Objective: We tested whether hesperetin stimulates production of nitric oxide (NO) from vascular endothelium and evaluated endothelial function in subjects with metabolic syndrome on oral hesperidin therapy. Design, Setting, and Interventions: Cellular mechanisms of action of hesperetin were evaluated in bovine aortic endothelial cells (BAEC) in primary culture. A randomized, placebo-controlled, double-blind, crossover trial examined whether oral hesperidin administration (500 mg once daily for 3 wk) improves endothelial function in individuals with metabolic syndrome (n = 24). Main Outcome Measure: We measured the difference in brachial artery flow-mediated dilation between placebo and hesperidin treatment periods. Results: Treatment of BAEC with hesperetin acutely stimulated phosphorylation of Src, Akt, AMP kinase, and endothelial NO synthase to produce NO; this required generation of H2O2. Increased adhesion of monocytes to BAEC and expression of vascular cell adhesion molecule-1 in response to TNF-α treatment was reduced by pretreatment with hesperetin. In the clinical study, when compared with placebo, hesperidin treatment increased flow-mediated dilation (10.26 ± 1.19 vs. 7.78 ± 0.76%; P = 0.02) and reduced concentrations of circulating inflammatory biomarkers (high-sensitivity C-reactive protein, serum amyloid A protein, soluble E-selectin). Conclusions: Novel mechanisms for hesperetin action in endothelial cells inform effects of oral hesperidin treatment to improve endothelial dysfunction and reduce circulating markers of inflammation in our exploratory clinical trial. Hesperetin has vasculoprotective actions that may explain beneficial cardiovascular effects of citrus consumption. PMID:21346065

  20. Monoclonal Antibody against Angiotensin-Converting Enzyme: Its Use as a Marker for Murine, Bovine, and Human Endothelial Cells

    NASA Astrophysics Data System (ADS)

    Auerbach, R.; Alby, L.; Grieves, J.; Joseph, J.; Lindgren, C.; Morrissey, L. W.; Sidky, Y. A.; Tu, M.; Watt, S. L.

    1982-12-01

    A monoclonal antibody has been prepared against rat angiotensin-converting enzyme (ACE). By selection for antibody binding to endothelial cells of bovine rather than rat origin we have obtained a reagent that has broad cross-species binding properties and that can at the same time serve as a useful marker for the surface of endothelial cells. The IgM-producing clone that we have established, α -ACE 3.1.1, has been grown in ascites form to yield ascites fluid that binds selectively to immobilized ACE at a >1:10,000 dilution. By use of enzyme-linked immunosorbent assays, immunofluorescence histology, and flow cytometry, we have demonstrated the presence of ACE on endothelial cells of murine, bovine, and human origin. By means of a fluorescence-activated cell sorter (FACS-IV) we have been able to selectively isolate viable endothelial cells from a mixture of endothelial cells and fibroblasts. We believe the antibody will be useful not only for the selection and in vitro cultivation of endothelial cells but also as a tool for the identification and pharmacological study of ACE.

  1. Chronic exposure to biomass fuel smoke and markers of endothelial inflammation.

    PubMed

    Caravedo, M A; Herrera, P M; Mongilardi, N; de Ferrari, A; Davila-Roman, V G; Gilman, R H; Wise, R A; Miele, C H; Miranda, J J; Checkley, W

    2016-10-01

    Indoor smoke exposure may affect cardiovascular disease (CVD) risk via lung-mediated inflammation, oxidative stress, and endothelial inflammation. We sought to explore the association between indoor smoke exposure from burning biomass fuels and a selected group of markers for endothelial inflammation. We compared serum concentrations of amyloid A protein, E-selectin, soluble intercellular adhesion molecule 1 (ICAM-1) and VCAM-1, von Willebrand factor (vWF), and high-sensitivity C-reactive protein (hs-CRP) in 228 biomass-exposed vs. 228 non-exposed participants living in Puno, Peru. Average age was 56 years (s.d. = 13), average BMI was 26.5 kg/m(2) (s.d. = 4.4), 48% were male, 59.4% completed high school, and 2% reported a physician diagnosis of CVD. In unadjusted analysis, serum levels of soluble ICAM-1 (330 vs. 302 ng/ml; P < 0.001), soluble VCAM-1 (403 vs. 362 ng/ml; P < 0.001), and E-selectin (54.2 vs. 52.7 ng/ml; P = 0.05) were increased in biomass-exposed vs. non-exposed participants, respectively, whereas serum levels of vWF (1148 vs. 1311 mU/ml; P < 0.001) and hs-CRP (2.56 vs. 3.12 mg/l; P < 0.001) were decreased, respectively. In adjusted analyses, chronic exposure to biomass fuels remained positively associated with serum levels of soluble ICAM-1 (P = 0.03) and VCAM-1 (P = 0.05) and E-selectin (P = 0.05), and remained negatively associated with serum levels of vWF (P = 0.02) and hs-CRP (P < 0.001). Daily exposure to biomass fuel smoke was associated with important differences in specific biomarkers of endothelial inflammation and may help explain accelerated atherosclerosis among those who are chronically exposed.

  2. Generation and Characterization of Telomerase-Transfected Human Lymphatic Endothelial Cells with an Extended Life Span

    PubMed Central

    Nisato, Riccardo E.; Harrison, Jillian A.; Buser, Raphaele; Orci, Lelio; Rinsch, Chris; Montesano, Roberto; Dupraz, Philippe; Pepper, Michael S.

    2004-01-01

    The study of lymphatic endothelial cells and lymphangiogenesis has, in the past, been hampered by the lack of lymphatic endothelial-specific markers. The recent discovery of several such markers has permitted the isolation of lymphatic endothelial cells (LECs) from human skin. However, cell numbers are limited and purity is variable with the different isolation procedures. To overcome these problems, we have transfected human dermal microvascular endothelial cells (HDMVECs) with a retrovirus containing the coding region of human telomerase reverse transcriptase (hTERT), and have produced a cell line, hTERT-HDLEC, with an extended lifespan. hTERT-HDLEC exhibit a typical cobblestone morphology when grown in culture, are contact-inhibited, and express endothelial cell-specific markers. hTERT-HDLEC also express the recognized lymphatic markers, Prox-1, LYVE-1 and podoplanin, as well as integrin α9, but do not express CD34. They also form tube-like structures in three-dimensional collagen gels when stimulated with vascular endothelial growth factors -A and -C. Based on these currently recognized criteria, these cells are LEC. Surprisingly, we also found that the widely studied HMEC-1 cell line expresses recognized lymphatic markers; however, these cells are also CD34-positive. In summary, the ectopic expression of hTERT increases the life span of LECs and does not affect their capacity to form tube-like structures in a collagen matrix. The production and characterization of hTERT-HDLEC will facilitate the study of the properties of lymphatic endothelium in vitro. PMID:15215158

  3. Endosomal recycling regulates Anthrax Toxin Receptor 1/Tumor Endothelial Marker 8-dependent cell spreading.

    PubMed

    Gu, Jingsheng; Faundez, Victor; Werner, Erica

    2010-07-15

    Mechanisms for receptor-mediated anthrax toxin internalization and delivery to the cytosol are well understood. However, far less is known about the fate followed by anthrax toxin receptors prior and after cell exposure to the toxin. We report that Anthrax Toxin Receptor 1/Tumor Endothelial Marker 8 (TEM8) localized at steady state in Rab11a-positive and transferrin receptor-containing recycling endosomes. TEM8 followed a slow constitutive recycling route of approximately 30min as determined by pulsed surface biotinylation and chase experiments. A Rab11a dominant negative mutant and Myosin Vb tail expression impaired TEM8 recycling by sequestering TEM8 in intracellular compartments. Sequestration of TEM8 in intracellular compartments with monensin coincided with increased TEM8 association with a multi-protein complex isolated with antibodies against transferrin receptor. Addition of the cell-binding component of anthrax toxin, Protective Antigen, reduced TEM8 half-life from 7 to 3 hours, without preventing receptor recycling. Pharmacological and molecular perturbation of recycling endosome function using monensin, dominant negative Rab11a, or myosin Vb tail, reduced PA binding efficiency and TEM8-dependent cell spreading on PA-coated surfaces without affecting toxin delivery to the cytosol. These results indicate that the intracellular fate of TEM8 differentially affect its cell adhesion and cell intoxication functions.

  4. Markers of endothelial dysfunction and evaluation of vascular reactivity tests in Behçet disease.

    PubMed

    Ozuguz, Pinar; Karabulut, Ayse Anil; Tulmac, Murat; Kisa, Ucler; Kocak, Mukadder; Gunduz, Ozgur

    2014-11-01

    We assessed endothelial dysfunction (ED) in patients with Behcet disease (BD; n=40) and healthy controls (n=20). Serum lipid, homocysteine, asymmetric dimethylarginine (ADMA) and high-sensitivity C-reactive protein (hsCRP) levels, erythrocyte sedimentation rates (ESRs), and ultrasonographic flow-mediated dilatation (FMD) were measured. Mean hsCRP, ESR, homocysteine, and ADMA were significantly higher in the BD group (P<.001 for all). Patients with active BD had higher serum levels of hsCRP, homocysteine, and ESR compared with those in remission (P<.001, P<.001, and P=.005, respectively). Flow-mediated dilatation was significantly lower in patients with BD than in controls (P=.001). Flow-mediated dilatation correlated negatively with BD duration and serum ADMA levels (P<.001, r=-.745 and P<.001, r=-.682); a positive correlation was seen between serum ADMA levels and BD duration (P<.001, r=.552). Only stepwise multivariate regression analysis revealed BD duration to have a significant effect on FMD. Flow-mediated dilatation, in conjunction with markers of inflammation, may evaluate ED in patients with BD.

  5. Markers of endothelial damage in patients with chronic kidney disease on hemodialysis.

    PubMed

    Carmona, Andrés; Agüera, Maria L; Luna-Ruiz, Carlos; Buendía, Paula; Calleros, Laura; García-Jerez, Andrea; Rodríguez-Puyol, Manuel; Arias, Manuel; Arias-Guillen, Marta; de Arriba, Gabriel; Ballarin, Jose; Bernis, Carmen; Fernández, Elvira; García-Rebollo, Sagrario; Mancha, Javier; Del Peso, Gloria; Pérez, Estefanía; Poch, Esteban; Portolés, Jose M; Rodríguez-Puyol, Diego; Sánchez-Villanueva, Rafael; Sarro, Felipe; Torres, Armando; Martín-Malo, Alejandro; Aljama, Pedro; Ramírez, Rafael; Carracedo, Julia

    2017-04-01

    Patients with Stage 5 chronic kidney disease who are on hemodialysis (HD) remain in a chronic inflammatory state, characterized by the accumulation of uremic toxins that induce endothelial damage and cardiovascular disease (CVD). Our aim was to examine microvesicles (MVs), monocyte subpopulations, and angiopoietins (Ang) to identify prognostic markers in HD patients with or without diabetes mellitus (DM). A total of 160 prevalent HD patients from 10 centers across Spain were obtained from the Biobank of the Nephrology Renal Network (Madrid, Spain): 80 patients with DM and 80 patients without DM who were matched for clinical and demographic criteria. MVs from plasma and several monocyte subpopulations (CD14(2+)/CD16(+), CD14(+)/CD16(2+)) were analyzed by flow cytometry, and the plasma concentrations of Ang1 and Ang2 were quantified by ELISA. Data on CVD were gathered over the 5.5 yr after these samples were obtained. MV level, monocyte subpopulations (CD14(+)/CD16(2+) and CD14(2+)/CD16(+)), and Ang2-to-Ang1 ratios increased in HD patients with DM compared with non-DM patients. Moreover, MV level above the median (264 MVs/µl) was associated independently with greater mortality. MVs, monocyte subpopulations, and Ang2-to-Ang1 ratio can be used as predictors for CVD. In addition, MV level has a potential predictive value in the prevention of CVD in HD patients. These parameters undergo more extensive changes in patients with DM.

  6. The brain-tumor related protein podoplanin regulates synaptic plasticity and hippocampus-dependent learning and memory

    PubMed Central

    Cicvaric, Ana; Yang, Jiaye; Krieger, Sigurd; Khan, Deeba; Kim, Eun-Jung; Dominguez-Rodriguez, Manuel; Cabatic, Maureen; Molz, Barbara; Acevedo Aguilar, Juan Pablo; Milicevic, Radoslav; Smani, Tarik; Breuss, Johannes M.; Kerjaschki, Dontscho; Pollak, Daniela D.; Uhrin, Pavel; Monje, Francisco J.

    2016-01-01

    Abstract Introduction: Podoplanin is a cell-surface glycoprotein constitutively expressed in the brain and implicated in human brain tumorigenesis. The intrinsic function of podoplanin in brain neurons remains however uncharacterized. Materials and methods: Using an established podoplanin-knockout mouse model and electrophysiological, biochemical, and behavioral approaches, we investigated the brain neuronal role of podoplanin. Results: Ex-vivo electrophysiology showed that podoplanin deletion impairs dentate gyrus synaptic strengthening. In vivo, podoplanin deletion selectively impaired hippocampus-dependent spatial learning and memory without affecting amygdala-dependent cued fear conditioning. In vitro, neuronal overexpression of podoplanin promoted synaptic activity and neuritic outgrowth whereas podoplanin-deficient neurons exhibited stunted outgrowth and lower levels of p-Ezrin, TrkA, and CREB in response to nerve growth factor (NGF). Surface Plasmon Resonance data further indicated a physical interaction between podoplanin and NGF. Discussion: This work proposes podoplanin as a novel component of the neuronal machinery underlying neuritogenesis, synaptic plasticity, and hippocampus-dependent memory functions. The existence of a relevant cross-talk between podoplanin and the NGF/TrkA signaling pathway is also for the first time proposed here, thus providing a novel molecular complex as a target for future multidisciplinary studies of the brain function in the physiology and the pathology.Key messagesPodoplanin, a protein linked to the promotion of human brain tumors, is required in vivo for proper hippocampus-dependent learning and memory functions.Deletion of podoplanin selectively impairs activity-dependent synaptic strengthening at the neurogenic dentate-gyrus and hampers neuritogenesis and phospho Ezrin, TrkA and CREB protein levels upon NGF stimulation.Surface plasmon resonance data indicates a physical interaction between podoplanin and NGF. On these

  7. Soya isoflavone-enriched cereal bars affect markers of endothelial function in postmenopausal women.

    PubMed

    Hallund, J; Bügel, S; Tholstrup, T; Ferrari, M; Talbot, D; Hall, W L; Reimann, M; Williams, C M; Wiinberg, N

    2006-06-01

    Soya isoflavones are thought to be cardioprotective due to their structural similarity to oestrogen. In order to investigate the effect of soya isoflavones on markers of endothelial function we conducted a randomised, double-blind, placebo-controlled, cross-over study with thirty healthy postmenopausal women. The women consumed cereal bars, with or without soya isoflavones (50 mg/d), for 8 weeks, separated by an 8-week washout period. Systemic arterial compliance (SAC), isobaric arterial compliance (IAC), flow-mediated endothelium-dependent vasodilation (FMD) and nitroglycerine-mediated endothelium-independent vasodilation (NMD) were measured at the beginning of the study and after each intervention period. Blood pressure (BP) and plasma concentrations of nitrite and nitrate (NOx) and endothelin-1 (ET-1) were measured at the beginning and end of each intervention period. NMD was 13.4 (SEM 2.0)% at baseline and 15.5 (SEM 1.1) % after isoflavone treatment compared with 12.4 (SEM 1.0)% after placebo treatment (P=0.03). NOx increased from 27.7 (SEM 2.7) to 31.1 (SEM 3.2) microM after isoflavones treatment compared with 25.4 (SEM 1.5) to 20.4 (SEM 1.1) microM after placebo treatment (P=0.003) and a significant increase in the NOx:ET-1 ratio (P=0.005) was observed after the isoflavone treatment compared with placebo. A significant difference in SAC after the isoflavone and placebo treatment was observed (P=0.04). No significant difference was found in FMD, IAC, BP and ET-1. In conclusion, 8 weeks' consumption of cereals bars enriched with 50 mg soya isoflavones/d increased plasma NOx concentrations and improved endothelium-independent vasodilation in healthy postmenopausal women.

  8. Effect of repetitive SCUBA diving on humoral markers of endothelial and central nervous system integrity.

    PubMed

    Bilopavlovic, Nada; Marinovic, Jasna; Ljubkovic, Marko; Obad, Ante; Zanchi, Jaksa; Pollock, Neal W; Denoble, Petar; Dujic, Zeljko

    2013-07-01

    During SCUBA diving decompression, there is a significant gas bubble production in systemic veins, with rather frequent bubble crossover to arterial side even in asymptomatic divers. The aim of the current study was to investigate potential changes in humoral markers of endothelial and brain damage (endothelin-1, neuron-specific enolase and S-100β) after repetitive SCUBA diving with concomitant assessment of venous gas bubble production and subsequent arterialization. Sixteen male divers performed four open-water no-decompression dives to 18 msw (meters of sea water) lasting 49 min in consecutive days during which they performed moderate-level exercise. Before and after dives 1 and 4 blood was drawn, and bubble production and potential arterialization were echocardiographically evaluated. In addition, a control dive to 5 msw was performed with same duration, water temperature and exercise load. SCUBA diving to 18 msw caused significant bubble production with arterializations in six divers after dive 1 and in four divers after dive 4. Blood levels of endothelin-1 and neuron-specific enolase did not change after diving, but levels of S-100β were significantly elevated after both dives to 18 msw and a control dive. Creatine kinase activity following a control dive was also significantly increased. Although serum S-100β levels were increased after diving, concomitant increase of creatine kinase during control, almost bubble-free, dive suggests the extracranial release of S-100β, most likely from skeletal muscles. Therefore, despite the significant bubble production and sporadic arterialization after open-water dives to 18 msw, the current study found no signs of damage to neurons or the blood-brain barrier.

  9. Quantitative analysis of lymphangiogenic markers in human colorectal cancer.

    PubMed

    Parr, C; Jiang, W G

    2003-08-01

    Lymphatic spread of colorectal cancer cells to regional lymph nodes is one of the early events in metastatic cancer, and is often associated with distant metastatic spread and a poor prognosis. This study examined lymphangiogenic factors, and in particular a panel of newly discovered lymphangiogenic markers, in colorectal cancer tissues from a cohort of patients. Paired samples (background normal mucosa and cancer) of colon tissue were obtained from patients with colorectal cancer. The expression and levels of the VEGF-C and VEGF-D cytokines, the VEGF receptors VEGFR-2 and VEGFR-3, and newly described lymphatic endothelial markers, LYVE-1, Prox-1, podoplanin and 5'-nucleotidase were assessed. RNA was extracted from the frozen colon tissues. The level of expression for each factor/marker was determined using RT-PCR and quantified using a real-time quantitative PCR (RT-QPCR) technique, with respective cloned cDNA plasmids as internal standards. VEGF-D was expressed to a significantly higher degree in the colon tumour tissues. There was no significant difference between the expression levels for both VEGF-C and its receptor, VEGFR-2, in background and cancer tissues. However, levels of the VEGFR-3 receptor were found to be significantly higher in colon cancer than the normal background tissues. LYVE-1 levels were below detection in most cases. There was a significant increase in the degree of Prox-1 and 5'-nucleotidase expression in colon cancer tissue. Podoplanin expression was also increased in the cancer samples. These markers indicate an increase in lymphangiogenesis in colon cancer, and may therefore have prognostic value for colon cancer patients.

  10. Lycopersicon esculentum lectin: an effective and versatile endothelial marker of normal and tumoral blood vessels in the central nervous system.

    PubMed

    Mazzetti, S; Frigerio, S; Gelati, M; Salmaggi, A; Vitellaro-Zuccarello, L

    2004-01-01

    The binding of Lycopersicon esculentum lectin (LEA) to the vascular endothelium was studied in the central nervous system of rat, mouse and guinea pig at different developmental ages, and in a gliosarcoma model. Our observations showed that LEA consistently stained the entire vascular tree in the spinal cord and in the brain of all animal species at all developmental ages investigated. In the tumor model, the staining of the vascular network was very reproducible, enabled an easy identification of vascular profiles and displayed a higher efficiency when compared to two other commonly used vascular marker (EHS laminin and PECAM-1). Moreover, our results showed that LEA staining was comparable in both vibratome and paraffin sections and could be easily combined with other markers in double labeling experiments. These observations indicate that LEA staining may represent an effective and versatile endothelial marker for the study of the vasculature of the central nervous system in different animal species and experimental conditions.

  11. ALDH1 and podoplanin expression patterns predict the risk of malignant transformation in oral leukoplakia

    PubMed Central

    Habiba, Umma; Hida, Kyoko; Kitamura, Tetsuya; Matsuda, Aya Yanagawa; Higashino, Fumihiro; Ito, Yoichi M.; Ohiro, Yoichi; Totsuka, Yasunori; Shindoh, Masanobu

    2017-01-01

    Oral leukoplakia (OL) is a clinically diagnosed preneoplastic lesion of the oral cavity with an increased oral cancer risk. However, the risk of malignant transformation is still difficult to assess. The objective of the present study was to examine the expression patterns of aldehyde dehydrogenase 1 (ALDH1) and podoplanin in OL, and to determine their roles in predicting oral cancer development. In the present study, the expression patterns of ALDH1 and podoplanin were determined in samples from 79 patients with OL. The association between protein expression and clinicopathological parameters, including oral cancer-free survival, was analyzed during a mean follow-up period of 3.4 years. Expression of ALDH1 and podoplanin was observed in 61 and 67% patients, respectively. Kaplan-Meier analysis demonstrated that the expression of the proteins was correlated with the risk of progression to oral cancer. Multivariate analysis revealed that expression of ALDH1 and podoplanin was associated with 3.02- and 2.62-fold increased risk of malignant transformation, respectively. The malignant transformation risk of OL was considerably higher in cases with expression of both proteins. Point-prevalence analysis revealed that 66% of patients with co-expression of ALDH1 and podoplanin developed oral cancer. Taken together, our data indicate that ALDH1 and podoplanin expression patterns in OL are associated with oral cancer development, suggesting that ALDH1 and podoplanin may be useful biomarkers to identify OL patients with a substantially high oral cancer risk. PMID:28123562

  12. ALDH1 and podoplanin expression patterns predict the risk of malignant transformation in oral leukoplakia.

    PubMed

    Habiba, Umma; Hida, Kyoko; Kitamura, Tetsuya; Matsuda, Aya Yanagawa; Higashino, Fumihiro; Ito, Yoichi M; Ohiro, Yoichi; Totsuka, Yasunori; Shindoh, Masanobu

    2017-01-01

    Oral leukoplakia (OL) is a clinically diagnosed preneoplastic lesion of the oral cavity with an increased oral cancer risk. However, the risk of malignant transformation is still difficult to assess. The objective of the present study was to examine the expression patterns of aldehyde dehydrogenase 1 (ALDH1) and podoplanin in OL, and to determine their roles in predicting oral cancer development. In the present study, the expression patterns of ALDH1 and podoplanin were determined in samples from 79 patients with OL. The association between protein expression and clinicopathological parameters, including oral cancer-free survival, was analyzed during a mean follow-up period of 3.4 years. Expression of ALDH1 and podoplanin was observed in 61 and 67% patients, respectively. Kaplan-Meier analysis demonstrated that the expression of the proteins was correlated with the risk of progression to oral cancer. Multivariate analysis revealed that expression of ALDH1 and podoplanin was associated with 3.02- and 2.62-fold increased risk of malignant transformation, respectively. The malignant transformation risk of OL was considerably higher in cases with expression of both proteins. Point-prevalence analysis revealed that 66% of patients with co-expression of ALDH1 and podoplanin developed oral cancer. Taken together, our data indicate that ALDH1 and podoplanin expression patterns in OL are associated with oral cancer development, suggesting that ALDH1 and podoplanin may be useful biomarkers to identify OL patients with a substantially high oral cancer risk.

  13. Podoplanin mediates ECM degradation by squamous carcinoma cells through control of invadopodia stability

    PubMed Central

    Martín-Villar, E; Borda-d'Agua, B; Carrasco-Ramirez, P; Renart, J; Parsons, M; Quintanilla, M; Jones, G E

    2015-01-01

    Invadopodia are actin-rich cell membrane projections used by invasive cells to penetrate the basement membrane. Control of invadopodia stability is critical for efficient degradation of the extracellular matrix (ECM); however, the underlying molecular mechanisms remain poorly understood. Here, we uncover a new role for podoplanin, a transmembrane glycoprotein closely associated with malignant progression of squamous cell carcinomas (SCCs), in the regulation of invadopodia-mediated matrix degradation. Podoplanin downregulation in SCC cells impairs invadopodia stability, thereby reducing the efficiency of ECM degradation. We report podoplanin as a novel component of invadopodia-associated adhesion rings, where it clusters prior to matrix degradation. Early podoplanin recruitment to invadopodia is dependent on lipid rafts, whereas ezrin/moesin proteins mediate podoplanin ring assembly. Finally, we demonstrate that podoplanin regulates invadopodia maturation by acting upstream of the ROCK-LIMK-Cofilin pathway through the control of RhoC GTPase activity. Thus, podoplanin has a key role in the regulation of invadopodia function in SCC cells, controlling the initial steps of cancer cell invasion. PMID:25486435

  14. LpMab-23: A Cancer-Specific Monoclonal Antibody Against Human Podoplanin.

    PubMed

    Yamada, Shinji; Ogasawara, Satoshi; Kaneko, Mika K; Kato, Yukinari

    2017-04-07

    Human podoplanin (hPDPN), the ligand of C-type lectin-like receptor-2, is involved in cancer metastasis. Until now, many monoclonal antibodies (mAbs) have been established against hPDPN. However, it is still difficult to develop a cancer-specific mAb (CasMab) against hPDPN because the protein sequence of hPDPN expressed in cancer cells is the same as that in normal cells. Herein, we report LpMab-23 of the mouse IgG1 subclass, a novel CasMab against hPDPN. In an immunohistochemical analysis, LpMab-23 reacted with tumor cells of human oral cancer, but did not react with normal cells such as lymphatic endothelial cells (LECs). In contrast, LpMab-17, another anti-hPDPN mAb, reacted with both tumor cells and LECs. Furthermore, flow cytometric analysis revealed that LpMab-23 reacted with hPDPN-expressing cancer cell lines (LN319, RERF-LC-AI/hPDPN, Y-MESO-14/hPDPN, and HSC3/hPDPN) but showed little reaction with normal cells (LECs and HEK-293T), although another anti-hPDPN mAb, LpMab-7, reacted with both hPDPN-expressing cancer cells and normal cells, indicating that LpMab-23 is a CasMab against hPDPN.

  15. Effects of a Physical Activity Program on Markers of Endothelial Dysfunction, Oxidative Stress, and Metabolic Status in Adolescents with Metabolic Syndrome

    PubMed Central

    Camarillo-Romero, Eneida; Dominguez-Garcia, Ma Victoria; Amaya-Chavez, Araceli; Camarillo-Romero, Maria del Socorro; Talavera-Piña, Juan; Huitron-Bravo, Gerardo; Majluf-Cruz, Abraham

    2012-01-01

    The metabolic syndrome (MetS) is a precursor of diabetes. Physical activity (PA) improves endothelial dysfunction and may benefit patients with MetS. Aims. To evaluate the effect of a physical activity (PA) program on markers of endothelial dysfunction and oxidative stress in adolescents with (MetS). Methods. We carried out a cohort study of 38 adolescents with and without MetS (18 females and 20 males). All participants completed a 3-month PA program. All variables of the MetS as well as markers of endothelial dysfunction and oxidative stress tests were evaluated. Results. Females with and without MetS showed significant differences for almost all components of the MetS, whereas males were significantly different in half of the components. After the PA program, components of the MetS were not different from baseline values except for HDL-C levels. Some baseline endothelial dysfunction markers were significantly different among adolescents with and without MetS; however, after the PA program, most of these markers significantly improved in subjects with and without MetS. Conclusion. PA improves the markers of endothelial dysfunction in adolescents with MetS although other changes in the components of the MetS were not observed. Perhaps the benefits of PA on all components of MetS would appear after a PA program with a longer duration. PMID:22888450

  16. Connection between telomerase activity in PBMC and markers of inflammation and endothelial dysfunction in patients with metabolic syndrome.

    PubMed

    Rentoukas, Elias; Tsarouhas, Konstantinos; Kaplanis, Ioannis; Korou, Eleni; Nikolaou, Maria; Marathonitis, George; Kokkinou, Stavroula; Haliassos, Alexander; Mamalaki, Avgi; Kouretas, Demetrios; Tsitsimpikou, Christina

    2012-01-01

    Metabolic syndrome (MS) is a constellation of metabolic derangements associated with vascular endothelial dysfunction and oxidative stress and is widely regarded as an inflammatory condition, accompanied by an increased risk for cardiovascular disease. The present study tried to investigate the implications of telomerase activity with inflammation and impaired endothelial function in patients with metabolic syndrome. Telomerase activity in circulating peripheral blood mononuclear cells (PBMC), TNF-α, IL-6 and ADMA were monitored in 39 patients with MS and 20 age and sex-matched healthy volunteers. Telomerase activity in PBMC, TNF-α, IL-6 and ADMA were all significantly elevated in patients with MS compared to healthy volunteers. PBMC telomerase was negatively correlated with HDL and positively correlated with ADMA, while no association between TNF-α and IL-6 was observed. IL-6 was increasing with increasing systolic pressure both in the patients with MS and in the healthy volunteers, while smoking and diabetes were positively correlated with IL-6 only in the patients' group. In conclusion, in patients with MS characterised by a strong dyslipidemic profile and low diabetes prevalence, significant telomerase activity was detected in circulating PBMC, along with elevated markers of inflammation and endothelial dysfunction. These findings suggest a prolonged activity of inflammatory cells in the studied state of this metabolic disorder that could represent a contributory pathway in the pathogenesis of atherosclerosis.

  17. Effects of Olive Oil on Markers of Inflammation and Endothelial Function—A Systematic Review and Meta-Analysis

    PubMed Central

    Schwingshackl, Lukas; Christoph, Marina; Hoffmann, Georg

    2015-01-01

    The aim of the present systematic review was to synthesize data from randomized controlled trials investigating the effects of olive oil on markers of inflammation or endothelial function. Literature search in electronic databases Cochrane Trial Register, EMBASE, and MEDLINE was performed. Thirty studies enrolling 3106 participants fulfilled the selection criteria. Pooled effects of different interventions were assessed as mean difference using a random effects model. Olive oil interventions (with daily consumption ranging approximately between 1 mg and 50 mg) resulted in a significantly more pronounced decrease in C-reactive protein (mean difference: −0.64 mg/L, (95% confidence interval (CI) −0.96 to −0.31), p < 0.0001, n = 15 trials) and interleukin-6 (mean difference: −0.29 (95% CI −0.7 to −0.02), p < 0.04, n = 7 trials) as compared to controls, respectively. Values of flow-mediated dilatation (given as absolute percentage) were significantly more increased in individuals subjected to olive oil interventions (mean difference: 0.76% (95% CI 0.27 to 1.24), p < 0.002, n = 8 trials). These results provide evidence that olive oil might exert beneficial effects on endothelial function as well as markers of inflammation and endothelial function, thus representing a key ingredient contributing to the cardiovascular-protective effects of a Mediterranean diet. However, due to the heterogeneous study designs (e.g., olive oil given as a supplement or as part of dietary pattern, variations in control diets), a conservative interpretation of the results is necessary. PMID:26378571

  18. Effects of Olive Oil on Markers of Inflammation and Endothelial Function-A Systematic Review and Meta-Analysis.

    PubMed

    Schwingshackl, Lukas; Christoph, Marina; Hoffmann, Georg

    2015-09-11

    The aim of the present systematic review was to synthesize data from randomized controlled trials investigating the effects of olive oil on markers of inflammation or endothelial function. Literature search in electronic databases Cochrane Trial Register, EMBASE, and MEDLINE was performed. Thirty studies enrolling 3106 participants fulfilled the selection criteria. Pooled effects of different interventions were assessed as mean difference using a random effects model. Olive oil interventions (with daily consumption ranging approximately between 1 mg and 50 mg) resulted in a significantly more pronounced decrease in C-reactive protein (mean difference: -0.64 mg/L, (95% confidence interval (CI) -0.96 to -0.31), p < 0.0001, n = 15 trials) and interleukin-6 (mean difference: -0.29 (95% CI -0.7 to -0.02), p < 0.04, n = 7 trials) as compared to controls, respectively. Values of flow-mediated dilatation (given as absolute percentage) were significantly more increased in individuals subjected to olive oil interventions (mean difference: 0.76% (95% CI 0.27 to 1.24), p < 0.002, n = 8 trials). These results provide evidence that olive oil might exert beneficial effects on endothelial function as well as markers of inflammation and endothelial function, thus representing a key ingredient contributing to the cardiovascular-protective effects of a Mediterranean diet. However, due to the heterogeneous study designs (e.g., olive oil given as a supplement or as part of dietary pattern, variations in control diets), a conservative interpretation of the results is necessary.

  19. Carotid Intima Media Thickness, Inflammatory Markers, and Endothelial Activation Markers in HIV Patients with Lipoatrophy Increased at 48 Weeks Regardless of Use of Rosiglitazone or Placebo

    PubMed Central

    Tungsiripat, Marisa; El-Bejjani, Dalia; Rizk, Nesrine; Dogra, Vikram; O'Riordan, Mary Ann; Ross, Allison C.; Hileman, Corrilynn; Storer, Norma; Harrill, Danielle

    2011-01-01

    Abstract Rosiglitazone may be useful for the treatment of antiretroviral therapy-associated lipoatrophy, but an association with cardiovascular disease (CVD) has been questioned in diabetics. We evaluated rosiglitazone's effect on surrogate markers of CVD in HIV-infected individuals with lipoatrophy. HIV+ patients with lipoatrophy on thymidine-sparing regimens were randomized to rosiglitazone vs. placebo for 48 weeks. We serially assessed carotid IMT, fasting metabolic profiles, tumor necrosis factor (TNF)-α, soluble receptors (sTNFRI and II), interleukin (IL)-6, high-sensitivity C-reactive protein (hsCRP), myeloperoxidase (MPO), and endothelial activation markers [von Willebrand factor (vWF), soluble intercellular cell adhesion molecules-1 (sICAM-1), and vascular cell adhesion molecules-1 (sVCAM-1)]. Seventy-one subjects enrolled: 17% were female and 51%were white. Baseline characteristics were similar between groups except for higher total cholesterol in the placebo group (p = 0.04). At 48 weeks, common carotid artery (CCA) IMT changed significantly (p ≤ 0.05) within but not between the groups (p = 0.36): the median (IQR) increase was 0.10 (0.05, 0.25) mm and 0.15 (0, 0.25) mm in the rosiglitazone and placebo groups, respectively. hsCRP, sTNFRI and II, sVCAM-1, and vWF changed significantly (p ≤ 0.02) within but not between groups. Total cholesterol increased significantly in the rosiglitazone group (p = 0.008). In our study of virologically controlled subjects with lipoatrophy, rosiglitazone did not independently increase carotid IMT, endothelial activation, and inflammatory cytokines. PMID:20969457

  20. The effects of interleukin-7 on the lymphangiogenic properties of human endothelial cells.

    PubMed

    Al-Rawi, Mahir A A; Watkins, Gareth; Mansel, Robert E; Jiang, Wen G

    2005-09-01

    Lymphangiogenesis (growth of new lymphatic vessels) is thought to play an important role in cancer lymphatic spread to the regional lymph nodes. However, the molecular pathways involved in lymphangiogenesis and their regulation are still unclear. Recently, there has been a significant advance in the studies of the lymphatic system and lymphangiogenesis as several novel specific lymphatic markers are discovered. Here, the effects of several cytokines on the lymphatic expression of human endothelial cells were studied. Amongst these cytokines, interleukin-7 (IL-7) was found to have significant impact on the lymphatic expression as it induced the expression of podoplanin, prox-1 and LYVE-1 in endothelial cells. Furthermore, IL-7 enhanced endothelial cell growth, migration and generation of lymphatic tubules in vitro via upregulating the expression of the lymphangiogenic growth factor and vascular endothelial growth factor-D. The specificity of these effects of IL-7 was confirmed using blocking anti-bodies and ribozyme transgene technology. These effects of IL-7 were totally abolished when IL-7R null endothelial cell mutants were used. IL-7 activated its transmembrane receptor, IL-7R, on endothelial cells, as well as its downstream signalling intermediates, Jak-1, Jak-3, PI3-K and Stat-5. Selective inhibition of these intermediates using specific inhibitors showed that IL-7 induced the afore-mentioned effects via a Wortmannin sensitive pathway. Collectively, these results demonstrate, for the first time, that IL-7 is a lymphangiogenic growth factor by inducing the lymphangiogenic properties of endothelial cells. This might have a significant impact on the lymphatic spread of solid tumours. Furthermore, interruption of IL-7 signalling might provide an attractive therapeutic option in cancer lymphatic metastasis.

  1. Down-regulation of tumor endothelial marker 8 suppresses cell proliferation mediated by ERK1/2 activity

    PubMed Central

    Cao, Chuangjie; Wang, Zhuo; Huang, Leilei; Bai, Lihong; Wang, Yuefeng; Liang, Yingjie; Dou, Chengyun; Wang, Liantang

    2016-01-01

    Tumor endothelial marker 8 (TEM8) was recently suggested as a putative anti-tumor target in several types of human cancer based on its selective overexpression in tumor versus normal endothelial cells. The objective of this study was to detect the potential functions of TEM8 in osteosarcoma. Overall, TEM8 was mainly located in cytoplasm and was up-regulated in osteosarcoma compared to benign bone lesions and adjacent non tumor tissue (ANT). High TEM8 expression group had a significant lower overall survival rate than that in the low TEM8 expression group. TEM8 knock-down by siRNA or shRNA results in significant reduction of osteosarcoma cell growth and proliferation both in vitro and in vivo. Ablation of TEM8 led to increasing of p21 and p27 and suppression of cyclin D1 mediated by Erk1/2 activity. These findings suggest that down-regulation of TEM8 play an important role in the inhibition of tumorigenesis and development of osteosarcoma. PMID:26996335

  2. Plasma angiopoietin-2 outperforms other markers of endothelial injury in prognosticating pediatric ARDS mortality.

    PubMed

    Zinter, Matt S; Spicer, Aaron; Orwoll, Benjamin O; Alkhouli, Mustafa; Dvorak, Christopher C; Calfee, Carolyn S; Matthay, Michael A; Sapru, Anil

    2016-02-01

    Angiopoietin-2 (Ang-2) is a key mediator of pulmonary vascular permeability. This study tested the association between plasma Ang-2 and mortality in pediatric acute respiratory distress syndrome (ARDS), with stratification for prior hematopoietic cellular transplantation (HCT), given the severe, yet poorly understood, ARDS phenotype of this subgroup. We enrolled 259 children <18 years of age with ARDS; 25 had prior HCT. Plasma Ang-2, von Willebrand Factor antigen (vWF), and vascular endothelial growth factor (VEGF) were measured on ARDS days 1 and 3 and correlated with patient outcomes. Day 1 and day 3 Ang-2 levels were associated with mortality independent of age, sex, race, and P/F ratio [odds ratio (OR) 3.7, 95% CI 1.1-11.5, P = 0.027; and OR 10.2, 95% confidence interval (CI) 2.2-46.5, P = 0.003, for each log10 increase in Ang-2]. vWF was associated with mortality (P = 0.027), but VEGF was not. The association between day 1 Ang-2 and mortality was independent of levels of both vWF and VEGF (OR 3.6, 95% CI 1.1-12.1, P = 0.039, for each log10 increase in Ang-2). 45% of the cohort had a rising Ang-2 between ARDS day 1 and 3 (adjusted mortality OR 3.3, 95% CI 1.2-9.2, P = 0.026). HCT patients with a rising Ang-2 had 70% mortality compared with 13% mortality for those without (OR 16.3, 95% CI 1.3-197.8, P = 0.028). Elevated plasma levels of Ang-2 were associated with mortality independent of vWF and VEGF. A rising Ang-2 between days 1 and 3 was strongly associated with mortality, particularly in pediatric HCT patients, suggesting vulnerability to ongoing endothelial damage.

  3. Early circulating levels of endothelial cell activation markers in aneurysmal subarachnoid haemorrhage: associations with cerebral ischaemic events and outcome

    PubMed Central

    Frijns, C J M; Fijnheer, R; Algra, A; van Mourik, J A; van Gijn, J; Rinkel, G J E

    2006-01-01

    Objective To investigate the relation of endothelial cell activation with delayed cerebral ischaemia (DCI) and outcome after subarachnoid haemorrhage (SAH). Methods Concentrations of soluble (s) intercellular adhesion molecule‐1, sE‐selectin, sP‐selectin, ED1‐fibronectin, von Willebrand Factor (vWf), and vWf propeptide were measured within three days of SAH onset. The associations with poor outcome were investigated at three months in 106 patients. In 90 patients in whom the occurrence of cerebral ischaemia could be dated accurately, two analyses were undertaken: one for all ischaemic events (n = 32), including those related to treatment, and another for spontaneous DCI (n = 11). Concentrations of markers were dichotomised at their medians. The associations of endothelial cell activation markers with outcome were expressed as odds ratios (OR) from logistic regression and those with ischaemic events as hazard ratios (HR) derived from Cox regression. Results Early vWf concentrations were associated with poor outcome (crude OR = 4.6 (95% CI, 2.0 to 10.9; adjusted OR = 3.3 (1.1 to 9.8). Early levels of vWf were also positively related to occurrence of all ischaemic events (crude HR = 2.3 (1.1 to 4.9); adjusted HR = 1.8 (0.8 to 3.9) and with occurrence of spontaneous DCI (crude HR = 3.5 (0.9 to 13.1); adjusted HR = 2.2 (0.5 to 9.8). None of the other markers showed any associations. Conclusions Concentrations of sICAM‐1, sP‐selectin, sE‐selectin, and ED1‐fibronectin do not predict the occurrence of DCI or outcome. The positive associations of raised early vWf concentrations with ischaemic events and poor outcome after SAH may reflect a predisposition to further ischaemic injury through formation of microthrombi in the cerebral circulation. PMID:16361599

  4. Effects of Complementary Creatine Monohydrate and Physical Training on Inflammatory and Endothelial Dysfunction Markers Among Heart Failure Patients

    PubMed Central

    Hemati, Farajollah; Rahmani, Asghar; Asadollahi, Khairollah; Soleimannejad, Koroush; Khalighi, Zahra

    2016-01-01

    Background: Previous studies have reported endothelial dysfunction and inflammatory cytokine in heart failure patients (HF). Objectives: The purpose of this study was to determine the effects of creatine monohydrate and exercise on inflammatory and endothelial dysfunction markers among HF patients. Patients and Methods: One hundred patients were prospectively randomized into two groups: Intervention group which received 5 grams/day creatine monohydrate and exercised for 8 weeks; and control group which did not receive any interventions. Interleukine-6 (IL-6), high sensitivity C reactive protein (hs-CRP), P-selectin, intercellular adhesion molecule-1 (ICAM-1), and vascular cell adhesion molecule 1 (VCAM-1) were measured at the start and end of the study for both groups. Results: In total, 100 patients including 50 controls and 50 intervention group (54% male, mean EF of 34.2 ± 10.5% and 52% male, mean EF of 35.6 ± 12.7%, respectively) were analyzed. The serum levels of hs-CRP and IL-6 increased at the end of the study in the control group compared to the baseline, (7.5 ± 1.5 mg/L vs. 6.9 ± 1.3 mg/L, P < 0.05 and 3.0 ± 0.75 ng/L vs. 2.55 ± 0.9 ng/L, P < 0.05, respectively). However, compared to the baseline, the level of both markers decreased at the end of the study in the intervention group (6.3 ± 1.6 mg/L vs.7.5 ± 1.5 mg/L, P < 0.05 and 2.1 ± 0.8 ng/L vs.2.5 ± 0.5 ng/L, P < 0.05). Also, P-selectin and ICAM-1 levels increased at the end of study (56.9 ± 1.8 ng/L vs. 51.9 ± 1.5 ng/L, P < 0.05 and 368.1 ± 25.4 µg/L vs. 353.1 ± 10.4 µg/L, P < 0.05 respectively). Inversely, the levels of these markers decreased in the intervention group, at the end of study (49.7 ± 1.9 ng/l vs. 51.4 ± 2.1 ng/l, P < 0.05 and 342.7 ± 16.5 µg/l vs. 350.4 ± 14.7 µg/l, P < 0.05, respectively). VCAM-1 level was not decreased significantly at the end of the study in the intervention group (570.5 ± 78.4 µg/L vs. 575.3 ± 86.5 µg/L, P > 0.05). Conclusions: Combination

  5. Immunohistochemical expression of endothelial markers CD31, CD34, von Willebrand factor, and Fli-1 in normal human tissues.

    PubMed

    Pusztaszeri, Marc P; Seelentag, Walter; Bosman, Fred T

    2006-04-01

    Few systematic studies have been published comparing the expression and distribution of endothelial cell (EC) markers in different vascular beds in normal human tissues. We investigated by immunohistochemistry the expression of CD31, CD34, von Willebrand factor (vWF), and Fli-1 in EC of the major organs and large vessels. Tissue samples obtained from autopsies and biopsy specimens were routinely processed and stained immunohistochemically for CD31, CD34, and vWF. Biopsy material was also stained immunohistochemically for Fli-1, D2-40, and Lyve-1. The expression pattern of the markers was heterogeneous in some of the organs studied. In the kidney, fenestrated endothelium of the glomeruli strongly expressed CD31 and CD34 but was only focally positive or completely negative for vWF. Alveolar wall capillaries of the lung strongly stained for CD31 and CD34 but were usually negative for vWF. The staining intensity for vWF increased gradually with the vessel caliber in the lung. Sinusoids of the spleen and liver were diffusely positive for CD31. They were negative for CD34 in the spleen and only expressed CD34 in the periportal area in the liver. Fli-1 was expressed in all types of EC but also in lymphocytes. D2-40 stained lymphatic endothelium only. Lyve-1 immunostaining was too variable to be applied to routinely processed tissues. The expression of EC markers CD31, CD34, and vWF in the vascular tree is heterogeneous with a specific pattern for individual vessel types and different anatomic compartments of the same organ. D2-40 labels lymphatic EC only.

  6. Targeting a novel domain in podoplanin for inhibiting platelet-mediated tumor metastasis

    PubMed Central

    Sekiguchi, Takaya; Takemoto, Ai; Takagi, Satoshi; Takatori, Kazuki; Sato, Shigeo; Takami, Miho; Fujita, Naoya

    2016-01-01

    Podoplanin/Aggrus is a sialoglycoprotein expressed in various cancers. We previously identified podoplanin as a key factor in tumor-induced platelet aggregation. Podoplanin-mediated platelet aggregation enhances tumor growth and metastasis by secreting growth factors and by forming tumor emboli in the microvasculature. Thus, precise analysis of the mechanisms of podoplanin-mediated platelet aggregation is critical for developing anti-tumor therapies. Here we report the discovery of a novel platelet aggregation-inducing domain, PLAG4 (81-EDLPT-85). PLAG4 has high homology to the previously reported PLAG3 and contributes to the binding of its platelet receptor CLEC-2. Mutant analyses indicated that PLAG4 exhibits a predominant platelet-aggregating function relative to PLAG3 and that conserved Glu81/Asp82/Thr85 residues in PLAG4 are indispensable for CLEC-2 binding. By establishing anti-PLAG4-neutralizing monoclonal antibodies, we confirmed its role in CLEC-2 binding, platelet aggregation, and tumor emboli formation. Our results suggest the requirement of simultaneous inhibition of PLAG3/4 for complete suppression of podoplanin-mediated tumor growth and metastasis. PMID:26684030

  7. Effect of exercise intensity on postprandial lipemia, markers of oxidative stress, and endothelial function after a high-fat meal.

    PubMed

    Lopes Krüger, Renata; Costa Teixeira, Bruno; Boufleur Farinha, Juliano; Cauduro Oliveira Macedo, Rodrigo; Pinto Boeno, Francesco; Rech, Anderson; Lopez, Pedro; Silveira Pinto, Ronei; Reischak-Oliveira, Alvaro

    2016-12-01

    The aim of this study was to compare the effects of 2 different exercise intensities on postprandial lipemia, oxidative stress markers, and endothelial function after a high-fat meal (HFM). Eleven young men completed 2-day trials in 3 conditions: rest, moderate-intensity exercise (MI-Exercise) and heavy-intensity exercise (HI-Exercise). Subjects performed an exercise bout or no exercise (Rest) on the evening of day 1. On the morning of day 2, an HFM was provided. Blood was sampled at fasting (0 h) and every hour from 1 to 5 h during the postprandial period for triacylglycerol (TAG), thiobarbituric acid reactive substance (TBARS), and nitrite/nitrate (NOx) concentrations. Flow-mediated dilatation (FMD) was also analyzed. TAG concentrations were reduced in exercise conditions compared with Rest during the postprandial period (P < 0.004). TAG incremental area under the curve (iAUC) was smaller after HI-Exercise compared with Rest (P = 0.012). TBARS concentrations were reduced in MI-Exercise compared with Rest (P < 0.041). FMD was higher in exercise conditions than Rest at 0 h (P < 0.02) and NOx concentrations were enhanced in MI-Exercise compared with Rest at 0 h (P < 0.01). These results suggest that acute exercise can reduce lipemia after an HFM. However, HI-Exercise showed to be more effective in reducing iAUC TAG, which might suggest higher protection against postprandial TAG enhancement. Conversely, MI-Exercise can be beneficial to attenuate the susceptibility of oxidative damage induced by an HFM and to increase endothelial function in the fasted state compared with Rest.

  8. Vascular Endothelial Growth Factor as an Angiogenic Marker in Malignant Astrocytoma and Oligodendroglioma: An Indian Scenario

    PubMed Central

    Vokuda, Ramya S; Srinivas, Bheemanathi Hanuman; Madhugiri, Venkatesh S

    2017-01-01

    Introduction The role of Vascular Endothelial Growth Factor (VEGF) in angiogenesis has been extensively studied in gliomas, such as astrocytoma and oligodendrogliomas, worldwide. However, there is limited information available with regard to the Indian population. Aim To study, whether VEGF is expressed in the Indian population in a pattern similar to that in other population. Materials and Methods In this prospective study approved by the Institute Ethics Committee for Human Studies at Jawaharlal Institute of Post Graduate Medical Education and Research (JIPMER) the patients operated for glioma in 2014 and 2015 (n = 60) were included. Tumours were graded as per the World Health Organization (WHO) grading system. VEGF expression in various grades was analysed using immunohistochemistry. Results Of the 60 patients included in this study, 15 were Grade II- (diffuse astrocytomas – 12; oligodendrogliomas- 3), 15 were Grade III-(anaplastic astrocytomas- 2; anaplastic oligodendrogliomas – 13) and 30 were Grade IV-glioblastomas. For VEGF antibody staining, two patients (3.33%) showed negative results and 58 patients (96%) showed positive results. VEGF positivity was 100% in Grade II and III, while it was 93.3% (28/30) in Grade IV tumours (p=0.012). Conclusion The expression of VEGF was associated with the grade of tumour, which gradually increased from Grade II to Grade IV. We conclude that VEGF-regulated angiogenesis plays an important role in tumour progression of astrocytomas and oligodendrogliomas in the Indian population as observed worldwide.

  9. Markers of Endothelial-to-Mesenchymal Transition: Evidence for Antibody-Endothelium Interaction during Antibody-Mediated Rejection in Kidney Recipients.

    PubMed

    Xu-Dubois, Yi-Chun; Peltier, Julie; Brocheriou, Isabelle; Suberbielle-Boissel, Caroline; Djamali, Arjang; Reese, Shannon; Mooney, Nuala; Keuylian, Zela; Lion, Julien; Ouali, Nacéra; Levy, Pierre P; Jouanneau, Chantal; Rondeau, Eric; Hertig, Alexandre

    2016-01-01

    Antibody-mediated rejection (ABMR) is a leading cause of allograft loss. Treatment efficacy depends on accurate diagnosis at an early stage. However, sensitive and reliable markers of antibody-endothelium interaction during ABMR are not available for routine use. Using immunohistochemistry, we retrospectively studied the diagnostic value of three markers of endothelial-to-mesenchymal transition (EndMT), fascin1, vimentin, and heat shock protein 47, for ABMR in 53 renal transplant biopsy specimens, including 20 ABMR specimens, 24 cell-mediated rejection specimens, and nine normal grafts. We validated our results in an independent set of 74 unselected biopsy specimens. Endothelial cells of the peritubular capillaries in grafts with ABMR expressed fascin1, vimentin, and heat shock protein 47 strongly, whereas those from normal renal grafts did not. The level of EndMT marker expression was significantly associated with current ABMR criteria, including capillaritis, glomerulitis, peritubular capillary C4d deposition, and donor-specific antibodies. These markers allowed us to identify C4d-negative ABMR and to predict late occurrence of disease. EndMT markers were more specific than capillaritis for the diagnosis and prognosis of ABMR and predicted late (up to 4 years after biopsy) renal graft dysfunction and proteinuria. In the independent set of 74 renal graft biopsy specimens, the EndMT markers for the diagnosis of ABMR had a sensitivity of 100% and a specificity of 85%. Fascin1 expression in peritubular capillaries was also induced in a rat model of ABMR. In conclusion, EndMT markers are a sensitive and reliable diagnostic tool for detecting endothelial activation during ABMR and predicting late loss of allograft function.

  10. Markers of endothelial and haemostatic function in the treatment of relapsed myeloma with the immunomodulatory agent Actimid (CC-4047) and their relationship with venous thrombosis.

    PubMed

    Streetly, Matthew; Hunt, Beverley J; Parmar, Kiran; Jones, Richard; Zeldis, Jerome; Schey, Steve

    2005-04-01

    We evaluated the serum/plasma levels of cytokines [interleukin (IL)-6, vascular endothelial growth factor (VEGF), transforming growth factor (TGF)-beta2] and markers of coagulation, fibrinolysis, endothelial and platelet activation during the first 4 wk of treatment with the thalidomide analogue Actimid (CC-4047) in 15 patients with relapsed/refractory myeloma. There was evidence of activation of endothelium (soluble vascular cell adhesion molecule, sVCAM), coagulation (prothrombin fragment 1 + 2, PF1 + 2) and fibrinolysis (D-dimers) but no evidence of platelet activation or endothelial cell damage in myeloma patients. These parameters were not affected by the use of CC-4047. Three of four patients with baseline D-dimers levels >500 microg/L subsequently developed deep vein thrombosis (DVT). The hypothesis that D-dimer level >500 microg/L may predict for those patients most at risk of thromboembolism with multiple myeloma undergoing treatment is worthy of further study.

  11. Prospective surface marker-based isolation and expansion of fetal endothelial colony-forming cells from human term placenta.

    PubMed

    Patel, Jatin; Seppanen, Elke; Chong, Mark S K; Yeo, Julie S L; Teo, Erin Y L; Chan, Jerry K Y; Fisk, Nicholas M; Khosrotehrani, Kiarash

    2013-11-01

    The term placenta is a highly vascularized tissue and is usually discarded upon birth. Our objective was to isolate clinically relevant quantities of fetal endothelial colony-forming cells (ECFCs) from human term placenta and to compare them to the well-established donor-matched umbilical cord blood (UCB)-derived ECFCs. A sorting strategy was devised to enrich for CD45-CD34+CD31Lo cells prior to primary plating to obtain pure placental ECFCs (PL-ECFCs) upon culture. UCB-ECFCs were derived using a well-described assay. PL-ECFCs were fetal in origin and expressed the same cell surface markers as UCB-ECFCs. Most importantly, a single term placenta could yield as many ECFCs as 27 UCB donors. PL-ECFCs and UCB-ECFCs had similar in vitro and in vivo vessel forming capacities and restored mouse hind limb ischemia in similar proportions. Gene expression profiles were only minimally divergent between PL-ECFCs and UCB-ECFCs, probably reflecting a vascular source versus a circulating source. Finally, PL-ECFCs and UCB-ECFCs displayed similar hierarchies between high and low proliferative colonies. We report a robust strategy to isolate ECFCs from human term placentas based on their cell surface expression. This yielded much larger quantities of ECFCs than UCB, but the cells were comparable in immunophenotype, gene expression, and in vivo functional ability. We conclude that PL-ECFCs have significant bio-banking and clinical translatability potential.

  12. Ligand-independent assembly of purified soluble magic roundabout (Robo4), a tumor-specific endothelial marker.

    PubMed

    Yoshikawa, Mai; Mukai, Yohei; Okada, Yoshiaki; Yoshioka, Yasuo; Tsunoda, Shin-Ichi; Tsutsumi, Yasuo; Okada, Naoki; Aird, William C; Doi, Takefumi; Nakagawa, Shinsaku

    2008-09-01

    Magic roundabout (Robo4) is the fourth recently identified member of the roundabout receptor family. Robo4 is predominantly expressed in embryonic or tumor vascular endothelium and is considered important for vascular development and as a candidate tumor endothelial marker. Much remains unknown about the Robo4 molecule, however, such as its ligands, structure, and the details of its function. Thus, we aimed to establish an expression and purification method for obtaining soluble recombinant human Robo4 (shRobo4) and mouse Robo4 (smRobo4) for use in Robo4 characterization studies. In this work, we expressed the extracellular domain of hRobo4 and mRobo4 in mammalian 293F cells and purified them by two-step chromatography. Based on gel-filtration chromatography and Blue Native polyacrylamide gel electrophoresis, these purified proteins exist as multimers. The shRobo4 and smRobo4 we obtained will be useful in advanced studies to determine the importance of multimerization, identify the ligands, and elucidate the ligand-receptor interactions and Robo4-mediated signaling. The results of these studies will help to elucidate the role of Robo4 in angiogenesis and perhaps eventually contribute to the development of novel vessel-targeting therapies.

  13. Identity of M2A (D2-40) antigen and gp36 (Aggrus, T1A-2, podoplanin) in human developing testis, testicular carcinoma in situ and germ-cell tumours.

    PubMed

    Sonne, Si Brask; Herlihy, Amy S; Hoei-Hansen, Christina E; Nielsen, John E; Almstrup, Kristian; Skakkebaek, Niels E; Marks, Alexander; Leffers, Henrik; Rajpert-De Meyts, Ewa

    2006-08-01

    Testicular germ-cell tumours of young adults are derived from a pre-invasive intratubular lesion, carcinoma in situ (CIS). In a recent genome-wide gene expression screening using cDNA microarrays, we found PDPN over-expressed in CIS compared to normal adult testis. PDPN encodes podoplanin (Aggrus, human gp36, T1A-2), a transmembrane glycoprotein expressed in lymphatic endothelium and various solid tumours. To examine a potential role for PDPN in testicular neoplasms and during testicular development, we investigated its expression pattern during the development of human testis and in a series of testicular CIS, gonadoblastoma and overt germ-cell tumours. We established by RT-PCR and by immunohistochemistry with a gp36 antibody that PDPN mRNA and the protein product were expressed in testes with germ-cell neoplasms but not in the normal adult testis. We also found gp36 expression in early foetal gonocytes and immature Sertoli cells, similar to the expression pattern of M2A antigen, a previously identified marker for CIS and seminoma. This reinforced our previous proposal that M2A (D2-40) antigen was identical to gp36 (podoplanin, Aggrus, T1A-2). Our findings also suggest that podoplanin has a function in developing testis, most likely at the level of cell-cell interactions among pre-meiotic germ cells and immature Sertoli cells.

  14. Cell adhesion markers are expressed by a stable human endothelial cell line transformed by the SV40 large T antigen under vimentin promoter control.

    PubMed

    Vicart, P; Testut, P; Schwartz, B; Llorens-Cortes, C; Perdomo, J J; Paulin, D

    1993-10-01

    Markers of endothelium have been studied in a new endothelial cell line derived from human umbilical cord vein cells by microinjection of a recombinant gene that includes a deletion mutant of the human vimentin gene regulatory region controlling the large T and small t antigen coding region of the SV40 virus. In culture, this immortalized venous endothelial cell line (IVEC) demonstrated morphological characteristics of endothelium; uptake of acetylated low density lipoprotein and presence of the Factor VIII-related antigen. Treatment of IVEC cells with Interleukin-1 beta (IL-1 beta) at 10 U.ml-1 activates the expression of cell adhesion molecules such as endothelial leucocyte adhesion molecule (ELAM-1), intercellular adhesion molecule-1 (ICAM-1), and vascular cell adhesion molecule-1 (VCAM-1), as observed in primary culture. Prostacyclin secretion was induced in the IVEC cells by 100 nM PMA treatment and thrombin at 0.5 U/ml. Angiotensin converting enzyme (ACE) activity detected in IVEC cells was present but lower than ACE activity in primary endothelial cells and was completely blocked by enalaprilat (1 microM), a specific ACE inhibitor. The presence of ACE mRNA was also demonstrated in IVEC cells by RT-PCR amplification. Our data demonstrate that endothelial cells immortalized by use of this recombinant gene retain the morphological organization and numerous differentiated properties of endothelium.

  15. Pathological lymphangiogenesis is modulated by galectin-8-dependent crosstalk between podoplanin and integrin-associated VEGFR-3

    PubMed Central

    Chen, Wei-Sheng; Cao, Zhiyi; Sugaya, Satoshi; Lopez, Maria J.; Sendra, Victor G.; Laver, Nora; Leffler, Hakon; Nilsson, Ulf J.; Fu, Jianxin; Song, Jianhua; Xia, Lijun; Hamrah, Pedram; Panjwani, Noorjahan

    2016-01-01

    Lymphangiogenesis plays a pivotal role in diverse pathological conditions. Here, we demonstrate that a carbohydrate-binding protein, galectin-8, promotes pathological lymphangiogenesis. Galectin-8 is markedly upregulated in inflamed human and mouse corneas, and galectin-8 inhibitors reduce inflammatory lymphangiogenesis. In the mouse model of corneal allogeneic transplantation, galectin-8-induced lymphangiogenesis is associated with an increased rate of corneal graft rejection. Further, in the murine model of herpes simplex virus keratitis, corneal pathology and lymphangiogenesis are ameliorated in Lgals8−/− mice. Mechanistically, VEGF-C-induced lymphangiogenesis is significantly reduced in the Lgals8−/− and Pdpn−/− mice; likewise, galectin-8-induced lymphangiogenesis is reduced in Pdpn−/− mice. Interestingly, knockdown of VEGFR-3 does not affect galectin-8-mediated lymphatic endothelial cell (LEC) sprouting. Instead, inhibiting integrins α1β1 and α5β1 curtails both galectin-8- and VEGF-C-mediated LEC sprouting. Together, this study uncovers a unique molecular mechanism of lymphangiogenesis in which galectin-8-dependent crosstalk among VEGF-C, podoplanin and integrin pathways plays a key role. PMID:27066737

  16. Podoplanin-rich stromal networks induce dendritic cell motility via activation of the C-type lectin receptor CLEC-2.

    PubMed

    Acton, Sophie E; Astarita, Jillian L; Malhotra, Deepali; Lukacs-Kornek, Veronika; Franz, Bettina; Hess, Paul R; Jakus, Zoltan; Kuligowski, Michael; Fletcher, Anne L; Elpek, Kutlu G; Bellemare-Pelletier, Angelique; Sceats, Lindsay; Reynoso, Erika D; Gonzalez, Santiago F; Graham, Daniel B; Chang, Jonathan; Peters, Anneli; Woodruff, Matthew; Kim, Young-A; Swat, Wojciech; Morita, Takashi; Kuchroo, Vijay; Carroll, Michael C; Kahn, Mark L; Wucherpfennig, Kai W; Turley, Shannon J

    2012-08-24

    To initiate adaptive immunity, dendritic cells (DCs) move from parenchymal tissues to lymphoid organs by migrating along stromal scaffolds that display the glycoprotein podoplanin (PDPN). PDPN is expressed by lymphatic endothelial and fibroblastic reticular cells and promotes blood-lymph separation during development by activating the C-type lectin receptor, CLEC-2, on platelets. Here, we describe a role for CLEC-2 in the morphodynamic behavior and motility of DCs. CLEC-2 deficiency in DCs impaired their entry into lymphatics and trafficking to and within lymph nodes, thereby reducing T cell priming. CLEC-2 engagement of PDPN was necessary for DCs to spread and migrate along stromal surfaces and sufficient to induce membrane protrusions. CLEC-2 activation triggered cell spreading via downregulation of RhoA activity and myosin light-chain phosphorylation and triggered F-actin-rich protrusions via Vav signaling and Rac1 activation. Thus, activation of CLEC-2 by PDPN rearranges the actin cytoskeleton in DCs to promote efficient motility along stromal surfaces.

  17. Circulating microparticles, protein C, free protein S and endothelial vascular markers in children with sickle cell anaemia

    PubMed Central

    Piccin, Andrea; Murphy, Ciaran; Eakins, Elva; Kunde, Jan; Corvetta, Daisy; Di Pierro, Angela; Negri, Giovanni; Guido, Mazzoleni; Sainati, Laura; Mc Mahon, Corrina; Smith, Owen Patrick; Murphy, William

    2015-01-01

    Introduction Circulating microparticles (MP) have been described in sickle cell anaemia (SCA); however, their interaction with endothelial markers remains unclear. We investigated the relationship between MP, protein C (PC), free protein S (PS), nitric oxide (NO), endothelin-1 (ET-1) and adrenomedullin (ADM) in a large cohort of paediatric patients. Method A total of 111 children of African ethnicity with SCA: 51 in steady state; 15 in crises; 30 on hydroxyurea (HU) therapy; 15 on transfusion; 17 controls (HbAA) of similar age/ethnicity. MP were analysed by flow cytometry using: Annexin V (AV), CD61, CD42a, CD62P, CD235a, CD14, CD142 (tissue factor), CD201 (endothelial PC receptor), CD62E, CD36 (TSP-1), CD47 (TSP-1 receptor), CD31 (PECAM), CD144 (VE-cadherin). Protein C, free PS, NO, pro-ADM and C-terminal ET-1 were also measured. Results Total MP AV was lower in crisis (1.26×106 ml−1; 0.56–2.44×106) and steady state (1.35×106 ml−1; 0.71–3.0×106) compared to transfusion (4.33×106 ml−1; 1.6–9.2×106, p<0.01). Protein C levels were significantly lower in crisis (median 0.52 IU ml−1; interquartile range 0.43–0.62) compared with all other groups: HbAA (0.72 IU ml−1; 0.66–0.82, p<0.001); HU (0.67 IU ml−1; 0.58–0.77, p<0.001); steady state (0.63 IU ml−1; 0.54–0.70, p<0.05) and transfusion (0.60 IU ml−1; 0.54–0.70, p<0.05). In addition, levels were significantly reduced in steady state (0.63 IU ml−1; 0.54–0.70) compared with HbAA (0.72 IU ml−1; 0.66–0.80, p<0.01). PS levels were significantly higher in HbAA (0.85 IU ml−1; 0.72–0.97) compared with crisis (0.49 IU ml−1; 0.42–0.64, p<0.001), HU (0.65 IU ml−1; 0.56–0.74, p<0.01) and transfusion (0.59 IU ml−1; 0.47–0.71, p<0.01). There was also a significant difference in crisis patients compared with steady state (0.49 IU ml−1; 0.42–0.64 vs. 0.68 IU ml−1; 0.58–0.79, p<0.05). There was high correlation (R>0.9, p<0.05) between total numbers of AV-positive MP

  18. The impact of decreases in air temperature and increases in ozone on markers of endothelial function in individuals having type-2 diabetes.

    PubMed

    Lanzinger, Stefanie; Breitner, Susanne; Neas, Lucas; Cascio, Wayne; Diaz-Sanchez, David; Hinderliter, Alan; Peters, Annette; Devlin, Robert B; Schneider, Alexandra

    2014-10-01

    Several studies have reported an association between air pollution and endothelial dysfunction, especially in individuals having diabetes. However, very few studies have examined the impact of air temperature on endothelial function. The objective of this analysis was to investigate short-term effects of temperature and ozone on endothelial function in individuals having diabetes. Moreover, we investigated interactive effects between air temperature and air pollution on markers of endothelial function. Between November 2004 and December 2005 flow-mediated dilatation (FMD), nitroglycerin-mediated dilatation (NTGMD) and several blood markers representing endothelial function were measured using brachial artery ultrasound on four consecutive days in 22 individuals with type-2 diabetes mellitus in Chapel Hill, North Carolina (USA). Daily measurements of meteorological parameters, ozone and particulate matter with an aerodynamic diameter ≤2.5 µm (PM2.5) were obtained from fixed monitoring sites. We used additive mixed-models adjusting for time trend, day of the week, relative humidity and barometric pressure to assess temperature and ozone associations with endothelial function. A 1 °C decrease in the 24-h temperature average was associated with a decrease in mean FMD on the same day (-2.2% (95%-confidence interval:[-4.7;0.3%])) and with a delay of one and four days. A temperature decrement also led to an immediate (-1.7%[-3.3;-0.04]) decrease in NTGMD. Moreover, we observed an immediate (-14.6%[-26.3;-2.9%]) and a one day delayed (-13.5%[-27.0; 0.04%]) decrease in FMD in association with a 0.01 ppm increase in the maximum 8-h moving average of ozone. Temperature effects on FMD strengthened when PM2.5 and ozone concentrations were high. The associations were similar during winter and summer. We detected an association between temperature decreases and ozone increases on endothelial dysfunction in individuals having diabetes. We conclude that endothelial dysfunction

  19. Ets-1 is required for the activation of VEGFR3 during latent Kaposi's sarcoma-associated herpesvirus infection of endothelial cells.

    PubMed

    Gutierrez, Kimberley D; Morris, Valerie A; Wu, David; Barcy, Serge; Lagunoff, Michael

    2013-06-01

    Kaposi's sarcoma-associated herpesvirus (KSHV), the etiologic agent of Kaposi's sarcoma (KS), is present in the predominant tumor cells of KS, the spindle cells. Spindle cells express markers of lymphatic endothelium and, interestingly, KSHV infection of blood endothelial cells reprograms them to a lymphatic endothelial cell phenotype. KSHV-induced reprogramming requires the activation of STAT3 and phosphatidylinositol 3 (PI3)/AKT through the activation of cellular receptor gp130. Importantly, KSHV-induced reprogramming is specific to endothelial cells, indicating that there are additional host genes that are differentially regulated during KSHV infection of endothelial cells that contribute to lymphatic reprogramming. We found that the transcription factor Ets-1 is highly expressed in KS spindle cells and is upregulated during KSHV infection of endothelial cells in culture. The KSHV latent vFLIP gene is sufficient to induce Ets-1 expression in an NF-κB-dependent fashion. Ets-1 is required for KSHV-induced expression of VEGFR3, a lymphatic endothelial-cell-specific receptor important for lymphangiogenesis, and Ets-1 activates the promoter of VEGFR3. Ets-1 knockdown does not alter the expression of another lymphatic-specific gene, the podoplanin gene, but does inhibit the expression of VEGFR3 in uninfected lymphatic endothelium, indicating that Ets-1 is a novel cellular regulator of VEGFR3 expression. Knockdown of Ets-1 affects the ability of KSHV-infected cells to display angiogenic phenotypes, indicating that Ets-1 plays a role in KSHV activation of endothelial cells during latent KSHV infection. Thus, Ets-1 is a novel regulator of VEGFR3 and is involved in the induction of angiogenic phenotypes by KSHV.

  20. Air pollution and markers of coagulation, inflammation and endothelial function: Associations and epigene-environment interactions in an elderly cohort

    PubMed Central

    Bind, Marie-Abele; Baccarelli, Andrea; Zanobetti, Antonella; Tarantini, Letizia; Suh, Helen; Vokonas, Pantel; Schwartz, Joel

    2012-01-01

    BACKGROUND Previous studies suggest that air pollution is related to thrombosis, inflammation, and endothelial dysfunction. Mechanisms and sources of susceptibility are still unclear. One possibility is that these associations can be modified by DNA methylation states. METHODS We conducted a cohort study with repeated measurements of fibrinogen, C-reactive protein, intercellular adhesion molecule-1 (ICAM-1), and vascular cell adhesion molecule-1 (VCAM-1) in 704 elderly men participating in the Veterans Administration Normative Aging Study (2000-2009). We investigated short- and intermediate-term air pollution effects on these blood markers, and epigene-environment interactions by DNA methylation of Alu, LINE-1, tissue factor (F3), Toll-Like Receptor 2 (TLR-2), and ICAM-1. RESULTS We found effects of particle number, black carbon, nitrogen dioxide (NO2), and carbon monoxide (CO) on fibrinogen. Ozone was a significant predictor of C-reactive protein and ICAM-1. Particle number, black carbon, NO2, CO, PM2.5, and sulfates were associated with ICAM-1and VCAM-1. An interquartile range increase in 24-hour exposure for NO2; was associated with a 1.7% (95% confidence interval = 0.2% to 3.3%) increase in fibrinogen for ozone a 10.8% (2.2% to 20.0%) increase in C-reactive protein for particle number, a 5.9% (3.6% to 8.3%) increase in ICAM-1; and for PM2.5 a 3.7% (1.7% to 5.8%) increase in VCAM-1. The air pollution effect was stronger among subjects having higher Alu, lower LINE-1, tissue factor, or TLR-2 methylation status. CONCLUSION We observed associations of traffic-related pollutants on fibrinogen, and both traffic and secondary particles on C-reactive protein, ICAM-1, and VCAM-1. There was effect modification by DNA methylation status, indicating that epigenetic states can convey susceptibility to air pollution. PMID:22237295

  1. Tumor endothelial marker 8 expression levels in dendritic cell-based cancer vaccines are related to clinical outcome.

    PubMed

    Venanzi, Franco Maria; Petrini, Massimiliano; Fiammenghi, Laura; Bolli, Elisabetta; Granato, Anna Maria; Ridolfi, Laura; Gabrielli, Federica; Barucca, Alessandra; Concetti, Antonio; Ridolfi, Ruggero; Riccobon, Angela

    2010-01-01

    Previous studies have shown that tumor endothelial markers (TEMs 1-9) are up modulated in immunosuppressive, pro-angiogenic dendritic cells (DCs) found in tumor microenvironments. We recently reported that monocyte-derived DCs used for vaccination trials may accumulate high levels of TEM8 gene transcripts. Here, we investigate whether TEM8 expression in DC preparations represents a specific tumor-associated change of potential clinical relevance. TEM8 expression at the mRNA and protein level was evaluated by quantitative real-time RT-PCR and cytofluorimetric analysis in human clinical grade DCs utilized for the therapeutic vaccination of 17 advanced cancer patients (13 melanoma and 4 renal cell carcinoma). The analyses revealed that DCs from patients markedly differ in their ability to up-modulate TEM8. Indeed, mDCs from eight non-progressing patients [median overall survival (OS) = 32 months, all positive to the delayed-type hypersensitivity test (DTH)], had similar TEM8 mRNA expression levels [mDCs vs. immature iDCs; mean fold increase (mfi) = 1.97] to those found in healthy donors (mfi = 2.7). Conversely, mDCs from nine progressing patients (OS < 5 months, all but one with negative DTH) showed an increase in TEM8 mRNA levels (mfi = 12.88, p = 0.0018). The present observations suggest that TEM8 expression levels in DC-based therapeutic vaccines would allow the selection of a subgroup of patients who are most likely to benefit from therapeutic vaccination.

  2. Markers

    ERIC Educational Resources Information Center

    Healthy Schools Network, Inc., 2011

    2011-01-01

    Dry erase whiteboards come with toxic dry erase markers and toxic cleaning products. Dry erase markers labeled "nontoxic" are not free of toxic chemicals and can cause health problems. Children are especially vulnerable to environmental health hazards; moreover, schools commonly have problems with indoor air pollution, as they are more densely…

  3. Endovascular treatment of chronic cerebro spinal venous insufficiency in patients with multiple sclerosis modifies circulating markers of endothelial dysfunction and coagulation activation: a prospective study.

    PubMed

    Napolitano, Mariasanta; Bruno, Aldo; Mastrangelo, Diego; De Vizia, Marcella; Bernardo, Benedetto; Rosa, Buonagura; De Lucia, Domenico

    2014-10-01

    We performed a monocentric observational prospective study to evaluate coagulation activation and endothelial dysfunction parameters in patients with multiple sclerosis undergoing endovascular treatment for cerebro-spinal-venous insufficiency. Between February 2011 and July 2012, 144 endovascular procedures in 110 patients with multiple sclerosis and chronical cerebro-spinal venous insufficiency were performed and they were prospectively analyzed. Each patient was included in the study according to previously published criteria, assessed by the investigators before enrollment. Endothelial dysfunction and coagulation activation parameters were determined before the procedure and during follow-up at 1, 3, 6, 9, 12, 15 and 18 months after treatment, respectively. After the endovascular procedure, patients were treated with standard therapies, with the addition of mesoglycan. Fifty-five percent of patients experienced a favorable outcome of multiple sclerosis within 1 month after treatment, 25% regressed in the following 3 months, 24.9% did not experience any benefit. In only 0.1% patients, acute recurrence was observed and it was treated with high-dose immunosuppressive therapy. No major complications were observed. Coagulation activation and endothelial dysfunction parameters were shown to be reduced at 1 month and stable up to 12-month follow-up, and they were furthermore associated with a good clinical outcome. Endovascular procedures performed by a qualified staff are well tolerated; they can be associated with other currently adopted treatments. Correlations between inflammation, coagulation activation and neurodegenerative disorders are here supported by the observed variations in plasma levels of markers of coagulation activation and endothelial dysfunction.

  4. Micro- and nano-topography to enhance proliferation and sustain functional markers of donor-derived primary human corneal endothelial cells.

    PubMed

    Muhammad, Rizwan; Peh, Gary S L; Adnan, Khadijah; Law, Jaslyn B K; Mehta, Jodhbir S; Yim, Evelyn K F

    2015-06-01

    One of the most common indications for corneal transplantation is corneal endothelium dysfunction, which can lead to corneal blindness. Due to a worldwide donor cornea shortage, alternative treatments are needed, but the development of new treatment strategies relies on the successful in vitro culture of primary human corneal endothelial cells (HCECs) because transformed cell lines and animal-derived corneal endothelial cells are not desirable for therapeutic applications. Primary HCECs are non-proliferative in vivo and challenging to expand in vitro while maintaining their characteristic cell morphology and critical markers. Biochemical cues such as growth factors and small molecules have been investigated to enhance the expansion of HCECs with a limited increase in proliferation. In this study, patterned tissue culture polystyrene (TCPS) was shown to significantly enhance the expansion of HCECs. The proliferation of HCECs increased up to 2.9-fold, and the expression amount and localization of cell-cell tight junction protein Zona Occludens-1 (ZO-1) was significantly enhanced when grown on 1 μm TCPS pillars. 250 nm pillars induced an optimal hexagonal morphology of HCEC cells. Furthermore, we demonstrated that the topographical effect on tight-junction expression and cell morphology could be maintained throughout each passage, and was effectively 'remembered' by the cells. Higher amount of tight-junction protein expression was maintained at cell junctions when topographic cues were removed in the successive seeding. This topographic memory suggested topography-exposed/induced cells would maintain the enhanced functional markers, which would be useful in cell-therapy based approaches to enable the in situ endothelial cell monolayer formation upon delivery. The development of patterned TCPS culture platforms could significantly benefit those researching human corneal endothelial cell cultivation for cell therapy, and tissue engineering applications.

  5. Markers of endothelial cell activation and immune activation are increased in patients with severe leptospirosis and associated with disease severity

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Objectives: Previous studies concluded that haemorrhage is one of the most accurate prognostic factors of mortality in leptospirosis. Therefore, endothelial cell activation was investigated in relation to disease severity in severe leptospirosis. Methods: Prospective cohort study of severe leptospi...

  6. Expression of Von Willebrand factor, an endothelial cell marker, is up-regulated by angiogenesis factors: a potential method for objective assessment of tumor angiogenesis.

    PubMed

    Zanetta, L; Marcus, S G; Vasile, J; Dobryansky, M; Cohen, H; Eng, K; Shamamian, P; Mignatti, P

    2000-01-15

    von Willebrand factor (vWF), a glycoprotein produced uniquely by endothelial cells and megakaryocytes, is routinely used to identify vessels in tissue sections. Vessel density in tumor specimens, as determined by immuno-histochemical staining for vWF or other endothelial cell markers, is a negative prognostic factor for many solid tumors. vWF is heterogeneously distributed throughout the vasculature, transcriptional control in response to the tissue microenvironment being responsible for local variations in endothelial cell levels of vWF. Here, we report that fibroblast growth factor-2 and vascular endothelial growth factor, potent angiogenesis inducers expressed in a variety of tumors, up-regulate expression of vWF mRNA and protein in cultured endothelial cells with a synergistic effect. Our data support the measurement of vWF mRNA in tumors to detect activated endothelium or angiogenesis. For this purpose, we developed a semi-quantitative RT-PCR for vWF mRNA. Preliminary results obtained with specimens from colon carcinoma and the corresponding normal colonic mucosa showed higher vWF mRNA levels in most tumors than in their normal counterparts. The differences in vWF mRNA levels were much larger than the differences in vessel counts between a tumor and the corresponding normal mucosa, indicating that high vWF mRNA levels in tumors may indeed be an early sign of activation of the endothelium. The rapidity, objectivity, sensitivity and specificity of this technique make it suitable for routine clinical application to identify aggressive, highly angiogenic tumors.

  7. A transcriptome signature of endothelial lymphatic cells coexists with the chronic oxidative stress signature in radiation-induced post-radiotherapy breast angiosarcomas.

    PubMed

    Hadj-Hamou, Nabila-Sandra; Laé, Marick; Almeida, Anna; de la Grange, Pierre; Kirova, Youlia; Sastre-Garau, Xavier; Malfoy, Bernard

    2012-07-01

    Radiation-induced breast angiosarcomas are rare but recognized complication of breast cancer radiotherapy and are of poor prognosis. Little is known about the genetic abnormalities present in these secondary tumors. Herein, we investigated the differences in the genome and in the transcriptome that discriminate these tumors as a function of their etiology. Seven primary breast angiosarcomas and 18 secondary breast angiosarcomas arising in the irradiation field of a radiotherapy were analyzed. Copy number alterations and gene expression were analyzed using Affymetrix SNP 6.0 Array and Affymetrix Exon Arrays, respectively. We showed that two transcriptome signatures of the radiation tumorigenesis coexisted in these tumors. One was histology specific and correctly discriminated 100% of the primary tumors from the radiation-induced tumors. The deregulation of marker genes, including podoplanin (PDPN), prospero homeobox 1 (PROX-1), vascular endothelial growth factor 3 (VEGFR3) and endothelin receptor A (EDNRA), suggests that the radiation-induced breast angiosarcomas developed from radiation-stimulated lymphatic endothelial cells. None of the genes of the histology-specific signature were present in our previously published signature of the radiation tumorigenesis which shows the presence of a chronic oxidative stress in radiation-induced sarcomas of various histologies. Nevertheless, this oxidative stress signature classified correctly 88% of the breast angiosarcomas as a function of the etiology. In contrast, MYC amplification, which is observed in all radiation-induced tumors but also at a low rate in primary tumors, was not a marker of the radiation tumorigenesis.

  8. Suppression of Aggrus/podoplanin-induced platelet aggregation and pulmonary metastasis by a single-chain antibody variable region fragment.

    PubMed

    Miyata, Kenichi; Takagi, Satoshi; Sato, Shigeo; Morioka, Hiroshi; Shiba, Kiyotaka; Minamisawa, Tamiko; Takami, Miho; Fujita, Naoya

    2014-12-01

    Almost all highly metastatic tumor cells possess high platelet aggregating abilities, thereby form large tumor cell-platelet aggregates in the microvasculature. Embolization of tumor cells in the microvasculature is considered to be the first step in metastasis to distant organs. We previously identified the platelet aggregation-inducing factor expressed on the surfaces of highly metastatic tumor cells and named as Aggrus. Aggrus was observed to be identical to the marker protein podoplanin (alternative names, T1α, OTS-8, and others). Aggrus is frequently overexpressed in several types of tumors and enhances platelet aggregation by interacting with the platelet receptor C-type lectin-like receptor 2 (CLEC-2). Here, we generated a novel single-chain antibody variable region fragment (scFv) by linking the variable regions of heavy and light chains of the neutralizing anti-human Aggrus monoclonal antibody MS-1 with a flexible peptide linker. Unfortunately, the generated KM10 scFv failed to suppress Aggrus-induced platelet aggregation in vitro. Therefore, we performed phage display screening and finally obtained a high-affinity scFv, K-11. K-11 scFv was able to suppress Aggrus-induced platelet aggregation in vitro. Moreover, K-11 scFv prevented the formation of pulmonary metastasis in vivo. These results suggest that K-11 scFv may be useful as metastasis inhibitory scFv and is expected to aid in the development of preclinical and clinical examinations of Aggrus-targeted cancer therapies.

  9. Effects of spinach nitrate on insulin resistance, endothelial dysfunction markers and inflammation in mice with high-fat and high-fructose consumption

    PubMed Central

    Li, Ting; Lu, Xinshan; Sun, Yanfei; Yang, Xingbin

    2016-01-01

    Background Insulin resistance, which is associated with an increased risk of cardiovascular morbidity and mortality, has become a leading nutrition problem. Inorganic nitrate enriched in spinach has been demonstrated to reverse the pathological features of insulin resistance and endothelial dysfunction. However, the effects of a direct intake of nitrate-enriched spinach on insulin resistance and endothelial dysfunction have not been studied. Objective To investigate the effects of spinach nitrate on insulin resistance, lipid metabolism, endothelial function, and inflammation in mice fed with a high-fat and high-fructose diet. Design A diet intervention of spinach with or without nitrate was performed in mice. A high-fat and high-fructose diet was used to cause insulin resistance, endothelial dysfunction, and inflammation in mice. The impacts of spinach nitrate on lipid profile, insulin resistance, markers of endothelial function, and inflammation were determined in mice. Results Spinach nitrate improved the vascular endothelial function of the mice with high-fat and high-fructose consumption, as evidenced by the elevated plasma nitrite level, increased serum nitric oxide (NO) level and decreased serum ET-1 level after spinach nitrate intervention. Spinach nitrate also reduced serum triglycerides, total cholesterol, and low-density lipoprotein-cholesterol levels and elevated serum high-density lipoprotein-cholesterol levels in the mice fed with a high-fat and high-fructose diet. Mice receiving spinach with 60 mg/kg of nitrate (1.02±0.34) showed a significantly low homeostasis model assessment-insulin resistance index as compared with the model mice (2.05±0.58), which is indicating that spinach nitrate could effectively improve the insulin resistance. In addition, spinach nitrate remarkably decreased the elevated serum C-reactive protein, tumor necrosis factor α, and interleukin-6 levels induced by a high-fat and high-fructose diet. Conclusions The intake of

  10. Co-Expression of Bmi-1 and Podoplanin Predicts Overall Survival in Patients With Squamous Cell Carcinoma of the Head and Neck Treated With Radio(chemo)therapy

    SciTech Connect

    Vormittag, Laurenz; Thurnher, Dietmar; Geleff, Silvana; Pammer, Johannes; Heiduschka, Gregor; Brunner, Markus; Grasl, Matthaeus Ch.; Erovic, Boban M.

    2009-03-01

    Purpose: This study was conducted to determine the expression of Bmi-1 and podoplanin in healthy oral mucosa and in untreated tumor tissues samples of patients with squamous cell carcinomas of the head and neck. All patients were treated by primary radio(chemo)therapy. Methods and Materials: The expression of Bmi-1 and podoplanin was immunohistochemically evaluated in 12 normal oral mucosa and 63 tumor specimens and correlated with patients' clinical data. Results: In healthy mucosa expression of Bmi-1 and podoplanin was restricted to the basal cell layer. Expression of both proteins was found in 79% and 86% of our tumor samples, respectively. In 17 and 8 samples, Bmi-1 and podoplanin were co-expressed at the invasive border or diffuse in the bulk of the tumor, respectively. Univariate analysis showed that the co-expression of Bmi-1 and podoplanin correlated to decreased overall survival (p = 0.044). Moreover, multivariate testing identified high expression of podoplanin (p = 0.044), co-expression of Bmi-1 and podoplanin (p = 0.007) and lack of response to therapy (p < 0.0001) as predictors of shortened overall survival in patients treated with primary radio(chemo)therapy. Conclusions: Bmi-1 and podoplanin are expressed at the invasive front of squamous cell carcinomas of the head and neck. Co-expression of Bmi-1 and podoplanin predicts significantly overall survival of patients treated with primary radio(chemo)therapy.

  11. Use of tritiated thymidine as a marker to compare the effects of matrix proteins on adult human vascular endothelial cell attachment: implications for seeding of vascular prostheses

    SciTech Connect

    Hasson, J.E.; Wiebe, D.H.; Sharefkin, J.B.; D'Amore, P.A.; Abbott, W.M.

    1986-11-01

    We have developed a technique to measure attachment of adult human vascular endothelial cells to test surfaces with tritiated thymidine used as a marker. With this technique, we measured attachment of adult human vascular endothelial cells to a series of extracellular matrix proteins, including fibronectin-coated (10 micrograms/cm/sup 2/), laminin-coated (10 micrograms/cm/sup 2/), and collagen-coated (1% gelatin) surfaces because of the role of these proteins in promoting cell attachment and growth. For a typical experiment, in the presence of serum, initial attachment (at 1 hour) was greatest on fibronectin-coated (63%) and gelatin-coated (60%) tissue culture plastic (polystyrene) and was least on laminin-coated (28%) or untreated polystyrene (18%). The data suggest that fibronectin, either alone, or with a more complex combination of extracellular components may need to be present on prosthetic surfaces to produce maximal cell attachment and subsequent growth to confluence in vivo. The described method of measuring attachment is independent of surface properties, ensures complete recovery of cells, and will allow systematic exploration of those properties that best support human endothelial cell attachment to vascular prosthetic surfaces.

  12. Epithelial deletion of podoplanin is dispensable for re-epithelialization of skin wounds.

    PubMed

    Baars, Sebastian; Bauer, Christine; Szabowski, Sibylle; Hartenstein, Bettina; Angel, Peter

    2015-10-01

    The mucin-like transmembrane protein podoplanin (PDPN) is prominently represented in tumor-associated gene expression signatures of numerous types of cancer including squamous cell carcinoma, and gain-of-function and knockdown approaches in tissue culture strongly suggested an important role of PDPN in cell proliferation, migration and adhesion. PDPN is absent during epidermal homeostasis but is highly expressed in basal keratinocytes during cutaneous wound healing. Enhanced motility of immortalized keratinocytes upon ectopic PDPN overexpression argues for wound healing defects upon podoplanin deficiency in keratinocytes; however, in vivo data that unequivocally define the impact of PDPN by functional studies in a physiologically relevant system are still missing. Here, we have applied an in vivo loss-of-function approach by generating a novel transgenic mouse line with keratinocyte-specific podoplanin deficiency. Performing cutaneous full-thickness excisional wounds to examine re-epithelialization capacity, unexpectedly, no defects were observed in wound healing properties of mutant mice. Similarly, PDPN-deficient primary keratinocytes showed no impairment in migration, adhesion or proliferation. Thus, PDPN function is not rate-limiting for re-epithelialization but may be functionally compensated by an as yet unknown protein. Our data also call for in vivo functional studies on PDPN in settings of skin tumor development and progression to clarify PDPN's role in skin pathology.

  13. Phosphorylated mTOR expression correlates with podoplanin expression and high tumor grade in esophageal squamous cell carcinoma

    PubMed Central

    Chuang, Wen-Yu; Chang, Yu-Sun; Chao, Yin-Kai; Yeh, Chi-Ju; Ueng, Shir-Hwa; Chang, Chiu-Yueh; Liu, Yun-Hen; Tseng, Chen-Kan; Chang, Hsien-Kun; Wan, Yung-Liang; Hsueh, Chuen

    2015-01-01

    Mechanistic (or mammalian) target of rapamycin (mTOR) plays important roles in cell growth and proliferation. In esophageal squamous cell carcinoma (SCC), high expression of phosphorylated (activated) mTOR (p-mTOR) has been reported as an adverse prognostic factor in some but not all studies. The signals of mTOR pathway and mitogen-activated protein kinase (MAPK) pathway converge on 4E-binding protein 1 (4EBP1), which drives the downstream proliferative signals. We previously found that high expression of phosphorylated 4EBP1 (p-4EBP1) is an adverse prognostic factor in esophageal SCC. Podoplanin is a type-1 transmembrane glycoprotein expressed in various normal human tissues, including lymphatic endothelium. Our previous study showed that high podoplanin expression correlates with clinical nodal metastasis, which is associated with short survival in esophageal SCC. In current study, we investigated p-mTOR expression by immunohistochemistry in 75 cases of surgically resected esophageal SCC. The result was correlated with p-4EBP1 expression, podoplanin expression, clinicopathologic features and patient survival. We found that high p-mTOR expression was significantly associated with high podoplanin expression (P = 0.0030) and high tumor grade (P = 0.0014). No correlation with p-4EBP1 expression, patient survival or other clinicopathologic features was found. Recently, podoplanin expression in astrocytic brain tumors was found to be regulated by the phosphatidylinositol 3-kinase (PI3K)/AKT/activator protein-1 (AP-1) pathway. Similarly, mTOR is activated by a PI3K/AKT/mTOR pathway. The association of p-mTOR and podoplanin expression in our study could be due to a common upstream pathway. Since both mTOR and podoplanin are potential therapeutic targets, the possible benefit of combined targeted therapy warrants further investigation. PMID:26722465

  14. Development and characterization of anti-glycopeptide monoclonal antibodies against human podoplanin, using glycan-deficient cell lines generated by CRISPR/Cas9 and TALEN.

    PubMed

    Kaneko, Mika K; Nakamura, Takuro; Honma, Ryusuke; Ogasawara, Satoshi; Fujii, Yuki; Abe, Shinji; Takagi, Michiaki; Harada, Hiroyuki; Suzuki, Hiroyoshi; Nishioka, Yasuhiko; Kato, Yukinari

    2017-02-01

    Human podoplanin (hPDPN), which binds to C-type lectin-like receptor-2 (CLEC-2), is involved in platelet aggregation and cancer metastasis. The expression of hPDPN in cancer cells or cancer-associated fibroblasts indicates poor prognosis. Human lymphatic endothelial cells, lung-type I alveolar cells, and renal glomerular epithelial cells express hPDPN. Although numerous monoclonal antibodies (mAbs) against hPDPN are available, they recognize peptide epitopes of hPDPN. Here, we generated a novel anti-hPDPN mAb, LpMab-21. To characterize the hPDPN epitope recognized by the LpMab-21, we established glycan-deficient CHO-S and HEK-293T cell lines, using the CRISPR/Cas9 or TALEN. Flow cytometric analysis revealed that the minimum hPDPN epitope, in which sialic acid is linked to Thr76, recognized by LpMab-21 is Thr76-Arg79. LpMab-21 detected hPDPN expression in glioblastoma, oral squamous carcinoma, and seminoma cells as well as in normal lymphatic endothelial cells. However, LpMab-21 did not react with renal glomerular epithelial cells or lung type I alveolar cells, indicating that sialylation of hPDPN Thr76 is cell-type-specific. LpMab-21 combined with other anti-hPDPN antibodies that recognize different epitopes may therefore be useful for determining the physiological function of sialylated hPDPN.

  15. A critical role of platelet TGF-β release in podoplanin-mediated tumour invasion and metastasis

    PubMed Central

    Takemoto, Ai; Okitaka, Mina; Takagi, Satoshi; Takami, Miho; Sato, Shigeo; Nishio, Makoto; Okumura, Sakae; Fujita, Naoya

    2017-01-01

    The tumour microenvironment is critical for various characteristics of tumour malignancies. Platelets, as part of the tumour microenvironment, are associated with metastasis formation via increasing the rate of tumour embolus formation in microvasculature. However, the mechanisms underlying the ability of tumour cells to acquire invasiveness and extravasate into target organs at the site of embolization remain unclear. In this study, we reported that platelet aggregation-inducing factor podoplanin expressed on tumour cell surfaces were found to not only promote the formation of tumour-platelet aggregates via interaction with platelets, but also induced the epithelial-mesenchymal transition (EMT) of tumour cells by enhancing transforming growth factor-β (TGF-β) release from platelets. In vitro and in vivo analyses revealed that podoplanin-mediated EMT resulted in increased invasiveness and extravasation of tumour cells. Treatment of mice with a TGF-β-neutralizing antibody statistically suppressed podoplanin-mediated distant metastasis in vivo, suggesting that podoplanin promoted haematogenous metastasis in part by releasing TGF-β from platelets that was essential for EMT of tumour cells. Therefore, our findings suggested that blocking the TGF-β signalling pathway might be a promising strategy for suppressing podoplanin-mediated haematogenous metastasis in vivo. PMID:28176852

  16. Expression of prox1, lymphatic endothelial nuclear transcription factor, in Kaposiform hemangioendothelioma and tufted angioma.

    PubMed

    Le Huu, Aude Rimella; Jokinen, Chris H; Rubin, Brian P; Ruben, Brian P; Mihm, Martin C; Weiss, Sharon W; North, Paula E; Dadras, Soheil S

    2010-11-01

    Kaposiform hemangioendothelioma (KHE) and tufted angioma (TA) are rare tumors mainly occurring in early childhood. Our recent results showed that ectopic overexpression of human Prox1 gene, a lymphatic endothelial nuclear transcription factor, promoted an aggressive behavior in 2 murine models of KHE. This dramatic Prox1-induced phenotype prompted us to investigate immunohistochemical staining pattern of Prox1, podoplanin (D2-40), LYVE-1, and Prox1/CD34 as well as double immunofluorescent staining pattern of LYVE-1/CD31 in KHE and TA, compared with other pediatric vascular tumors. For this purpose, we examined 75 vascular lesions: KHE (n=18), TA (n=13), infantile hemangioma (n=13), pyogenic granuloma (n=18), and granulation tissue (n=13). Overall, KHE and TA shared an identical endothelial immunophenotype: the neoplastic spindle cells were Prox1, podoplanin, LYVE-1, CD31, and CD34, whereas endothelial cells within glomeruloid foci were Prox1, podoplanin, LYVE-1, CD31, and CD34. The lesional cells of all infantile hemangiomas and pyogenic granulomas were negative for Prox1 in the presence of positive internal control. These findings provide immunophenotypic evidence to support a preexisting notion that KHE and TA are closely related, if not identical. Overall, our results show, for the first time, that Prox1 is an immunohistochemical biomarker helpful in confirming the diagnosis of KHE/TA and in distinguishing it from infantile hemangioma and pyogenic granuloma.

  17. Physical exercise, fitness and dietary pattern and their relationship with circadian blood pressure pattern, augmentation index and endothelial dysfunction biological markers: EVIDENT study protocol

    PubMed Central

    2010-01-01

    Background Healthy lifestyles may help to delay arterial aging. The purpose of this study is to analyze the relationship of physical activity and dietary pattern to the circadian pattern of blood pressure, central and peripheral blood pressure, pulse wave velocity, carotid intima-media thickness and biological markers of endothelial dysfunction in active and sedentary individuals without arteriosclerotic disease. Methods/Design Design: A cross-sectional multicenter study with six research groups. Subjects: From subjects of the PEPAF project cohort, in which 1,163 who were sedentary became active, 1,942 were sedentary and 2,346 were active. By stratified random sampling, 1,500 subjects will be included, 250 in each group. Primary measurements: We will evaluate height, weight, abdominal circumference, clinical and ambulatory blood pressure with the Radial Pulse Wave Acquisition Device (BPro), central blood pressure and augmentation index with Pulse Wave Application Software (A-Pulse) and SphymgoCor System Px (Pulse Wave Analysis), pulse wave velocity (PWV) with SphymgoCor System Px (Pulse Wave Velocity), nutritional pattern with a food intake frequency questionnaire, physical activity with the 7-day PAR questionnaire and accelerometer (Actigraph GT3X), physical fitness with the cycle ergometer (PWC-170), carotid intima-media thickness by ultrasound (Micromax), and endothelial dysfunction biological markers (endoglin and osteoprotegerin). Discussion Determining that sustained physical activity and the change from sedentary to active as well as a healthy diet improve circadian pattern, arterial elasticity and carotid intima-media thickness may help to propose lifestyle intervention programs. These interventions could improve the cardiovascular risk profile in some parameters not routinely assessed with traditional risk scales. From the results of this study, interventional approaches could be obtained to delay vascular aging that combine physical exercise and diet

  18. Podoplanin is a component of extracellular vesicles that reprograms cell-derived exosomal proteins and modulates lymphatic vessel formation

    PubMed Central

    Andrés, Germán; Gopal, Shashi K.; Martín-Villar, Ester; Renart, Jaime; Simpson, Richard J.; Quintanilla, Miguel

    2016-01-01

    Podoplanin (PDPN) is a transmembrane glycoprotein that plays crucial roles in embryonic development, the immune response, and malignant progression. Here, we report that cells ectopically or endogenously expressing PDPN release extracellular vesicles (EVs) that contain PDPN mRNA and protein. PDPN incorporates into membrane shed microvesicles (MVs) and endosomal-derived exosomes (EXOs), where it was found to colocalize with the canonical EV marker CD63 by immunoelectron microscopy. We have previously found that expression of PDPN in MDCK cells induces an epithelial-mesenchymal transition (EMT). Proteomic profiling of MDCK-PDPN cells compared to control cells shows that PDPN-induced EMT is associated with upregulation of oncogenic proteins and diminished expression of tumor suppressors. Proteomic analysis of exosomes reveals that MDCK-PDPN EXOs were enriched in protein cargos involved in cell adhesion, cytoskeletal remodeling, signal transduction and, importantly, intracellular trafficking and EV biogenesis. Indeed, expression of PDPN in MDCK cells stimulated both EXO and MV production, while knockdown of endogenous PDPN in human HN5 squamous carcinoma cells reduced EXO production and inhibited tumorigenesis. EXOs released from MDCK-PDPN and control cells both stimulated in vitro angiogenesis, but only EXOs containing PDPN were shown to promote lymphatic vessel formation. This effect was mediated by PDPN on the surface of EXOs, as demonstrated by a neutralizing specific monoclonal antibody. These results contribute to our understanding of PDPN-induced EMT in association to tumor progression, and suggest an important role for PDPN in EV biogenesis and/or release and for PDPN-EXOs in modulating lymphangiogenesis. PMID:26893367

  19. Identification and expression of troponin T, a new marker on the surface of cultured tumor endothelial cells by aptamer ligand

    PubMed Central

    Ara, Mst Naznin; Hyodo, Mamoru; Ohga, Noritaka; Akiyama, Kosuke; Hida, Kyoko; Hida, Yasuhiro; Shinohara, Nobuo; Harashima, Hideyoshi

    2014-01-01

    The identification of a specific biomarker involves the development of new clinical diagnostic tools, and an in-depth understanding of the disease at the molecular level. When new blood vessels form in tumor cells, endothelial cell production is induced, a process that plays a key role in disease progression and metastasis to distinct organs for solid tumor types. The present study reports on the identification of a new biomarker on primary cultured mouse tumor endothelial cells (mTECs) using our recently developed high-affinity DNA aptamer AraHH001 (Kd = 43 nmol/L) assisted proteomics approach. We applied a strategy involving aptamer-facilitated biomarker discovery. Biotin-tagged AraHH001 was incubated with lysates of mTECs and the aptamer-proteins were then conjugated with streptavidin magnetic beads. Finally, the bound proteins were separated by sodiumdodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) with silver staining. We identified troponin T via matrix assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry, the molecular target of aptamer AraHH001, and its presence was confirmed by measuring mRNA, protein levels, western blot, immunostaining, a gel shift assay of AraHH001 with troponin T. We first report here on the discovery of troponin T on mTECs, a promising and interesting diagnostic tool in the development of antiangiogenic therapy techniques the involves the targeting of the tumor vasculature. PMID:24810801

  20. Antiglycopeptide Mouse Monoclonal Antibody LpMab-21 Exerts Antitumor Activity Against Human Podoplanin Through Antibody-Dependent Cellular Cytotoxicity and Complement-Dependent Cytotoxicity.

    PubMed

    Kato, Yukinari; Kunita, Akiko; Fukayama, Masashi; Abe, Shinji; Nishioka, Yasuhiko; Uchida, Hiroaki; Tahara, Hideaki; Yamada, Shinji; Yanaka, Miyuki; Nakamura, Takuro; Saidoh, Noriko; Yoshida, Kanae; Fujii, Yuki; Honma, Ryusuke; Takagi, Michiaki; Ogasawara, Satoshi; Murata, Takeshi; Kaneko, Mika K

    2017-02-01

    The interaction between podoplanin (PDPN) and C-type lectin-like receptor 2 (CLEC-2) is involved in tumor malignancy. We have established many monoclonal antibodies (mAbs) against human podoplanin using the cancer-specific mAb (CasMab) technology. LpMab-21, one of the mouse antipodoplanin mAbs, is of the IgG2a subclass, and its minimum epitope was determined to be Thr76-Arg79 of the human podoplanin. Importantly, sialic acid is linked to Thr76; therefore, LpMab-21 is an antiglycopeptide mAb (GpMab). In this study, we investigated whether LpMab-21 shows antibody-dependent cellular cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC) against human podoplanin-expressing cancer cell lines in vitro and also studied its antitumor activities using a xenograft model. LpMab-21 showed high ADCC and CDC activities against not only podoplanin-expressing Chinese hamster ovary cells but also LN319 glioblastoma cells and PC-10 lung cancer cells, both of which endogenously express podoplanin. Furthermore, LpMab-21 decreased tumor growth in vivo, indicating that LpMab-21 could be useful for antibody therapy against human podoplanin-expressing cancers.

  1. Association of Urinary N-Domain Angiotensin I-Converting Enzyme with Plasma Inflammatory Markers and Endothelial Function

    PubMed Central

    Fernandes, Fernanda B; Plavnik, Frida L; Teixeira, Andressa MS; Christofalo, Dejaldo MJ; Ajzen, Sergio A; Higa, Elisa MS; Ronchi, Fernanda A; Sesso, Ricardo CC; Casarini, Dulce E

    2008-01-01

    The aim of this study was to investigate the association between urinary 90 kDa N-domain Angiotensin I-converting enzyme (ACE) form with C-reactive protein (CRP) and homocysteine plasma levels (Hcy), urinary nitric oxide (NOu), and endothelial function (EF) in normotensive subjects. Forty healthy subjects were evaluated through brachial Doppler US to test the response to reactive hyperemia and a panel of blood tests to determine CRP and Hcy levels, NOu, and urinary ACE. They were divided into groups according to the presence (ACE90+) or absence (ACE90–) of the 90 kDa ACE, the presence (FH+) or absence (FH–) of family history of hypertension, and the presence or absence of these two variables FH+/ACE90+ and FH–/ACE90–. We found an impaired endothelial dilatation in subjects who presented the 90 kDa N-domain ACE as follows: 11.4% ± 5.3% in ACE90+ compared with 17.6% ± 7.1% in ACE90– group and 12.4% ± 5.6% in FH+/ACE90+ compared with 17.7% ± 6.2% in FH–/ACE90– group, P < 0.05. Hcy and CRP levels were statistically significantly lower in FH+/ACE90+ than in FH–/ACE90– group, as follows: 10.0 ± 2.3 μM compared with 12.7 ± 1.5 μM, and 1.3 ± 1.8 mg/L compared with 3.6 ± 2.0 mg/L, respectively. A correlation between flow-mediated dilatation (FMD) and CRP, Hcy, and NOu levels was not found. Our study suggests a reduction in the basal NO production confirmed by NOu analysis in subjects with the 90 kDa N-domain ACE isoform alone or associated with a family history of hypertension. Our data suggest that the presence of the 90 kDa N-domain ACE itself may have a negative impact on flow-mediated dilatation stimulated by reactive hyperemia. PMID:18475311

  2. Circulating and progenitor endothelial cells are abnormal in patients with different types of von Willebrand disease and correlate with markers of angiogenesis.

    PubMed

    Gritti, Giuseppe; Cortelezzi, Agostino; Bucciarelli, Paolo; Rezzonico, Francesca; Lonati, Silvia; La Marca, Silvia; Silvestris, Ilaria; Federici, Augusto B

    2011-08-01

    von Willebrand disease (VWD) is the most common inherited bleeding disorder and is caused by quantitative or qualitative defects of von Willebrand factor (VWF). VWF, synthesized by endothelium and megakaryocytes (MK), circulates in plasma and is present in subendothelium and platelets. Circulating endothelial cells (CEC) and progenitor endothelial cells (EPC) have been recently proposed as markers of peripheral and bone marrow-derived angiogenesis. To evaluate the association of CEC/EPC with known inherited defects of cellular and circulating VWF, we have measured the number of CEC/EPC together with cytokines involved in angiogenesis in different VWD types. A group of 74 patients was composed by the following VWD types: VWD1 (n = 22), VWD2A (n = 9), VWD2B (n = 19), VWD2M (n = 17), and VWD3 (n = 7). Healthy individuals (n = 20) were used as controls. CEC (CD146(+) , CD31(+) , and CD45(-) ) and EPC (CD34(+) , CD133(+) , and CD45(-) ) were evaluated by flow cytometry. Circulating serum levels of VEGF, E-selectin, P-selectin, EPO, and TPO were determined by ELISA. CEC, VEGF, E-selectin, and EPO were higher and EPC lower in VWD patients than in controls (P < 0.01). Among the five groups of VWD patients and controls, a significant difference was found for CEC (one-way ANOVA: P = 0.005), EPC (P = 0.001), E-Selectin (P < 0.0001), EPO (P = 0.021), and TPO (P = 0.004): the latter was high in VWD3 patients. In VWD1, we found an inverse relationship between CEC and VWF:Ag levels (P = 0.048; R(2) = 0.19). Based on these data, CEC are increased in VWD and are associated with the high levels of cytokines involved in angiogenesis (up-regulation). EPC are decreased, suggesting down-regulation of bone marrow-derived angiogenesis in VWD.

  3. Comparison of cardiovascular disease risk markers in HIV-infected patients receiving abacavir and tenofovir: the nucleoside inflammation, coagulation and endothelial function (NICE) study

    PubMed Central

    Wohl, David A; Arnoczy, Gretchen; Fichtenbaum, Carl J; Campbell, Thomas; Taiwo, Babafemi; Hicks, Charles; McComsey, Grace A; Koletar, Susan; Sax, Paul; Tebas, Pablo; Ha, Belinda; Massengale, Kelly; Walsh, Kendall; Stein, James H

    2015-01-01

    Background The association between abacavir (ABC) and cardiovascular disease (CVD) risk in HIV-infected individuals is unclear. Putative mechanisms for an effect of ABC on CVD risk including endothelial dysfunction have been proposed; however, a biological mechanism has not been established. Methods This was a cross-sectional study of HIV-infected subjects with HIV RNA levels <400 copies/ml, who were randomly assigned to ABC or tenofovir (TDF) as initial therapy during a prior clinical trial. A small cohort of subjects on zidovudine (AZT; not randomly assigned) were studied to explore long-term exposure to this agent. All underwent brachial artery ultrasound for flow-mediated dilation (FMD), and D-dimer, high-sensitivity C-reactive protein (hsCRP), interleukin-6 (IL-6) and fasting lipids were measured. Between-arm differences were evaluated by multivariable linear or logistic regression modelling. Results There were 148 subjects (46 on ABC, 72 on TDF and 30 on AZT). Demographic characteristics were balanced across the groups except, as expected, AZT-treated participants were older, had higher CD4+ T-cell counts, and longer antiretroviral therapy duration. After adjusting for age, brachial artery diameter, and treatment duration, FMD was similar in those on ABC (3.9%) and TDF (5.4%; P=0.181). FMD was higher in those on AZT (6.1%; P<0.005). Levels of IL-6, hsCRP and detectable D-dimer were similar between groups. Conclusions Among individuals assigned to ABC or TDF in randomized clinical trials there were no significant differences in FMD or markers of inflammation and coagulation. Whether ABC contributes to risk of CVD remains unclear, but our results suggest that endothelial dysfunction, heightened inflammation, and altered coagulation are unlikely to be mechanisms by which the drug could increase CVD risk above that seen with TDF. PMID:23985706

  4. Weight loss, but not fish oil consumption, improves fasting and postprandial serum lipids, markers of endothelial function, and inflammatory signatures in moderately obese men.

    PubMed

    Plat, Jogchum; Jellema, Annemarie; Ramakers, Julian; Mensink, Ronald P

    2007-12-01

    Overweight persons are at risk for cardiovascular diseases, which may relate to a disturbed endothelial function and pro-inflammatory serum profiles. Indeed, weight loss lowers cardiovascular disease risk, but weight maintenance is difficult. Therefore, dietary supplements such as fish oil, which improve endothelial function, are useful. In this study, we evaluated effects of fish oil and moderate weight loss in the same population. For this, 11 normolipidemic healthy, moderately obese men (BMI 30-35 kg/m2) received in random order 1.1 g/d eicosapentanoic acid (EPA) + docosahexanoic acid (DHA) or oleic acid (control) for 6 wk. In the 3rd period, 8 of the 11 subjects consumed low-energy diets (2 MJ/d) for 4 wk followed by 4 wk weight stabilization. Their body weight was reduced by 9.4 +/- 2.0 kg (P < 0.05). On the final day of all 3 periods, a postprandial test was conducted. Weight loss lowered fasting and postprandial plasma triacylglycerol (TG) responses (P < 0.001), whereas fish oil reduced only postprandial TG (P = 0.006). Fish oil did not affect soluble intercellular adhesion molecule (s-ICAM), whereas weight loss reduced fasting (P = 0.009) and postprandial s-ICAM responses (P < 0.001). Fasting s-ICAM and TG correlated (r = 0.68; P = 0.029), as did changes in fasting s-ICAM and TG during weight loss (r = 0.80; P = 0.029) and fish oil treatment (r = 0.76; P = 0.009). Fasting (P = 0.027) and postprandial (P < 0.001) serum C-reactive protein were lowered by weight loss. The postprandial monocyte chemoattractant protein-1 response was lowered by fish oil and after weight loss (P < 0.001). This indicates that 1.1 g/d EPA+DHA supplied for 6 wk, in contrast to approximately 10 kg weight loss, does not improve markers of endothelial function and inflammation.

  5. The impact of decreases in air temperature and increases in ozone on markers of endothelial function in individuals having type-2 diabetes

    EPA Science Inventory

    Several studies have reported an association between air pollution and endothelial dysfunction, especially in individuals having diabetes. However, very few studies have examined the impact of air temperature on endothelial function. The objective of this analysis was to investig...

  6. Podoplanin expression in tumor-free resection margins of oral squamous cell carcinomas: an immunohistochemical and fractal analysis study.

    PubMed

    Margaritescu, C; Raica, M; Pirici, D; Simionescu, C; Mogoanta, L; Stinga, A C; Stinga, A S; Ribatti, D

    2010-06-01

    Podoplanin is involved in tumorigenesis and cancer progression in head and neck malignancies and its expression is not restricted to lymphatic vessel endothelium. The aim of this study was to establish podoplanin expression in the tumor-free resection margins of oral squamous cell carcinomas (OSCCs) and to evaluate the geometric complexity of the lymphatic vessels in oral mucosa by utilizing fractal analysis. As concerns the podoplanin expression in noncancerous tissue, forty tumor-free resection margins from OSCCs were investigated utilizing immunohistochemistry for D2-40 antibody and image densitometry analysis. Podoplanin expression was extremely low in basal cells, especially in resection margins of OSCCs developed in the lower lip regions. However, a highly variable D2-40 expression in tumor-free resection margins associated with hyperplastic or dysplastic lesions was identified. Moreover, podoplanin expression also extended to the basal layer of the lower lip skin appendages, the myoepithelial cells of acini and ducts of minor salivary glands, and other structures from the oral cavity. As concerns the study of the density and complexity of oral lymphatic vessels architecture by means of immunohistochemistry (D2-40, CD31 and Ki-67 antibodies) and fractal analysis, we demonstrated that in normal oral mucosa the geometry of the lymphatic vessels was less complex at the level of the lower lip compared to the anterior part of the oral floor mucosa or the tongue. A comparative analysis between the normal and pathological aspects revealed statistically significant differences between the fractal dimension (FD) of the vessels' outline, especially in the tongue. Fractal analysis proved an increasing lymphatic network complexity from normal to premalignant oral mucosal lesions, providing additional prognostic information in oral malignant tumors.

  7. Detection of Autoantibodies to Vascular Endothelial Growth Factor Receptor-3 in Bile Duct Ligated Rats and Correlations with a Panel of Traditional Markers of Liver Diseases

    PubMed Central

    Duval, Florent; Cruz-Vega, Delia Elva; González-Gamboa, Ivonne; González-Garza, María Teresa; Ponz, Fernando; Sánchez, Flora; Alarcón-Galván, Gabriela; Moreno-Cuevas, Jorge E.

    2016-01-01

    There is a need for new noninvasive biomarkers (NIBMs) able to assess cholestasis and fibrosis in chronic cholestatic liver diseases (CCLDs). Tumorigenesis can arise from CCLDs. Therefore, autoantibodies to tumor-associated antigens (TAA) may be early produced in response to abnormal self-antigen expression caused by cholestatic injury. Vascular endothelial growth factor receptor-3 (VEGFR-3) has TAA potential since it is involved in cholangiocytes and lymphatic vessels proliferations during CCLDs. This study aims to detect autoantibodies directed at VEGFR-3 during bile duct ligation- (BDL-) induced cholestatic injury in rat sera and investigate whether they could be associated with traditional markers of liver damage, cholestasis, and fibrosis. An ELISA was performed to detect anti-VEGFR-3 autoantibodies in sera of rats with different degree of liver injury and results were correlated with aminotransferases, total bilirubin, and the relative fibrotic area. Mean absorbances of anti-VEGFR-3 autoantibodies were significantly increased from week one to week five after BDL. The highest correlation was observed with total bilirubin (R2 = 0.8450, P = 3.04e − 12). In conclusion, anti-VEGFR-3 autoantibodies are early produced during BDL-induced cholestatic injury, and they are closely related to cholestasis, suggesting the potential of anti-VEGFR-3 autoantibodies as NIBMs of cholestasis in CCLDs and justifying the need for further investigations in patients with CCLD. PMID:27212785

  8. Potential anti-inflammatory effects of maraviroc in HIV-positive patients: a pilot study of inflammation, endothelial dysfunction, and coagulation markers.

    PubMed

    Francisci, Daniela; Falcinelli, Emanuela; Baroncelli, Silvia; Petito, Eleonora; Cecchini, Enisia; Weimer, Liliana Elena; Floridia, Marco; Gresele, Paolo; Baldelli, Franco

    2014-06-01

    Persistent immune activation and chronic inflammation significantly contribute to non-AIDS morbidity in HIV-infected patients. The HIV inhibitor maraviroc (MVC) targets the cellular chemokine CCR5 HIV co-receptor, which is involved in important inflammatory pathways. MVC could have significant anti-inflammatory and anti-atherosclerotic effects, also reducing immune activation. We designed a pilot study to determine which plasma biomarkers of inflammation, endothelial dysfunction, and hypercoagulability were modified by MVC in 2 groups of 10 patients starting MVC-free or MVC-containing regimens. Ten age- and gender-matched healthy controls were also included. We found higher levels of all inflammatory biomarkers in HIV-infected patients compared to healthy controls. Both groups showed decreasing levels of interleukin (IL)-17, IL-10, and macrophage inflammatory protein (MIP)-1a following the achievement of viral suppression. Vascular cell adhesion molecule (VCAM)-1 levels were decreased in the MVC group and increased in the MVC-free group. In conclusion, some inflammatory biomarkers tend to decrease with the salvage regimen; MVC was not associated with a better impact on these measured markers.

  9. The effect of Citrus Aurantifolia (Lemon) peels on cardiometabolic risk factors and markers of endothelial function in adolescents with excess weight: A triple-masked randomized controlled trial

    PubMed Central

    Hashemipour, Mahin; Kargar, Maryam; Ghannadi, Alireza; Kelishadi, Roya

    2016-01-01

    Background: Childhood obesity is becoming a global problem and its incidence is increasing. The role of dietary intervention with fruits containing vitamin C and flavonoid to control obesity consequences in childhood has not been yet defined. Lemon (Citrus aurantifolia) peels contain flavonoid, pectin and vitamin C. We aimed to compare the effects of lemon peels and placebo on cardiometabolic risk factors and markers of endothelial function among adolescents with overweight and obesity. Methods: In this triple-masked, randomized controlled trial, 60 overweight/obese adolescents were enrolled in a 4-week trial. Eligible participants were randomly assigned into two groups of equal number receiving daily oral capsules containing lemon powder or placebo. Fasting blood sugar, lipid profile, ICAM-1 and VCAM-1, as well as systolic and diastolic blood pressure were compared between the two groups before and after administration of medication and placebo. Results: Of the total 60 enrolled patients, 30 and 29 patients in the lemon and control groups completed the study, respectively. The results of within-group analysis demonstrated a slight reduction in body mass index, LDL-C and systolic blood pressure in the lemon group, but no between group differences existed in the studied variables. Conclusion: This study revealed that consumption of lemon peel extract has some beneficial effects for childhood obesity; however, no considerable effect was documented on anthropometric measures and biochemical factors. Future studies with longer follow up are highly recommended. PMID:28210594

  10. The effect of Citrus Aurantifolia (Lemon) peels on cardiometabolic risk factors and markers of endothelial function in adolescents with excess weight: A triple-masked randomized controlled trial.

    PubMed

    Hashemipour, Mahin; Kargar, Maryam; Ghannadi, Alireza; Kelishadi, Roya

    2016-01-01

    Background: Childhood obesity is becoming a global problem and its incidence is increasing. The role of dietary intervention with fruits containing vitamin C and flavonoid to control obesity consequences in childhood has not been yet defined. Lemon (Citrus aurantifolia) peels contain flavonoid, pectin and vitamin C. We aimed to compare the effects of lemon peels and placebo on cardiometabolic risk factors and markers of endothelial function among adolescents with overweight and obesity. Methods: In this triple-masked, randomized controlled trial, 60 overweight/obese adolescents were enrolled in a 4-week trial. Eligible participants were randomly assigned into two groups of equal number receiving daily oral capsules containing lemon powder or placebo. Fasting blood sugar, lipid profile, ICAM-1 and VCAM-1, as well as systolic and diastolic blood pressure were compared between the two groups before and after administration of medication and placebo. Results: Of the total 60 enrolled patients, 30 and 29 patients in the lemon and control groups completed the study, respectively. The results of within-group analysis demonstrated a slight reduction in body mass index, LDL-C and systolic blood pressure in the lemon group, but no between group differences existed in the studied variables. Conclusion: This study revealed that consumption of lemon peel extract has some beneficial effects for childhood obesity; however, no considerable effect was documented on anthropometric measures and biochemical factors. Future studies with longer follow up are highly recommended.

  11. Markers of endothelial dysfunction and inflammation predict progression of diabetic nephropathy in African Americans with type 1 diabetes.

    PubMed

    Roy, Monique S; Janal, Malvin N; Crosby, Juan; Donnelly, Robert

    2015-02-01

    African Americans with early-onset type 1 diabetes mellitus are at a high risk for severe diabetic nephropathy and end-stage renal disease. In order to determine whether baseline plasma levels of inflammatory markers predict incidence of overt proteinuria or renal failure in African Americans with type 1 diabetes mellitus, we re-examined data of 356 participants in our observational follow-up study of 725 New Jersey African Americans with type 1 diabetes. At baseline and 6-year follow-up, a detailed structured clinical interview was conducted to document medical history including kidney dialysis or transplant, other diabetic complications, and renal-specific mortality. Plasma levels of 28 inflammatory biomarkers were measured using a multiplex bead analysis system. After adjusting for baseline age, glycohemoglobin, and other confounders, the baseline plasma levels of soluble intercellular adhesion molecule-1 (sICAM-1) in the upper two quartiles were, respectively, associated with a three- to fivefold increase in the risk of progression from no albuminuria or microalbuminuria to overt proteinuria. Baseline plasma levels of the chemokine eotaxin in the upper quartile were significantly associated with a sevenfold increase in risk of incident renal failure. These associations were independent of traditional risk factors for progression of diabetic nephropathy. Thus, in type 1 diabetic African Americans, sICAM-1 predicted progression to overt proteinuria and eotaxin-predicted progression to renal failure.

  12. The CLEC-2-podoplanin axis controls the contractility of fibroblastic reticular cells and lymph node microarchitecture.

    PubMed

    Astarita, Jillian L; Cremasco, Viviana; Fu, Jianxin; Darnell, Max C; Peck, James R; Nieves-Bonilla, Janice M; Song, Kai; Kondo, Yuji; Woodruff, Matthew C; Gogineni, Alvin; Onder, Lucas; Ludewig, Burkhard; Weimer, Robby M; Carroll, Michael C; Mooney, David J; Xia, Lijun; Turley, Shannon J

    2015-01-01

    In lymph nodes, fibroblastic reticular cells (FRCs) form a collagen-based reticular network that supports migratory dendritic cells (DCs) and T cells and transports lymph. A hallmark of FRCs is their propensity to contract collagen, yet this function is poorly understood. Here we demonstrate that podoplanin (PDPN) regulates actomyosin contractility in FRCs. Under resting conditions, when FRCs are unlikely to encounter mature DCs expressing the PDPN receptor CLEC-2, PDPN endowed FRCs with contractile function and exerted tension within the reticulum. Upon inflammation, CLEC-2 on mature DCs potently attenuated PDPN-mediated contractility, which resulted in FRC relaxation and reduced tissue stiffness. Disrupting PDPN function altered the homeostasis and spacing of FRCs and T cells, which resulted in an expanded reticular network and enhanced immunity.

  13. Markers of endothelial dysfunction and leucocyte activation in Saudi and non-Saudi haplotypes of sickle cell disease.

    PubMed

    Al Najjar, Salwa; Adam, Soheir; Ahmed, Nessar; Qari, Mohamed

    2017-01-01

    Sickle cell disease (SCD) is an autosomal recessive inherited hemoglobinopathy, characterized by chronic hemolysis and recurrent vaso-occlusive crisis (VOC). This study investigates changes in leucocyte subsets and the relationship between cell adhesion molecule expression and disease manifestations in patients during steady state and acute VOC. We compared soluble E-selectin and P-selectin levels in 84 SCD patients, in steady state and during VOC to 84 healthy controls. Using immunophenotyping, we also compared lymphocyte subsets in these three groups. Further, we compared E-selectin and P-selectin levels in patients of Saudi ethnicity to non-Saudi patients, in all three groups. Lymphocyte subsets showed high percentages of total T lymphocytes, T helper and suppressor lymphocytes, B lymphocytes as well as NK cells in patients with SCD during steady state, while B lymphocytes and NK cells were significantly higher during acute VOC crisis. High levels of both soluble E-selectin (sE-selectin) and soluble P-selectin (sP-selectin) markers were demonstrated in the serum of patients with SCD during both steady state and acute VOC. Levels of selectins were significantly higher in acute VOC. The immunophenotypic expression of L-selectin, on leucocytes, was high in SCD both during steady state and during acute VOC in comparison to normal control subjects. There was no significant difference in all three study groups between Saudi and non-Saudi patients. These findings suggest that patients with SCD have increased expression of adhesion molecules: E-selectin and P-selectin, which play an important role in the pathogenesis of VOC. Despite the distinct phenotype of Saudi patients with SCD, there was no significant difference in levels of soluble E-selectin and soluble P-selectin between Saudi and non-Saudi patients in all three groups. While sickle cell disease is a well-recognized state of chronic inflammation, the role of specific adhesion molecules is steadily unraveling

  14. Incorporation of podoplanin into HIV released from HEK-293T cells, but not PBMC, is required for efficient binding to the attachment factor CLEC-2

    PubMed Central

    2010-01-01

    Background Platelets are associated with HIV in the blood of infected individuals and might modulate viral dissemination, particularly if the virus is directly transmitted into the bloodstream. The C-type lectin DC-SIGN and the novel HIV attachment factor CLEC-2 are expressed by platelets and facilitate HIV transmission from platelets to T-cells. Here, we studied the molecular mechanisms behind CLEC-2-mediated HIV-1 transmission. Results Binding studies with soluble proteins indicated that CLEC-2, in contrast to DC-SIGN, does not recognize the viral envelope protein, but a cellular factor expressed on kidney-derived 293T cells. Subsequent analyses revealed that the cellular mucin-like membranous glycoprotein podoplanin, a CLEC-2 ligand, was expressed on 293T cells and incorporated into virions released from these cells. Knock-down of podoplanin in 293T cells by shRNA showed that virion incorporation of podoplanin was required for efficient CLEC-2-dependent HIV-1 interactions with cell lines and platelets. Flow cytometry revealed no evidence for podoplanin expression on viable T-cells and peripheral blood mononuclear cells (PBMC). Podoplanin was also not detected on HIV-1 infected T-cells. However, apoptotic bystander cells in HIV-1 infected cultures reacted with anti-podoplanin antibodies, and similar results were obtained upon induction of apoptosis in a cell line and in PBMCs suggesting an unexpected link between apoptosis and podoplanin expression. Despite the absence of detectable podoplanin expression, HIV-1 produced in PBMC was transmitted to T-cells in a CLEC-2-dependent manner, indicating that T-cells might express an as yet unidentified CLEC-2 ligand. Conclusions Virion incorporation of podoplanin mediates CLEC-2 interactions of HIV-1 derived from 293T cells, while incorporation of a different cellular factor seems to be responsible for CLEC-2-dependent capture of PBMC-derived viruses. Furthermore, evidence was obtained that podoplanin expression is

  15. HuR and podoplanin expression is associated with a high risk of malignant transformation in patients with oral preneoplastic lesions.

    PubMed

    Habiba, Umma; Kitamura, Tetsuya; Yanagawa-Matsuda, Aya; Higashino, Fumihiro; Hida, Kyoko; Totsuka, Yasunori; Shindoh, Masanobu

    2016-11-01

    The risk of malignant transformation in oral preneoplastic lesions (OPLs) is challenging to assess. The objective of the present study was to determine the expression of ELAV like RNA binding protein 1 (HuR) and podoplanin in OPLs, and to evaluate the use of each protein as biomarkers for the risk assessment of malignant transformations. Immunohistochemistry for HuR and podoplanin was performed on the tissues of 51 patients with OPL, including cases of low grade dysplasia (LGD) and high grade dysplasia (HGD). The association between the protein expression patterns and clinicopathological parameters, including oral cancer free survival (OCFS) time, was analyzed during the follow-up period. HuR and podoplanin expression was observed in 28 (55%) and 36 (71%) of 51 patients, respectively. Kaplan-Meier analysis showed that the expression of HuR and podoplanin was associated with the risk of progression to oral cancer (P<0.05). Multivariate analysis revealed that HuR and podoplanin expression was associated with a 2.93-fold (95% confidence interval (CI), 0.98-10.34; P=0.055) and 2.06-fold (95% CI, 0.55-8.01; P=0.283) increase in risk of malignant transformation, respectively. The risk of OPL malignant transformation was considerably increased with the coexpression of HuR and podoplanin compared with the histological grading (95% CI, 1.64-23.59; P=0.005). The results of the present study demonstrated that the expression of HuR and podoplanin associates with malignant transformation and suggests that the proteins may be used as biomarkers to identify OPL patients with an increased risk of cancer development.

  16. HuR and podoplanin expression is associated with a high risk of malignant transformation in patients with oral preneoplastic lesions

    PubMed Central

    Habiba, Umma; Kitamura, Tetsuya; Yanagawa-Matsuda, Aya; Higashino, Fumihiro; Hida, Kyoko; Totsuka, Yasunori; Shindoh, Masanobu

    2016-01-01

    The risk of malignant transformation in oral preneoplastic lesions (OPLs) is challenging to assess. The objective of the present study was to determine the expression of ELAV like RNA binding protein 1 (HuR) and podoplanin in OPLs, and to evaluate the use of each protein as biomarkers for the risk assessment of malignant transformations. Immunohistochemistry for HuR and podoplanin was performed on the tissues of 51 patients with OPL, including cases of low grade dysplasia (LGD) and high grade dysplasia (HGD). The association between the protein expression patterns and clinicopathological parameters, including oral cancer free survival (OCFS) time, was analyzed during the follow-up period. HuR and podoplanin expression was observed in 28 (55%) and 36 (71%) of 51 patients, respectively. Kaplan-Meier analysis showed that the expression of HuR and podoplanin was associated with the risk of progression to oral cancer (P<0.05). Multivariate analysis revealed that HuR and podoplanin expression was associated with a 2.93-fold (95% confidence interval (CI), 0.98–10.34; P=0.055) and 2.06-fold (95% CI, 0.55–8.01; P=0.283) increase in risk of malignant transformation, respectively. The risk of OPL malignant transformation was considerably increased with the coexpression of HuR and podoplanin compared with the histological grading (95% CI, 1.64–23.59; P=0.005). The results of the present study demonstrated that the expression of HuR and podoplanin associates with malignant transformation and suggests that the proteins may be used as biomarkers to identify OPL patients with an increased risk of cancer development. PMID:27899983

  17. Short communication: Effect of supplementation with Lactobacillus casei Shirota on insulin sensitivity, β-cell function, and markers of endothelial function and inflammation in subjects with metabolic syndrome--a pilot study.

    PubMed

    Tripolt, N J; Leber, B; Blattl, D; Eder, M; Wonisch, W; Scharnagl, H; Stojakovic, T; Obermayer-Pietsch, B; Wascher, T C; Pieber, T R; Stadlbauer, V; Sourij, H

    2013-01-01

    Based on animal studies, intake of probiotic bacteria was suggested to improve insulin sensitivity by reducing endotoxinemia and inflammation. The objective of this study was to determine the effects of supplementation with the probiotic strain Lactobacillus casei Shirota (LcS) over 12 wk on insulin sensitivity, β-cell function, inflammation, and endothelial dysfunction parameters in subjects with metabolic syndrome. In a randomized-controlled study, 30 subjects with metabolic syndrome either received Lactobacillus casei Shirota 3 times daily for 12 wk or served as controls with standard medical therapy. Fasting blood samples were taken and a 75-g oral glucose tolerance test was performed to derive indices for insulin sensitivity and β-cell function. In addition, parameters to assess endothelial function and inflammation markers were determined. Even though the insulin sensitivity index significantly improved after 3 mo of probiotic supplementation (0.058±0.021 vs. 0.038±0.025), the change was not significantly different compared with the control group. No improvements were seen in additional indices of insulin sensitivity (quantitative insulin sensitivity check index, insulin sensitivity by oral glucose tolerance test, and homeostasis model assessment for insulin resistance) and β-cell function (first and second phase insulin secretion, and homeostasis model assessment for β-cell function). Probiotic supplementation resulted in a significant reduction in soluble vascular cell adhesion molecule-1 (sVCAM-1) level (1,614±343 vs. 1,418±265 ng/mL). No significant changes in parameters used to assess low-grade inflammation or endothelial dysfunction were observed. Intake of LcS for 12 wk in subjects with metabolic syndrome did not improve insulin sensitivity, β-cell function, endothelial function, or inflammation markers in this trial.

  18. Platelets Promote Tumor Growth and Metastasis via Direct Interaction between Aggrus/Podoplanin and CLEC-2

    PubMed Central

    Takagi, Satoshi; Sato, Shigeo; Oh-hara, Tomoko; Takami, Miho; Koike, Sumie; Mishima, Yuji; Hatake, Kiyohiko; Fujita, Naoya

    2013-01-01

    The platelet aggregation-inducing factor Aggrus, also known as podoplanin, is frequently upregulated in several types of tumors and enhances hematogenous metastasis by interacting with and activating the platelet receptor CLEC-2. Thus, Aggrus–CLEC-2 binding could be a therapeutic molecular mechanism for cancer therapy. We generated a new anti-human Aggrus monoclonal antibody, MS-1, that suppressed Aggrus–CLEC-2 binding, Aggrus-induced platelet aggregation, and Aggrus-mediated tumor metastasis. Interestingly, the MS-1 monoclonal antibody attenuated the growth of Aggrus-positive tumors in vivo. Moreover, the humanized chimeric MS-1 antibody, ChMS-1, also exhibited strong antitumor activity against Aggrus-positive lung squamous cell carcinoma xenografted into NOD-SCID mice compromising antibody-dependent cellular cytotoxic and complement-dependent cytotoxic activities. Because Aggrus knockdown suppressed platelet-induced proliferation in vitro and tumor growth of the lung squamous cell carcinoma in vivo, Aggrus may be involved in not only tumor metastasis but also tumor growth by promoting platelet-tumor interaction, platelet activation, and secretion of platelet-derived factors in vivo. Our results indicate that molecular target drugs inhibiting specific platelet–tumor interactions can be developed as antitumor drugs that suppress both metastasis and proliferation of tumors such as lung squamous cell carcinoma. PMID:23991201

  19. Expression of Aggrus/podoplanin in bladder cancer and its role in pulmonary metastasis

    PubMed Central

    Takagi, Satoshi; Oh-hara, Tomoko; Sato, Shigeo; Gong, Bo; Takami, Miho; Fujita, Naoya

    2014-01-01

    Platelet aggregation-inducing factor Aggrus, also known as podoplanin, is associated with tumor malignancy by promoting hematogenous metastasis. Aggrus overexpression has been reported in some tumor tissues including lung, esophagus, head and neck and brain. We here found the frequent upregulation of aggrus mRNA in urinary bladder cancers using cancer tissue panels from various organs. Immunohistochemical analysis confirmed Aggrus protein expression in urinary bladder cancers and suggested a positive correlation between Aggrus expression and metastatic tendency in bladder cancers. Endogenous expression of Aggrus protein on the cell surface was found in the mouse bladder cancer MBT-2 cell line and human bladder cancer SCaBER cell lines. Knockdown of Aggrus expression in MBT-2 cells decreased their ability to induce platelet aggregation and form pulmonary metastasis in syngeneic mouse models. Knockdown of Aggrus expression in the human bladder cancer SCaBER cells also attenuated their ability to induce platelet aggregation and form pulmonary metastasis in mice. Moreover, pulmonary metastasis of SCaBER cells was prevented by prior administration of our generated anti-Aggrus neutralizing monoclonal antibodies by attenuating their retention in lung. These results indicate that Aggrus plays an important role in bladder cancer metastasis. Thus, anti-Aggrus neutralizing antibodies would be useful for the prevention of hematogenous metastasis of Aggrus-positive bladder cancer. PMID:24222607

  20. CXCR4 pos circulating progenitor cells coexpressing monocytic and endothelial markers correlating with fibrotic clinical features are present in the peripheral blood of patients affected by systemic sclerosis.

    PubMed

    Campioni, Diana; Lo Monaco, Andrea; Lanza, Francesco; Moretti, Sabrina; Ferrari, Luisa; Fotinidi, Maria; La Corte, Renato; Cuneo, Antonio; Trotta, Francesco

    2008-08-01

    There is still controversy regarding the role of circulating endothelial and progenitor cells (CECs/CEPs) in the pathogenesis of systemic sclerosis (SSc). Using a sequential Boolean gating strategy based on a 4-color flow cytometric protocol, an increased number of CD31(pos)/CD184(pos)(CXCR4)/CD34(pos)/CD45(pos) and CD31(pos)/CD117(pos) (c-kit-R) /CD34(pos)/ CD45(pos) hematopoietic circulating progenitor cells (HCPCs) was detected in SSc patients compared with healthy subjects. In SSc, no circulating mature and progenitor endothelial cells were observed, while an enhanced generation of erythroid progenitor cells was found to be correlated with the presence of CD117+ HCPCs. The presence of freshly detected CXCR4posHCPC was correlated either to the in vitro cultured spindle-shaped endothelial like cells (SELC) with an endo/myelomonocytic profile or to SDF-1 and VEGF serum level. These data are related to more fibrotic clinical features of the disease, thus supporting a possible role of these cells in fibrosis.

  1. Single automated donor plateletpheresis increases the plasma level of proinflammatory cytokine tumor necrosis factor-alpha which does not associate with endothelial release markers von Willebrand factor and fibronectin.

    PubMed

    Karadoğan, I; Ozdoğan, M; Undar, L

    2000-12-01

    The effect of plateletpheresis on endothelium, which has strong effects on blood coagulation, fibrinolysis and platelet function, is not known. Activation of leukocytes and subsequent generation of proinflammatory cytokines during the extracorporeal circulation may activate the endothelium. To test this hypothesis we measured plasma levels of tumor necrosis factor (TNF)-alpha as a prototype of the proinflammatory cytokines, and von Willebrand factor (vWF) and fibronectin as endothelial release/damage markers before and after a single plateletpheresis procedure on an intermittent-flow machine Haemonetics MCS 3p in 17 healthy donors. We found a significant increase in median plasma level of TNF-alpha following plateletpheresis (3.5 vs 26.5 pg/ml, P=0.02). Such increases in vWF and fibronectin were not observed. The increase in plasma TNF-alpha indicates that a single plateletpheresis procedure causes leukocyte activation which does not seemingly impair endothelial cell function. The relation of plateletpheresis-induced proinflammatory cytokine release to some adverse effects observed in both donors and recipients, and the effect of repeated plateletpheresis on endothelium deserve further studies.

  2. The role of vascular endothelial growth factor gene as the genetic marker of atherothrombotic disorders and in the gene therapy of coronary artery disease.

    PubMed

    Petrovic, Daniel

    2010-01-01

    Many human diseases are characterized by vasculature disorders. Out of the many players in the angiogenic network, the vascular endothelial growth genes are by far the best characterized. The vascular endothelial growth factor (VEGF) has been implicated in the pathogenesis of coronary artery disease (CAD) and in its complication, the acute myocardial infarction (AMI). Several common polymorphisms in the promoter region of the VEGF gene have been reported, but only few single nucleotide polymorphisms (SNPs) have been demonstrated to be associated with variations in VEGF serum concentrations and with a susceptibility to CAD and its complications-acute coronary syndromes. Moreover, the -634 C/G VEGF SNP (rs2010963) has been demonstrated to be associated with AMI and the development of heart failure after AMI. Gene-based therapy for patients with refractory CAD has been the subject of extensive investigation. Preclinical studies have shown promise for the delivery of VEGF gene for treating CAD, whereas the results of randomized placebo-controlled trials have not demonstrated unequivocal evidence of efficacy. To conclude, at present the role of VEGF and VEGF SNPs in pathogenesis of AMI and the development of heart failure after AMI is still uncertain and remains to be determined. Obviously, larger studies as well as functional studies are needed to confirm the role of VEGF SNPs in AMI and its complications after AMI.

  3. Vascular Endothelial Growth Factor Gene Polymorphism (rs2010963) and Its Receptor, Kinase Insert Domain-Containing Receptor Gene Polymorphism (rs2071559), and Markers of Carotid Atherosclerosis in Patients with Type 2 Diabetes Mellitus

    PubMed Central

    Merlo, Sebastjan; Starčević, Jovana Nikolajević; Mankoč, Sara; Šantl Letonja, Marija; Cokan Vujkovac, Andreja; Zorc, Marjeta; Petrovič, Daniel

    2016-01-01

    Background. The current study was designed to reveal possible associations between the polymorphisms of the vascular endothelial growth factor (VEGF) gene (rs2010963) and its receptor, kinase insert domain-containing receptor (KDR) gene polymorphism (rs2071559), and markers of carotid atherosclerosis in patients with type 2 diabetes mellitus (T2DM). Patients and Methods. 595 T2DM subjects and 200 control subjects were enrolled. The carotid intima-media thickness (CIMT) and plaque characteristics (presence and structure) were assessed ultrasonographically. Biochemical analyses were performed using standard biochemical methods. Genotyping of VEGF/KDR polymorphisms (rs2010963, rs2071559) was performed using KASPar assays. Results. Genotype distributions and allele frequencies of the VEGF/KDR polymorphisms (rs2010963, rs2071559) were not statistically significantly different between diabetic patients and controls. In our study, we demonstrated an association between the rs2071559 of KDR and either CIMT or the sum of plaque thickness in subjects with T2DM. We did not, however, demonstrate any association between the tested polymorphism of VEGF (rs2010963) and either CIMT, the sum of plaque thickness, the number of involved segments, hsCRP, the presence of carotid plaques, or the presence of unstable carotid plaques. Conclusions. In the present study, we demonstrated minor effect of the rs2071559 of KDR on markers of carotid atherosclerosis in subjects with T2DM. PMID:26881237

  4. Vascular Endothelial Growth Factor Gene Polymorphism (rs2010963) and Its Receptor, Kinase Insert Domain-Containing Receptor Gene Polymorphism (rs2071559), and Markers of Carotid Atherosclerosis in Patients with Type 2 Diabetes Mellitus.

    PubMed

    Merlo, Sebastjan; Starčević, Jovana Nikolajević; Mankoč, Sara; Šantl Letonja, Marija; Cokan Vujkovac, Andreja; Zorc, Marjeta; Petrovič, Daniel

    2016-01-01

    Background. The current study was designed to reveal possible associations between the polymorphisms of the vascular endothelial growth factor (VEGF) gene (rs2010963) and its receptor, kinase insert domain-containing receptor (KDR) gene polymorphism (rs2071559), and markers of carotid atherosclerosis in patients with type 2 diabetes mellitus (T2DM). Patients and Methods. 595 T2DM subjects and 200 control subjects were enrolled. The carotid intima-media thickness (CIMT) and plaque characteristics (presence and structure) were assessed ultrasonographically. Biochemical analyses were performed using standard biochemical methods. Genotyping of VEGF/KDR polymorphisms (rs2010963, rs2071559) was performed using KASPar assays. Results. Genotype distributions and allele frequencies of the VEGF/KDR polymorphisms (rs2010963, rs2071559) were not statistically significantly different between diabetic patients and controls. In our study, we demonstrated an association between the rs2071559 of KDR and either CIMT or the sum of plaque thickness in subjects with T2DM. We did not, however, demonstrate any association between the tested polymorphism of VEGF (rs2010963) and either CIMT, the sum of plaque thickness, the number of involved segments, hsCRP, the presence of carotid plaques, or the presence of unstable carotid plaques. Conclusions. In the present study, we demonstrated minor effect of the rs2071559 of KDR on markers of carotid atherosclerosis in subjects with T2DM.

  5. Effects of high-intensity interval training and moderate-intensity continuous training on endothelial function and cardiometabolic risk markers in obese adults.

    PubMed

    Sawyer, Brandon J; Tucker, Wesley J; Bhammar, Dharini M; Ryder, Justin R; Sweazea, Karen L; Gaesser, Glenn A

    2016-07-01

    We hypothesized that high-intensity interval training (HIIT) would be more effective than moderate-intensity continuous training (MICT) at improving endothelial function and maximum oxygen uptake (V̇o2 max) in obese adults. Eighteen participants [35.1 ± 8.1 (SD) yr; body mass index = 36.0 ± 5.0 kg/m(2)] were randomized to 8 wk (3 sessions/wk) of either HIIT [10 × 1 min, 90-95% maximum heart rate (HRmax), 1-min active recovery] or MICT (30 min, 70-75% HRmax). Brachial artery flow-mediated dilation (FMD) increased after HIIT (5.13 ± 2.80% vs. 8.98 ± 2.86%, P = 0.02) but not after MICT (5.23 ± 2.82% vs. 3.05 ± 2.76%, P = 0.16). Resting artery diameter increased after MICT (3.68 ± 0.58 mm vs. 3.86 ± 0.58 mm, P = 0.02) but not after HIIT (4.04 ± 0.70 mm vs. 4.09 ± 0.70 mm; P = 0.63). There was a significant (P = 0.02) group × time interaction in low flow-mediated constriction (L-FMC) between MICT (0.63 ± 2.00% vs. -2.79 ± 3.20%; P = 0.03) and HIIT (-1.04 ± 4.09% vs. 1.74 ± 3.46%; P = 0.29). V̇o2 max increased (P < 0.01) similarly after HIIT (2.19 ± 0.65 l/min vs. 2.64 ± 0.88 l/min) and MICT (2.24 ± 0.48 l/min vs. 2.55 ± 0.61 l/min). Biomarkers of cardiovascular risk and endothelial function were unchanged. HIIT and MICT produced different vascular adaptations in obese adults, with HIIT improving FMD and MICT increasing resting artery diameter and enhancing L-FMC. HIIT required 27.5% less total exercise time and ∼25% less energy expenditure than MICT.

  6. Evaluation of the Effects of Mesoglycan on Some Markers of Endothelial Damage and Walking Distance in Diabetic Patients with Peripheral Arterial Disease

    PubMed Central

    Giuseppe, Derosa; Angela, D’Angelo; Romano, Davide; Pamela, Maffioli

    2017-01-01

    The aim of this study was to evaluate the variation of some parameters involved in peripheral artery disease progression in diabetic patients with peripheral artery disease after six months of mesoglycan. We enrolled 64 Caucasian, type 2 diabetic patients, with stage IIa peripheral artery disease. They were randomized to mesoglycan (Prisma®), 50 mg twice a day, or placebo, for six months. We evaluated: glycemic control, metalloproteinase-2, and -9 (MMP-2, and -9), soluble intercellular adhesion molecule-1 (sICAM-1), soluble vascular cell adhesion protein-1 (sVCAM-1), interleukin-6 (IL-6), soluble E-selectin (sE-selectin), high sensitivity C-reactive protein (hs-CRP), tumor necrosis factor-α (TNF-α), and plasminogen activator inhibitor-1 (PAI-1). We recorded a decrease of MMP-2, MMP-9, sE-selectin, TNF-α, sVCAM-1, and IL-6 compared to baseline, and to placebo in the group treated with mesoglycan. Regarding sICAM-1, and hs-CRP, instead, we recorded a decrease with mesoglycan only compared to baseline. Preliminary results seem to suggest an improvement of pain free walking distance with mesoglycan in 18 patients both compared to baseline and to placebo, even if data should be taken cautiously. Our study showed that supplementation with mesoglycan improved endothelial dysfunction in type 2 diabetic patients with peripheral artery disease. Regarding the preliminary data suggesting also a slight improvement of clinical parameters such as pain free walking distance, more data and a bigger sample of patients are necessary to better verify this aspect. PMID:28272312

  7. [The vascular endothelial growth factor (VEGF): a model of gene regulation and a marker of tumour aggressiveness. An obvious therapeutic target?].

    PubMed

    Grépin, Renaud; Pagès, Gilles

    2009-01-01

    VEGF represents a model of gene expression regulation. RAS/RAF/MEK/ERK and PI3 Kinase pathways, activated in response to growth factors stimulation or by oncogenes, contribute to its expression by activating transcription factors or inactivating proteins implicated in degradation of its mRNA. These factors (Sp1/Sp3, HIF-1 and TTP) constitute molecular markers of tumor aggressiveness. VEGF is overexpressed in solid or hematologic tumors. Thus, numerous compounds regulating angiogenesis by targeting VEGF have been developed. However, their effects are not as spectacular as expected. The existence of anti-angiogenic isoforms of VEGF could be a cause of their less potent activity. These different points are discussed in this review article.

  8. Evaluation of Vascular Endothelial Growth Factor as a Prognostic Marker for Local Relapse in Early-Stage Breast Cancer Patients Treated With Breast-Conserving Therapy

    SciTech Connect

    Moran, Meena S.; Yang Qifeng; Goyal, Sharad; Harris, Lyndsay; Chung, Gina; Haffty, Bruce G.

    2011-12-01

    Purpose: Vascular endothelial growth factor (VEGF) is an important protein involved in the process of angiogenesis that has been found to correlate with relapse-free and overall survival in breast cancer, predominantly in locally advanced and metastatic disease. A paucity of data is available on the prognostic implications of VEGF in early-stage breast cancer; specifically, its prognostic value for local relapse after breast-conserving therapy (BCT) is largely unknown. The purpose of our study was to assess VEGF expression in a cohort of early-stage breast cancer patients treated with BCT and to correlate the clinical and pathologic features and outcomes with overexpression of VEGF. Methods and Materials: After obtaining institutional review board approval, the paraffin specimens of 368 patients with early-stage breast cancer treated with BCT between 1975 and 2005 were constructed into tissue microarrays with twofold redundancy. The tissue microarrays were stained for VEGF and read by a trained pathologist, who was unaware of the clinical details, as positive or negative according the standard guidelines. The clinical and pathologic data, long-term outcomes, and results of VEGF staining were analyzed. Results: The median follow-up for the entire cohort was 6.5 years. VEGF expression was positive in 56 (15%) of the 368 patients. Although VEGF expression did not correlate with age at diagnosis, tumor size, nodal status, histologic type, family history, estrogen receptor/progesterone receptor status, or HER-2 status, a trend was seen toward increased VEGF expression in the black cohort (26% black vs. 13% white, p = .068). Within the margin-negative cohort, VEGF did not predict for local relapse-free survival (RFS) (96% vs. 95%), nodal RFS (100% vs. 100%), distant metastasis-free survival (91% vs. 92%), overall survival (92% vs. 97%), respectively (all p >.05). Subset analysis revealed that VEGF was highly predictive of local RFS in node-positive, margin

  9. Endothelial progenitor cells in chronic obstructive pulmonary disease and emphysema.

    PubMed

    Doyle, Margaret F; Tracy, Russell P; Parikh, Megha A; Hoffman, Eric A; Shimbo, Daichi; Austin, John H M; Smith, Benjamin M; Hueper, Katja; Vogel-Claussen, Jens; Lima, Joao; Gomes, Antoinette; Watson, Karol; Kawut, Steven; Barr, R Graham

    2017-01-01

    Endothelial injury is implicated in the pathogenesis of COPD and emphysema; however the role of endothelial progenitor cells (EPCs), a marker of endothelial cell repair, and circulating endothelial cells (CECs), a marker of endothelial cell injury, in COPD and its subphenotypes is unresolved. We hypothesized that endothelial progenitor cell populations would be decreased in COPD and emphysema and that circulating endothelial cells would be increased. Associations with other subphenotypes were examined. The Multi-Ethnic Study of Atherosclerosis COPD Study recruited smokers with COPD and controls age 50-79 years without clinical cardiovascular disease. Endothelial progenitor cell populations (CD34+KDR+ and CD34+KDR+CD133+ cells) and circulating endothelial cells (CD45dimCD31+CD146+CD133-) were measured by flow cytometry. COPD was defined by standard spirometric criteria. Emphysema was assessed qualitatively and quantitatively on CT. Full pulmonary function testing and expiratory CTs were measured in a subset. Among 257 participants, both endothelial progenitor cell populations, and particularly CD34+KDR+ endothelial progenitor cells, were reduced in COPD. The CD34+KDR+CD133+ endothelial progenitor cells were associated inversely with emphysema extent. Both endothelial progenitor cell populations were associated inversely with extent of panlobular emphysema and positively with diffusing capacity. Circulating endothelial cells were not significantly altered in COPD but were inversely associated with pulmonary microvascular blood flow on MRI. There was no consistent association of endothelial progenitor cells or circulating endothelial cells with measures of gas trapping. These data provide evidence that endothelial repair is impaired in COPD and suggest that this pathological process is specific to emphysema.

  10. Endothelial progenitor cells in chronic obstructive pulmonary disease and emphysema

    PubMed Central

    Tracy, Russell P.; Parikh, Megha A.; Hoffman, Eric A.; Shimbo, Daichi; Austin, John H. M.; Smith, Benjamin M.; Hueper, Katja; Vogel-Claussen, Jens; Lima, Joao; Gomes, Antoinette; Watson, Karol; Kawut, Steven; Barr, R. Graham

    2017-01-01

    Endothelial injury is implicated in the pathogenesis of COPD and emphysema; however the role of endothelial progenitor cells (EPCs), a marker of endothelial cell repair, and circulating endothelial cells (CECs), a marker of endothelial cell injury, in COPD and its subphenotypes is unresolved. We hypothesized that endothelial progenitor cell populations would be decreased in COPD and emphysema and that circulating endothelial cells would be increased. Associations with other subphenotypes were examined. The Multi-Ethnic Study of Atherosclerosis COPD Study recruited smokers with COPD and controls age 50–79 years without clinical cardiovascular disease. Endothelial progenitor cell populations (CD34+KDR+ and CD34+KDR+CD133+ cells) and circulating endothelial cells (CD45dimCD31+CD146+CD133-) were measured by flow cytometry. COPD was defined by standard spirometric criteria. Emphysema was assessed qualitatively and quantitatively on CT. Full pulmonary function testing and expiratory CTs were measured in a subset. Among 257 participants, both endothelial progenitor cell populations, and particularly CD34+KDR+ endothelial progenitor cells, were reduced in COPD. The CD34+KDR+CD133+ endothelial progenitor cells were associated inversely with emphysema extent. Both endothelial progenitor cell populations were associated inversely with extent of panlobular emphysema and positively with diffusing capacity. Circulating endothelial cells were not significantly altered in COPD but were inversely associated with pulmonary microvascular blood flow on MRI. There was no consistent association of endothelial progenitor cells or circulating endothelial cells with measures of gas trapping. These data provide evidence that endothelial repair is impaired in COPD and suggest that this pathological process is specific to emphysema. PMID:28291826

  11. Aging-associated oxidized albumin promotes cellular senescence and endothelial damage

    PubMed Central

    Luna, Carlos; Alique, Matilde; Navalmoral, Estefanía; Noci, Maria-Victoria; Bohorquez-Magro, Lourdes; Carracedo, Julia; Ramírez, Rafael

    2016-01-01

    Increased levels of oxidized proteins with aging have been considered a cardiovascular risk factor. However, it is unclear whether oxidized albumin, which is the most abundant serum protein, induces endothelial damage. The results of this study indicated that with aging processes, the levels of oxidized proteins as well as endothelial microparticles release increased, a novel marker of endothelial damage. Among these, oxidized albumin seems to play a principal role. Through in vitro studies, endothelial cells cultured with oxidized albumin exhibited an increment of endothelial damage markers such as adhesion molecules and apoptosis levels. In addition, albumin oxidation increased the amount of endothelial microparticles that were released. Moreover, endothelial cells with increased oxidative stress undergo senescence. In addition, endothelial cells cultured with oxidized albumin shown a reduction in endothelial cell migration measured by wound healing. As a result, we provide the first evidence that oxidized albumin induces endothelial injury which then contributes to the increase of cardiovascular disease in the elderly subjects. PMID:27042026

  12. Circulating endothelial cells: a new biomarker of endothelial dysfunction in hematological diseases.

    PubMed

    Gendron, Nicolas; Smadja, David M

    2016-08-01

    The endothelium and its integrity are in the center of numerous cardiovascular, pulmonary and tumoral diseases. Several studies identified different circulating cellular sub-populations, which allow a noninvasive exploration of endothelial dysfunction. Furthermore, angiogenesis plays a major role in the biology of benign and malignant hematologic diseases. Among these biomarkers, circulating endothelial cells could be considered as a marker of endothelial injury and/or endothelial activation as well as vascular remodeling, whereas circulating endothelial progenitor cells would be only involved in the vascular regeneration. In the future, the quantification of circulating endothelial cells in many diseases could be a noninvasive biomarker used in diagnosis, prognostic and therapeutic follow-up of lung vasculopathy and/or residual disease of hematological malignancies.

  13. Immunohistochemical study of the lymphatic vessels in major salivary glands of the rat.

    PubMed

    Aiyama, Shigeo; Kikuchi, Kenichiro; Takada, Kiyomi; Ikeda, Rie; Sato, Sumie; Kuroki, Jyunya

    2011-02-01

    This study was designed to examine whether lymphatic vessels are present in the lobules of major salivary glands in the rat. Immunostaining with an antibody against podoplanin, a lymphatic endothelial cell marker, was performed on sections of the submandibular, sublingual and parotid glands. Light microscopy demonstrated podoplanin-positive lymphatic vessels around the interlobular ducts and the interlobular arteries and veins in the interlobular connective tissue in all of the major salivary glands. No podoplanin-positive lymphatic vessels were found in the lobules. Electron microscopy also demonstrated lymphatic endothelial cells showing podoplanin expression only in the interlobular connective tissue. These findings suggest that the lymphatic system of the rat major salivary glands originates in the interlobular connective tissue, and not in the lobules.

  14. Bubble-Induced Endothelial Microparticles Promote Endothelial Dysfunction

    PubMed Central

    Huang, Guoyang; Zhang, Kun; Qing, Long; Liu, Wenwu; Xu, Weigang

    2017-01-01

    Decompression sickness is a systemic pathophysiological process caused by bubbles and endothelial microparticles (EMPs) are established markers reflecting competency of endothelial function and vascular biology. Here, we investigated the effects of bubble-induced EMPs on endothelial cells in vitro and vivo. Rat pulmonary microvascular endothelial cells (PMVECs) were isolated and stimulated by bubbles and bubble-induced EMPs were collected and incubated with normal PMVECs in vitro. Cell viability and apoptosis were detected using Cell Counting Kit-8 assay and Annexin V FITC/PI double staining, respectively. Cell permeability and pro-inflammatory cytokines were determined by electric cell substrate impedance sensing and enzyme-linked immunosorbent assay, respectively. Intracellular nitric oxide and reactive oxygen species production were analyzed microscopically. In vivo study, bubble-induced EMPs were intravenously injected to the rats and soluble thrombomodulin, intercellular adhesion molecule 1, and vascullar adhesion molecule 1 were involved in evaluating endothelial dysfunction. In our study, bubble stimulus resulted in a significant increase of EMPs release by 3 fold. Bubble-induced EMPs significantly decreased cell viability and increased cell apoptosis. Moreover, bubble-induced EMPs induced abnormal increase of cell permeability and over-expression of pro-inflammatory cytokines. Intracellular ROS production increased while NO production decreased. These negative effects caused by bubble-induced EMPs were remarkably suppressed when EMPs pretreated with surfactant FSN-100. Finally, intravenous injection of bubble-induced EMPs caused elevations of soluble thrombomodulin and pro-inflammatory cytokines in the circulation. Altogether, our results demonstrated that bubble-induced EMPs can mediate endothelial dysfunction in vitro and vivo, which can be attenuated by EMPs abatement strategy. These data expanded our horizon of the detrimental effects of bubble

  15. Isolation and Characterization of Circulating Lymphatic Endothelial Colony Forming Cells

    PubMed Central

    DiMaio, Terri A.; Wentz, Breanna L.; Lagunoff, Michael

    2016-01-01

    Rationale The identification of circulating endothelial progenitor cells has led to speculation regarding their origin as well as their contribution to neovascular development. Two distinct types of endothelium make up the blood and lymphatic vessel system. However, it has yet to be determined whether there are distinct lymphatic-specific circulating endothelial progenitor cells. Objective This study aims to isolate and characterize the cellular properties and global gene expression of lymphatic-specific endothelial progenitor cells. Methods and Results We isolated circulating endothelial colony forming cells (ECFCs) from whole peripheral blood. These cells are endothelial in nature, as defined by their expression of endothelial markers and their ability to undergo capillary morphogenesis in three-dimensional culture. A subset of isolated colonies express markers of lymphatic endothelium, including VEGFR-3 and Prox-1, with low levels of VEGFR-1, a blood endothelial marker, while the bulk of the isolated cells express high VEGFR-1 levels with low VEGFR-3 and Prox-1 expression. The different isolates have differential responses to VEGF-C, a lymphatic endothelial specific cytokine, strongly suggesting that there are lymphatic specific and blood specific ECFCs. Global analysis of gene expression revealed key differences in the regulation of pathways involved in cellular differentiation between blood and lymphatic-specific ECFCs. Conclusion These data indicate that there are two distinguishable circulating ECFC types, blood and lymphatic, which are likely to have discrete functions during neovascularization. PMID:26597759

  16. Monocytes can be induced to express lymphatic phenotypes.

    PubMed

    Changming, W; Xin, L; Hua, T; Shikun, W; Qiong, X; Zhigeng, Z; Xueying, W

    2011-06-01

    Although it has been recently shown that monocytes can transdifferentiate into blood vascular endothelial cells which are involved in angiogenesis, little attention has been paid to their potential to transdifferentiate into lymphatic endothelial cells. Therefore, we examined this question in our study. We first stimulated monocytes with either fibronectin (FN), VEGF-C, TNF-alpha, LPS, or IL-3 for 24h. Then we examined the expression of several markers of lymphatic endothelium and found that the monocytes expressed specific lymphatic endothelial markers, LYVE-1, Podoplanin, and Prox-1, but not common endothelial markers vWF or eNOS. Next, monocytes were incubated in endothelial growth medium with FN and VEGF-C for 6d. These monocytes were also found to express LYVE-1, Podoplanin and Prox-1, but not vWF or eNOS. Our results indicate that monocytes in vitro can be easily induced to present lymphatic phenotypes in an inflammatory environment.

  17. [Vascular endothelial Barrier Function].

    PubMed

    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.

  18. Marker development

    SciTech Connect

    Adams, M.R.

    1987-05-01

    This report is to discuss the marker development for radioactive waste disposal sites. The markers must be designed to last 10,000 years, and place no undue burdens on the future generations. Barriers cannot be constructed that preclude human intrusion. Design specifications for surface markers will be discussed, also marker pictograms will also be covered.

  19. Effect of cinnamon, cardamom, saffron and ginger consumption on blood pressure and a marker of endothelial function in patients with type 2 diabetes mellitus: A randomized controlled clinical trial.

    PubMed

    Azimi, Paria; Ghiasvand, Reza; Feizi, Awat; Hosseinzadeh, Javad; Bahreynian, Maryam; Hariri, Mitra; Khosravi-Boroujeni, Hossein

    2016-06-01

    Herbal medicines with high amounts of phytochemicals have been shown to have beneficial effects on blood pressure (BP), endothelial function and anthropometric measures. This study aimed to determine the effect of herbal treatment on BP, endothelial function and anthropometric measures in patients with type 2 diabetes mellitus (T2DM). This clinical trial included 204 T2DM patients randomly assigned to four intervention groups receiving 3 g cinnamon, 3 g cardamom, 1 g saffron or 3 g ginger with three glasses of black tea, and one control group consuming only three glasses of tea without any herbals, for 8 weeks. Intercellular adhesion molecule-1 (ICAM-1), systolic and diastolic BP and anthropometric measures were collected at baseline and after 8 weeks. No significant difference was found between various medicinal plants in terms of influencing BP, serum soluble (s)ICAM-1 concentrations and anthropometric measures. However, in within-group comparison saffron and ginger intakes significantly reduced sICAM-1 concentrations (340.9 ± 14.4 vs 339.69 ± 14.4 ng/ml, p = 0.01, and 391.78 ± 16.0 vs 390.97 ± 15.8 ng/ml, p = 0.009, respectively) and ginger intake affected systolic BP (143.06 ± 0.2 vs 142.07 ± 0.2 mmHg, p = 0.02). Although administration of these herbal medicines as supplementary remedies could affect BP and sICAM-1 concentrations, there was no significant difference between the plants in terms of influencing anthropometric measures, BP and endothelial function.

  20. Identification of a novel platelet antagonist that binds to CLEC-2 and suppresses podoplanin-induced platelet aggregation and cancer metastasis

    PubMed Central

    Chang, Yu-Tsui; Lu, Meng-Hong; Huang, Tur-Fu; Chong, Kowit-Yu; Liao, Hsiang-Ruei; Cheng, Ju-Chien; Tseng, Ching-Ping

    2015-01-01

    Podoplanin (PDPN) enhances tumor metastases by eliciting tumor cell-induced platelet aggregation (TCIPA) through activation of platelet C-type lectin-like receptor 2 (CLEC-2). A novel and non-cytotoxic 5-nitrobenzoate compound 2CP was synthesized that specifically inhibited the PDPN/CLEC-2 interaction and TCIPA with no effect on platelet aggregation stimulated by other platelet agonists. 2CP possessed anti-cancer metastatic activity in vivo and augmented the therapeutic efficacy of cisplatin in the experimental animal model without causing a bleeding risk. Analysis of the molecular action of 2CP further revealed that Akt1/PDK1 and PKCμ were two alternative CLEC-2 signaling pathways mediating PDPN-induced platelet activation. 2CP directly bound to CLEC-2 and, by competing with the same binding pocket of PDPN in CLEC-2, inhibited PDPN-mediated platelet activation. This study provides evidence that 2CP is the first defined platelet antagonist with CLEC-2 binding activity. The augmentation in the therapeutic efficacy of cisplatin by 2CP suggests that a combination of a chemotherapeutic agent and a drug with anti-TCIPA activity such as 2CP may prove clinically effective. PMID:26528756

  1. Islet Endothelial Cells Derived From Mouse Embryonic Stem Cells.

    PubMed

    Jain, Neha; Lee, Eun Jung

    2016-01-01

    The islet endothelium comprises a specialized population of islet endothelial cells (IECs) expressing unique markers such as nephrin and α-1 antitrypsin (AAT) that are not found in endothelial cells in surrounding tissues. However, due to difficulties in isolating and maintaining a pure population of these cells, the information on these islet-specific cells is currently very limited. Interestingly, we have identified a large subpopulation of endothelial cells exhibiting IEC phenotype, while deriving insulin-producing cells from mouse embryonic stem cells (mESCs). These cells were identified by the uptake of low-density lipoprotein (LDL) and were successfully isolated and subsequently expanded in endothelial cell culture medium. Further analysis demonstrated that the mouse embryonic stem cell-derived endothelial cells (mESC-ECs) not only express classical endothelial markers, such as platelet endothelial cell adhesion molecule (PECAM1), thrombomodulin, intercellular adhesion molecule-1 (ICAM-1), and endothelial nitric oxide synthase (eNOS) but also IEC-specific markers such as nephrin and AAT. Moreover, mESC-ECs secrete basement membrane proteins such as collagen type IV, laminin, and fibronectin in culture and form tubular networks on a layer of Matrigel, demonstrating angiogenic activity. Further, mESC-ECs not only express eNOS, but also its eNOS expression is glucose dependent, which is another characteristic phenotype of IECs. With the ability to obtain highly purified IECs derived from pluripotent stem cells, it is possible to closely examine the function of these cells and their interaction with pancreatic β-cells during development and maturation in vitro. Further characterization of tissue-specific endothelial cell properties may enhance our ability to formulate new therapeutic angiogenic approaches for diabetes.

  2. Endothelial Dysfunction and Inflammation: Immunity in Rheumatoid Arthritis

    PubMed Central

    Yang, XueZhi; Chang, Yan; Wei, Wei

    2016-01-01

    Inflammation, as a feature of rheumatoid arthritis (RA), leads to the activation of endothelial cells (ECs). Activated ECs induce atherosclerosis through an increased expression of leukocyte adhesion molecules. Endothelial dysfunction (ED) is recognized as a failure of endothelial repair mechanisms. It is also an early preclinical marker of atherosclerosis and is commonly found in RA patients. RA is now established as an independent cardiovascular risk factor, while mechanistic determinants of ED in RA are still poorly understood. An expanding body of study has shown that EC at a site of RA is both active participant and regulator of inflammatory process. Over the last decade, a role for endothelial dysfunction in RA associated with cardiovascular disease (CVD) has been hypothesized. At the same time, several maintenance drugs targeting this phenomenon have been tested, which has promising results. Assessment of endothelial function may be a useful tool to identify and monitor RA patients. PMID:27122657

  3. Focally regulated endothelial proliferation and cell death in human synovium.

    PubMed Central

    Walsh, D. A.; Wade, M.; Mapp, P. I.; Blake, D. R.

    1998-01-01

    Angiogenesis and vascular insufficiency each may support the chronic synovial inflammation of rheumatoid arthritis. We have shown by quantitative immunohistochemistry and terminal uridyl deoxynucleotide nick end labeling that endothelial proliferation and cell death indices were each increased in synovia from patients with rheumatoid arthritis compared with osteoarthritic and noninflamed controls, whereas endothelial fractional areas did not differ significantly among disease groups. Markers of proliferation were associated with foci immunoreactive for vascular endothelial growth factor and integrin alpha(v)beta3, whereas cell death was observed in foci in which immunoreactivities for these factors were weak or absent. No association was found with thrombospondin immunoreactivity. The balance between angiogenesis and vascular regression in rheumatoid synovitis may be determined by the focal expression of angiogenic and endothelial survival factors. Increased endothelial cell turnover may contribute to microvascular dysfunction and thereby facilitate persistent synovitis. Images Figure 1 Figure 3 Figure 4 PMID:9502411

  4. [Endothelial cell adhesion molecules].

    PubMed

    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.

  5. Lipopolysaccharide induces a fibrotic-like phenotype in endothelial cells.

    PubMed

    Echeverría, César; Montorfano, Ignacio; Sarmiento, Daniela; Becerra, Alvaro; Nuñez-Villena, Felipe; Figueroa, Xavier F; Cabello-Verrugio, Claudio; Elorza, Alvaro A; Riedel, Claudia; Simon, Felipe

    2013-06-01

    Endothelial dysfunction is crucial in endotoxaemia-derived sepsis syndrome pathogenesis. It is well accepted that lipopolysaccharide (LPS) induces endothelial dysfunction through immune system activation. However, LPS can also directly generate actions in endothelial cells (ECs) in the absence of participation by immune cells. Although interactions between LPS and ECs evoke endothelial death, a significant portion of ECs are resistant to LPS challenge. However, the mechanism that confers endothelial resistance to LPS is not known. LPS-resistant ECs exhibit a fibroblast-like morphology, suggesting that these ECs enter a fibrotic programme in response to LPS. Thus, our aim was to investigate whether LPS is able to induce endothelial fibrosis in the absence of immune cells and explore the underlying mechanism. Using primary cultures of ECs and culturing intact blood vessels, we demonstrated that LPS is a crucial factor to induce endothelial fibrosis. We demonstrated that LPS was able and sufficient to promote endothelial fibrosis, in the absence of immune cells through an activin receptor-like kinase 5 (ALK5) activity-dependent mechanism. LPS-challenged ECs showed an up-regulation of both fibroblast-specific protein expression and extracellular matrix proteins secretion, as well as a down-regulation of endothelial markers. These results demonstrate that LPS is a crucial factor in inducing endothelial fibrosis in the absence of immune cells through an ALK5-dependent mechanism. It is noteworthy that LPS-induced endothelial fibrosis perpetuates endothelial dysfunction as a maladaptive process rather than a survival mechanism for protection against LPS. These findings are useful in improving current treatment against endotoxaemia-derived sepsis syndrome and other inflammatory diseases.

  6. Marker chromosomes.

    PubMed

    Rao, Kiran Prabhaker; Belogolovkin, Victoria

    2013-04-01

    Marker chromosomes are a morphologically heterogeneous group of structurally abnormal chromosomes that pose a significant challenge in prenatal diagnosis. Phenotypes associated with marker chromosomes are highly variable and range from normal to severely abnormal. Clinical outcomes are very difficult to predict when marker chromosomes are detected prenatally. In this review, we outline the classification, etiology, cytogenetic characterization, and clinical consequences of marker chromosomes, as well as practical approaches to prenatal diagnosis and genetic counseling.

  7. [Assessment of endothelial function in autoimmune diseases].

    PubMed

    Benhamou, Y; Bellien, J; Armengol, G; Gomez, E; Richard, V; Lévesque, H; Joannidès, R

    2014-08-01

    Numerous autoimmune-inflammatory rheumatic diseases have been associated with accelerated atherosclerosis or other types of vasculopathy leading to an increase in cardiovascular disease incidence. In addition to traditional cardiovascular risk factors, endothelial dysfunction is an important early event in the pathogenesis of atherosclerosis, contributing to plaque initiation and progression. Endothelial dysfunction is characterized by a shift of the actions of the endothelium toward reduced vasodilation, a proinflammatory and a proadhesive state, and prothrombic properties. Therefore, assessment of endothelial dysfunction targets this vascular phenotype using several biological markers as indicators of endothelial dysfunction. Measurements of soluble adhesion molecules (ICAM-1, VCAM-1, E-selectin), pro-thrombotic factors (thrombomodulin, von Willebrand factor, plasminogen activator inhibitor-1) and inflammatory cytokines are most often performed. Regarding the functional assessment of the endothelium, the flow-mediated dilatation of conduit arteries is a non-invasive method widely used in pathophysiological and interventional studies. In this review, we will briefly review the most relevant information upon endothelial dysfunction mechanisms and explorations. We will summarize the similarities and differences in the biological and functional assessments of the endothelium in different autoimmune diseases.

  8. Vascular endothelial-cadherin downregulation as a feature of endothelial transdifferentiation in monocrotaline-induced pulmonary hypertension.

    PubMed

    Nikitopoulou, Ioanna; Orfanos, Stylianos E; Kotanidou, Anastasia; Maltabe, Violetta; Manitsopoulos, Nikolaos; Karras, Panagiotis; Kouklis, Panos; Armaganidis, Apostolos; Maniatis, Nikolaos A

    2016-08-01

    Increased pulmonary vascular resistance in pulmonary hypertension (PH) is caused by vasoconstriction and obstruction of small pulmonary arteries by proliferating vascular cells. In analogy to cancer, subsets of proliferating cells may be derived from endothelial cells transitioning into a mesenchymal phenotype. To understand phenotypic shifts transpiring within endothelial cells in PH, we injected rats with alkaloid monocrotaline to induce PH and measured lung tissue levels of endothelial-specific protein and critical differentiation marker vascular endothelial (VE)-cadherin. VE-cadherin expression by immonoblotting declined significantly 24 h and 15 days postinjection to rebound to baseline at 30 days. There was a concomitant increase in transcriptional repressors Snail and Slug, along with a reduction in VE-cadherin mRNA. Mesenchymal markers α-smooth muscle actin and vimentin were upregulated by immunohistochemistry and immunoblotting, and α-smooth muscle actin was colocalized with endothelial marker platelet endothelial cell adhesion molecule-1 by confocal microscopy. Apoptosis was limited in this model, especially in the 24-h time point. In addition, monocrotaline resulted in activation of protein kinase B/Akt, endothelial nitric oxide synthase (eNOS), nuclear factor (NF)-κB, and increased lung tissue nitrotyrosine staining. To understand the etiological relationship between nitrosative stress and VE-cadherin suppression, we incubated cultured rat lung endothelial cells with endothelin-1, a vasoconstrictor and pro-proliferative agent in pulmonary arterial hypertension. This resulted in activation of eNOS, NF-κB, and Akt, in addition to induction of Snail, downregulation of VE-cadherin, and synthesis of vimentin. These effects were blocked by eNOS inhibitor N(ω)-nitro-l-arginine methyl ester. We propose that transcriptional repression of VE-cadherin by nitrosative stress is involved in endothelial-mesenchymal transdifferentiation in experimental PH.

  9. Glucose transporter 1-positive endothelial cells in infantile hemangioma exhibit features of facultative stem cells.

    PubMed

    Huang, Lan; Nakayama, Hironao; Klagsbrun, Michael; Mulliken, John B; Bischoff, Joyce

    2015-01-01

    Endothelial glucose transporter 1 (GLUT1) is a definitive and diagnostic marker for infantile hemangioma (IH), a vascular tumor of infancy. To date, GLUT1-positive endothelial cells in IH have not been quantified nor directly isolated and studied. We isolated GLUT1-positive and GLUT1-negative endothelial cells from IH specimens and characterized their proliferation, differentiation, and response to propranolol, a first-line therapy for IH, and to rapamycin, an mTOR pathway inhibitor used to treat an increasingly wide array of proliferative disorders. Although freshly isolated GLUT1-positive cells, selected using anti-GLUT1 magnetic beads, expressed endothelial markers CD31, VE-Cadherin, and vascular endothelial growth factor receptor 2, they converted to a mesenchymal phenotype after 3 weeks in culture. In contrast, GLUT1-negative endothelial cells exhibited a stable endothelial phenotype in vitro. GLUT1-selected cells were clonogenic when plated as single cells and could be induced to redifferentiate into endothelial cells, or into pericytes/smooth muscle cells or into adipocytes, indicating a stem cell-like phenotype. These data demonstrate that, although they appear and function in the tumor as bona fide endothelial cells, the GLUT1-positive endothelial cells display properties of facultative stem cells. Pretreatment with rapamycin for 4 days significantly slowed proliferation of GLUT1-selected cells, whereas propranolol pretreatment had no effect. These results reveal for the first time the facultative nature of GLUT1-positive endothelial cells in IH.

  10. Central Role of eNOS in the Maintenance of Endothelial Homeostasis

    PubMed Central

    Rodriguez-Mateos, Ana; Kelm, Malte

    2015-01-01

    Abstract Significance: Disruption of endothelial function is considered a key event in the development and progression of atherosclerosis. Endothelial nitric oxide synthase (eNOS) is a central regulator of cellular function that is important to maintain endothelial homeostasis. Recent Advances: Endothelial homeostasis encompasses acute responses such as adaption of flow to tissue's demand and more sustained responses to injury such as re-endothelialization and sprouting of endothelial cells (ECs) and attraction of circulating angiogenic cells (CAC), both of which support repair of damaged endothelium. The balance and the intensity of endothelial damage and repair might be reflected by changes in circulating endothelial microparticles (EMP) and CAC. Flow-mediated vasodilation (FMD) is a generally accepted clinical read-out of NO-dependent vasodilation, whereas EMP are upcoming prognostically validated markers of endothelial injury and CAC are reflective of the regenerative capacity with both expressing a functional eNOS. These markers can be integrated in a clinical endothelial phenotype, reflecting the net result between damage from risk factors and endogenous repair capacity with NO representing a central signaling molecule. Critical Issues: Improvements of reproducibility and observer independence of FMD measurements and definitions of relevant EMP and CAC subpopulations warrant further research. Future Directions: Endothelial homeostasis may be a clinical therapeutic target for cardiovascular health maintenance. Antioxid. Redox Signal. 22, 1230–1242. PMID:25330054

  11. Matrix Gla protein regulates differentiation of endothelial cells derived from mouse embryonic stem cells.

    PubMed

    Yao, Jiayi; Guihard, Pierre J; Blazquez-Medela, Ana M; Guo, Yina; Liu, Ting; Boström, Kristina I; Yao, Yucheng

    2016-01-01

    Matrix Gla protein (MGP) is an antagonist of bone morphogenetic proteins and expressed in vascular endothelial cells. Lack of MGP causes vascular abnormalities in multiple organs in mice. The objective of this study is to define the role of MGP in early endothelial differentiation. We find that expression of endothelial markers is highly induced in Mgp null organs, which, in wild type, contain high MGP expression. Furthermore, Mgp null embryonic stem cells express higher levels of endothelial markers than wild-type controls and an abnormal temporal pattern of expression. We also find that the Mgp-deficient endothelial cells adopt characteristics of mesenchymal stem cells. We conclude that loss of MGP causes dysregulation of early endothelial differentiation.

  12. Adult cutaneous hemangiomas are composed of nonreplicating endothelial cells.

    PubMed

    Tuder, R M; Young, R; Karasek, M; Bensch, K

    1987-12-01

    Thirty-four human "cherry" dermal hemangiomas were studied by electron microscopy, immunohistochemistry, and cell culture to assess the neoplastic nature of these lesions. Electron microscopy of nine hemangiomas revealed a pronounced thickening of the basement membrane (0.6 to 14 micron) in 93% of the total 158 vascular structures examined within the lesions. This increase was caused mainly by multiple layers of basal lamina, which were irregular in outline and frequently associated with pericytes. Basement membrane changes were present both in the periphery of the hemangiomas, as well as in the center of the lesions. Immature vessels could not be identified and mitoses were absent in all endothelial cells. Using an immunohistochemical marker (Ki67) specific for proliferating cells in G2 and S phases, positive staining was not found in the endothelial cells lining the hemangiomatous vessels, whereas basal epidermal keratinocytes in the same preparations and cultured microvascular endothelial cells expressed the antigen. Endothelial cells of nine hemangiomas did not stain with an activation-related antibody (E12) specific for endothelial cells. When endothelial cells from 14 hemangiomas were isolated and cultured under conditions that support the growth of normal human skin microvascular endothelial cells, the cells of hemangiomatous origin failed to grow. We conclude that the adult hemangiomas may not be true neoplasms, but a tissue overgrowth composed of mature vessels resembling dermal venules, lined by endothelial cells with virtually no turnover.

  13. Novel markers of endothelial dysfunction and inflammation in Behçet's disease patients with ocular involvement: epicardial fat thickness, carotid intima media thickness, serum ADMA level, and neutrophil-to-lymphocyte ratio.

    PubMed

    Yuksel, Murat; Yildiz, Abdulkadir; Oylumlu, Mustafa; Turkcu, Fatih Mehmet; Bilik, Mehmet Zihni; Ekinci, Aysun; Elbey, Bilal; Tekbas, Ebru; Alan, Sait

    2016-03-01

    The etiology of Behçet's disease (BD) has not been fully elucidated. However, immunological and environmental factors, endothelial dysfunction (ED), and genetic susceptibility have been proposed to play a role. In this study, we aimed to evaluate epicardial fat thickness (EFT) together with serum asymmetric dimethylarginine (ADMA), carotid intima media thickness (CIMT), and neutrophil-to-lymphocyte ratio (NLR) in BD patients with ocular involvement. Thirty-six ocular BD patients (17 active and 19 inactive ocular involvement), and 35 age and sex-matched healthy controls were enrolled to this cross-sectional study. All patients underwent examinations with transthoracic echocardiography and carotid Doppler ultrasound. Serum ADMA levels, CIMT, EFT, and NLR were compared between groups, and their association with disease activity was evaluated. Behçet's disease patients had higher WBC counts, neutrophil counts, NLR, CIMT, EFT values, and serum ADMA levels than do healthy controls. The other biochemical, hematological, and echocardiographic parameters were comparable between the two groups. Behçet's disease duration was positively correlated with EFT and CIMT. Multivariate logistic regression analysis revealed that increased serum ADMA concentration and CIMT are independently associated with BD. Neutrophil counts, NLR, and serum ADMA level were higher, and lymphocyte count was lower in patients with active ocular BD compared to those of inactive ocular BD group. Carotid intima media thickness, serum ADMA level, EFT, and NLR were increased in ocular BD patients compared to healthy subjects. In addition, both serum ADMA level and NLR were associated with disease activity of ocular involvement. Increase in disease duration was associated with increase in CIMT and EFT which suggests that anatomical changes occur in time during the disease course. Increased CIMT, serum ADMA level, EFT, and NLR may provide new clues about the role of ED and inflammation in the

  14. Increased Detection of Lymphatic Vessel Invasion by D2-40 (Podoplanin) in Early Breast Cancer: Possible Influence on Patient Selection for Accelerated Partial Breast Irradiation

    SciTech Connect

    Debald, Manuel; Poelcher, Martin; Flucke, Uta; Walgenbach-Bruenagel, Gisela

    2010-07-15

    Purpose: Several international trials are currently investigating accelerated partial breast irradiation (APBI) for patients with early-stage breast cancer. According to existing guidelines, patients with lymphatic vessel invasion (LVI) do not qualify for APBI. D2-40 (podoplanin) significantly increases the frequency of LVI detection compared with conventional hematoxylin and eosin (HE) staining in early-stage breast cancer. Our purpose was to retrospectively assess the hypothetical change in management from APBI to whole breast radiotherapy with the application of D2-40. Patients and Methods: Immunostaining with D2-40 was performed on 254 invasive breast tumors of 247 patients. The following criteria were used to determine the eligibility for APBI: invasive ductal adenocarcinoma of {<=}3 cm, negative axillary node status (N0), and unifocal disease. Of the 247 patients, 74 with available information concerning LVI, as detected by D2-40 immunostaining and routine HE staining, formed our study population. Results: Using D2-40, our results demonstrated a significantly greater detection rate (p = .031) of LVI compared with routine HE staining. LVI was correctly identified by D2-40 (D2-40-positive LVI) in 10 (13.5%) of 74 tumors. On routine HE staining, 4 tumors (5.4%) were classified as HE-positive LVI. Doublestaining of these specimens with D2-40 unmasked false-positive LVI status in 2 (50%) of the 4 tumors. According to the current recommendations for APBI, immunostaining with D2-40 would have changed the clinical management from APBI to whole breast radiotherapy in 8 (10.8%) of 74 patients and from whole breast radiotherapy to APBI in 2 patients (2.7%). Conclusion: These data support the implementation of D2-40 immunostaining in the routine workup to determine a patient's eligibility for APBI.

  15. The chimeric antibody chLpMab-7 targeting human podoplanin suppresses pulmonary metastasis via ADCC and CDC rather than via its neutralizing activity

    PubMed Central

    Ogasawara, Satoshi; Fujii, Yuki; Oki, Hiroharu; Fukayama, Masashi; Nishioka, Yasuhiko; Kaneko, Mika K.

    2015-01-01

    Podoplanin (PDPN/Aggrus/T1α) binds to C-type lectin-like receptor-2 (CLEC-2) and induces platelet aggregation. PDPN is associated with malignant progression, tumor metastasis, and poor prognosis in several types of cancer. Although many anti-human PDPN (hPDPN) monoclonal antibodies (mAbs), such as D2-40 and NZ-1, have been established, these epitopes are limited to the platelet aggregation-stimulating (PLAG) domain (amino acids 29-54) of hPDPN. Recently, we developed a novel mouse anti-hPDPN mAb, LpMab-7, which is more sensitive than D2-40 and NZ-1, using the Cancer-specific mAb (CasMab) method. The epitope of LpMab-7 was shown to be entirely different from that of NZ-1, a neutralizing mAb against the PLAG domain according to an inhibition assay and lectin microarray analysis. In the present study, we produced a mouse-human chimeric anti-hPDPN mAb, chLpMab-7. ChLpMab-7 showed high antibody-dependent cellular cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC). Furthermore, chLpMab-7 inhibited the growth of hPDPN-expressing tumors in vivo. Although chLpMab-7 recognizes a non-PLAG domain of hPDPN, it suppressed the hematogenous metastasis of hPDPN-expressing tumors. These results indicated that chLpMab-7 suppressed tumor development and hematogenous metastasis in a neutralization-independent manner. In conclusion, hPDPN shows promise as a target in the development of a novel antibody-based therapy. PMID:26416352

  16. Synergism of matrix stiffness and vascular endothelial growth factor on mesenchymal stem cells for vascular endothelial regeneration.

    PubMed

    Wingate, Kathryn; Floren, Michael; Tan, Yan; Tseng, Pi Ou Nancy; Tan, Wei

    2014-09-01

    Mesenchymal stem cells (MSCs) hold tremendous potential for vascular tissue regeneration. Research has demonstrated that individual factors in the cell microenvironment such as matrix elasticity and growth factors regulate MSC differentiation to vascular lineage. However, it is not well understood how matrix elasticity and growth factors combine to direct the MSC fate. This study examines the combined effects of matrix elasticity and vascular endothelial growth factor (VEGF) on both MSC differentiation into endothelial lineage and MSC paracrine signaling. MSCs were seeded in soft nanofibrous matrices with or without VEGF, and in Petri dishes with or without VEGF. Only MSCs seeded in three-dimensional soft matrices with VEGF showed significant increases in the expression of endothelial markers (vWF, eNOS, Flt-1, and Flk-1), while eliminating the expression of smooth muscle marker (SM-α-actin). MSCs cultured in VEGF alone on two-dimensional dishes showed increased expression of both early-stage endothelial and smooth muscle markers, indicating immature vascular differentiation. Furthermore, MSCs cultured in soft matrices with VEGF showed faster upregulation of endothelial markers compared with MSCs cultured in VEGF alone. Paracrine signaling studies found that endothelial cells cultured in the conditioned media from MSCs differentiated in the soft matrix and VEGF condition exhibited increased migration and formation of capillary-like structures. These results demonstrate that VEGF and soft matrix elasticity act synergistically to guide MSC differentiation into mature endothelial phenotype while enhancing paracrine signaling. Therefore, it is critical to control both mechanical and biochemical factors to safely regenerate vascular tissues with MSCs.

  17. Endothelial Activation and Repair During Hantavirus Infection: Association with Disease Outcome

    PubMed Central

    Connolly-Andersen, Anne-Marie; Thunberg, Therese; Ahlm, Clas

    2014-01-01

    Background.  Endothelial activation and dysfunction play a central role in the pathogenesis of sepsis and viral hemorrhagic fevers. Hantaviral disease is a viral hemorrhagic fever and is characterized by capillary dysfunction, although the underlying mechanisms for hantaviral disease are not fully elucidated. Methods.  The temporal course of endothelial activation and repair were analyzed during Puumala hantavirus infection and associated with disease outcome and a marker for hypoxia, insulin-like growth factor binding protein 1 (IGFBP-1). The following endothelial activation markers were studied: endothelial glycocalyx degradation (syndecan-1) and leukocyte adhesion molecules (soluble vascular cellular adhesion molecule 1, intercellular adhesion molecule 1, and endothelial selectin). Cytokines associated with vascular repair were also analyzed (vascular endothelial growth factor, erythropoietin, angiopoietin, and stromal cell-derived factor 1). Results.  Most of the markers we studied were highest during the earliest phase of hantaviral disease and associated with clinical and laboratory surrogate markers for disease outcome. In particular, the marker for glycocalyx degradation, syndecan-1, was significantly associated with levels of thrombocytes, albumin, IGFBP-1, decreased blood pressure, and disease severity. Conclusions.  Hantaviral disease outcome was associated with endothelial dysfunction. Consequently, the endothelium warrants further investigation when designing future medical interventions. PMID:25734100

  18. Glucose transporter 1-positive endothelial cells in infantile hemangioma exhibit features of facultative stem cells

    PubMed Central

    Huang, Lan; Nakayama, Hironao; Klagsbrun, Michael; Mulliken, John B.; Bischoff, Joyce

    2014-01-01

    Endothelial glucose transporter 1 (GLUT1) is a definitive and diagnostic marker for infantile hemangioma (IH), a vascular tumor of infancy. To date, GLUT1-positive endothelial cells in IH have not been quantified nor directly isolated and studied. We isolated GLUT1-positive and GLUT1-negative endothelial cells from IH specimens and characterized their proliferation, differentiation and response to propranolol, a first-line therapy for IH, and to rapamycin, an mTOR pathway inhibitor used to treat an increasingly wide array of proliferative disorders. Although freshly isolated GLUT1-positive cells, selected using anti-GLUT1 magnetic beads, expressed endothelial markers CD31, VE-Cadherin and VEGFR2, they converted to a mesenchymal phenotype after three weeks in culture. In contrast, GLUT1-negative endothelial cells exhibited a stable endothelial phenotype in vitro. GLUT1-selected cells were clonogenic when plated as single cells and could be induced to re-differentiate into endothelial cells, or into pericyte/smooth muscle cells or into adipocytes, indicating a stem cell-like phenotype. These data demonstrate that, although they appear and function in the tumor as bona fide endothelial cells, the GLUT1-positive endothelial cells display properties of facultative stem cells. Pretreatment with rapamycin for 4 days significantly slowed proliferation of GLUT1-selected cells, whereas propranolol pretreatment had no effect. These results reveal for the first time the facultative nature of GLUT1-positive endothelial cells in infantile hemangioma. PMID:25187207

  19. HDL and endothelial protection

    PubMed Central

    Tran-Dinh, A; Diallo, D; Delbosc, S; Varela-Perez, L Maria; Dang, QB; Lapergue, B; Burillo, E; Michel, JB; Levoye, A; Martin-Ventura, JL; Meilhac, O

    2013-01-01

    High-density lipoproteins (HDLs) represent a family of particles characterized by the presence of apolipoprotein A-I (apoA-I) and by their ability to transport cholesterol from peripheral tissues back to the liver. In addition to this function, HDLs display pleiotropic effects including antioxidant, anti-apoptotic, anti-inflammatory, anti-thrombotic or anti-proteolytic properties that account for their protective action on endothelial cells. Vasodilatation via production of nitric oxide is also a hallmark of HDL action on endothelial cells. Endothelial cells express receptors for apoA-I and HDLs that mediate intracellular signalling and potentially participate in the internalization of these particles. In this review, we will detail the different effects of HDLs on the endothelium in normal and pathological conditions with a particular focus on the potential use of HDL therapy to restore endothelial function and integrity. PMID:23488589

  20. Differentiation of mesenchymal stem cells from human amniotic fluid to vascular endothelial cells.

    PubMed

    Tancharoen, Waleephan; Aungsuchawan, Sirinda; Pothacharoen, Peraphan; Markmee, Runchana; Narakornsak, Suteera; Kieodee, Junjira; Boonma, Nonglak; Tasuya, Witoon

    2017-03-01

    Endothelial dysfunction is a principle feature of vascular-related disease. Endothelial cells have been acquired for the purposes of the restoration of damaged tissue in therapeutic angiogenesis. However, their use is limited by expansion capacity and the small amount of cells that are obtained. Human amniotic fluid mesenchymal stem cells (hAF-MSCs) are considered an important source for vascular tissue engineering. In this study, hAF-MSCs were characterized and then induced in order to differentiate into the endothelial-like cells. Human amniotic fluid cells (hAFCs) were obtained from amniocentesis at the second trimester of gestation. The cells were characterized as mesenchymal stem cells by flow cytometry. The results showed that the cells were positive for mesenchymal stem cell markers CD44, CD73, CD90 and HLA-ABC, and negative for CD31, Amniotic fluid stem cells marker: CD117, anti-human fibroblasts, HLA-DR and hematopoietic differentiation markers CD34 and CD45. The hAF-MSCs were differentiated into endothelial cells under the induction of vascular endothelial growth factor (VEGF) and analyzed for the expression of the endothelial-specific markers and function. The expression of the endothelial-specific markers was determined by reverse transcriptase-quantitative PCR (RT-qPCR), while immunofluorescent analysis demonstrated that the induced hAF-MSCs expressed von Willebrand factor (vWF), vascular endothelial growth factor receptor 2 (VEGFR2), CD31 and endothelial nitric oxide synthase (eNOS). The network formation assay showed that the induced hAF-MSCs formed partial networks. All results indicated that hAF-MSCs have the potential to be differentiated into endothelial-like cells, while human amniotic fluid might be a suitable source of MSCs for vascularized tissue engineering.

  1. Biomarkers of endothelial activation/dysfunction in infectious diseases

    PubMed Central

    Page, Andrea V; Liles, W Conrad

    2013-01-01

    Endothelial dysfunction contributes to the pathogenesis of a variety of potentially serious infectious diseases and syndromes, including sepsis and septic shock, hemolytic-uremic syndrome, severe malaria, and dengue hemorrhagic fever. Because endothelial activation often precedes overt endothelial dysfunction, biomarkers of the activated endothelium in serum and/or plasma may be detectable before classically recognized markers of disease, and therefore, may be clinically useful as biomarkers of disease severity or prognosis in systemic infectious diseases. In this review, the current status of mediators of endothelial cell function (angiopoietins-1 and -2), components of the coagulation pathway (von Willebrand Factor, ADAMTS13, and thrombomodulin), soluble cell-surface adhesion molecules (soluble E-selectin, sICAM-1, and sVCAM-1), and regulators of vascular tone and permeability (VEGF and sFlt-1) as biomarkers in severe infectious diseases is discussed in the context of sepsis, E. coli O157:H7 infection, malaria, and dengue virus infection. PMID:23669075

  2. New methods to evaluate endothelial function: A search for a marker of nitric oxide (NO) in vivo: re-evaluation of NOx in plasma and red blood cells and a trial to detect nitrosothiols.

    PubMed

    Ishibashi, Takaharu; Yoshida, Junko; Nishio, Matomo

    2003-12-01

    Although plasma NOx (NO(2)(-) and NO(3)(-)) has been used as an index of nitric oxide (NO) formation in vivo, many unreasonable results appeared even after active elimination of NOx contamination from laboratory ware. For example, plasma NOx concentrations did not increase during vasodilation mediated by the NO/cGMP pathway or after organ perfusion. A possible shift of NOx from plasma to erythrocytes (RBCs) as a cause of these phenomena has been excluded, leaving the destination of NOx (after leaving plasma) unknown. Kinetic analyses have revealed that steady state NOx concentrations in plasma and whole blood did not correlate with the NOx formation rate, but rather with the NOx elimination rate. Therefore, the supposition that the NO status is directly reflected by plasma NOx concentrations appears untenable. As nitrosothiols (R-SNOs), possible carriers of NO bioactivity, have been flagged as alternative indices of NO status in vivo, efforts have been made to detect these substances. When interference by ultrafiltration was eliminated, low molecular weight R-SNOs such as nitrosocystein and nitrosogluthathione were undetectable. However, a high-molecular weight R-SNO, nitrosoalbumin, was detected in human blood. Further research is required into the significance and practical use of nitrosoalbumin as a marker of NO in vivo.

  3. Assessment of Endothelial Dysfunction in Childhood Obesity and Clinical Use

    PubMed Central

    Hoymans, Vicky Y.; Van Craenenbroeck, Amaryllis H.; Vissers, Dirk K.; Vrints, Christiaan J.; Conraads, Viviane M.

    2013-01-01

    The association of obesity with noncommunicable diseases, such as cardiovascular complications and diabetes, is considered a major threat to the management of health care worldwide. Epidemiological findings show that childhood obesity is rapidly rising in Western society, as well as in developing countries. This pandemic is not without consequences and can affect the risk of future cardiovascular disease in these children. Childhood obesity is associated with endothelial dysfunction, the first yet still reversible step towards atherosclerosis. Advanced research techniques have added further insight on how childhood obesity and associated comorbidities lead to endothelial dysfunction. Techniques used to measure endothelial function were further brought to perfection, and novel biomarkers, including endothelial progenitor cells, were discovered. The aim of this paper is to provide a critical overview on both in vivo as well as in vitro markers for endothelial integrity. Additionally, an in-depth description of the mechanisms that disrupt the delicate balance between endothelial damage and repair will be given. Finally, the effects of lifestyle interventions and pharmacotherapy on endothelial dysfunction will be reviewed. PMID:23691262

  4. Endothelial cells derived from human embryonic stem cells

    NASA Astrophysics Data System (ADS)

    Levenberg, Shulamit; Golub, Justin S.; Amit, Michal; Itskovitz-Eldor, Joseph; Langer, Robert

    2002-04-01

    Human embryonic stem cells have the potential to differentiate into various cell types and, thus, may be useful as a source of cells for transplantation or tissue engineering. We describe here the differentiation steps of human embryonic stem cells into endothelial cells forming vascular-like structures. The human embryonic-derived endothelial cells were isolated by using platelet endothelial cell-adhesion molecule-1 (PECAM1) antibodies, their behavior was characterized in vitro and in vivo, and their potential in tissue engineering was examined. We show that the isolated embryonic PECAM1+ cells, grown in culture, display characteristics similar to vessel endothelium. The cells express endothelial cell markers in a pattern similar to human umbilical vein endothelial cells, their junctions are correctly organized, and they have high metabolism of acetylated low-density lipoprotein. In addition, the cells are able to differentiate and form tube-like structures when cultured on matrigel. In vivo, when transplanted into SCID mice, the cells appeared to form microvessels containing mouse blood cells. With further studies, these cells could provide a source of human endothelial cells that could be beneficial for potential applications such as engineering new blood vessels, endothelial cell transplantation into the heart for myocardial regeneration, and induction of angiogenesis for treatment of regional ischemia.

  5. Stress-Induced Premature Senescence of Endothelial and Endothelial Progenitor Cells

    PubMed Central

    Goligorsky, M.S.; Hirschi, K.

    2016-01-01

    This brief overview of premature senescence of dysfunctional endothelial and endothelial progenitor cells provides information on endothelial cell differentiation and specialization, their ontogeny, and controversies related to endothelial stem and progenitor cells. Stressors responsible for the dysfunction of endothelial and endothelial progenitor cells, as well as cellular mechanisms and consequences of endothelial cell dysfunction are presented. Metabolic signatures of dysfunctional endothelial cells and senescence pathways are described. Emerging strategies to rejuvenate endothelial and endothelial progenitor cells conclude the review. PMID:27451101

  6. Fuchs Endothelial Corneal Dystrophy

    PubMed Central

    Elhalis, Hussain; Azizi, Behrooz; Jurkunas, Ula V.

    2011-01-01

    Fuchs endothelial corneal dystrophy (FECD) is characterized by progressive loss of corneal endothelial cells, thickening of Descement’s membrane and deposition of extracellular matrix in the form of guttae. When the number of endothelial cells becomes critically low, the cornea swells and causes loss of vision. The clinical course of FECD usually spans 10–20 years. Corneal transplantation is currently the only modality used to restore vision. Over the last several decades genetic studies have detected several genes, as well as areas of chromosomal loci associated with the disease. Proteomic studies have given rise to several hypotheses regarding the pathogenesis of FECD. This review expands upon the recent findings from proteomic and genetic studies and builds upon recent advances in understanding the causes of this common corneal disorder. PMID:20964980

  7. PDGFRβ Is a Novel Marker of Stromal Activation in Oral Squamous Cell Carcinomas

    PubMed Central

    Han, Rong; Haines, Paul; Gallagher, George; Noonan, Vikki; Kukuruzinska, Maria; Monti, Stefano; Trojanowska, Maria

    2016-01-01

    Carcinoma associated fibroblasts (CAFs) form the main constituents of tumor stroma and play an important role in tumor growth and invasion. The presence of CAFs is a strong predictor of poor prognosis of head and neck squamous cell carcinoma. Despite significant progress in determining the role of CAFs in tumor progression, the mechanisms contributing to their activation remain poorly characterized, in part due to fibroblast heterogeneity and the scarcity of reliable fibroblast surface markers. To search for such markers in oral squamous cell carcinoma (OSCC), we applied a novel approach that uses RNA-sequencing data derived from the cancer genome atlas (TCGA). Specifically, our strategy allowed for an unbiased identification of genes whose expression was closely associated with a set of bona fide stroma-specific transcripts, namely the interstitial collagens COL1A1, COL1A2, and COL3A1. Among the top hits were genes involved in cellular matrix remodeling and tumor invasion and migration, including platelet-derived growth factor receptor beta (PDGFRβ), which was found to be the highest-ranking receptor protein genome-wide. Similar analyses performed on ten additional TCGA cancer datasets revealed that other tumor types shared CAF markers with OSCC, including PDGFRβ, which was found to significantly correlate with the reference collagen expression in ten of the 11 cancer types tested. Subsequent immunostaining of OSCC specimens demonstrated that PDGFRβ was abundantly expressed in stromal fibroblasts of all tested cases (12/12), while it was absent in tumor cells, with greater specificity than other known markers such as alpha smooth muscle actin or podoplanin (3/11). Overall, this study identified PDGFRβ as a novel marker of stromal activation in OSCC, and further characterized a list of promising candidate CAF markers that may be relevant to other carcinomas. Our novel approach provides for a fast and accurate method to identify CAF markers without the need for

  8. Asymmetric Dimethylarginine, Endothelial Dysfunction and Renal Disease

    PubMed Central

    Aldámiz-Echevarría, Luis; Andrade, Fernando

    2012-01-01

    l-Arginine (Arg) is oxidized to l-citrulline and nitric oxide (NO) by the action of endothelial nitric oxide synthase (NOS). In contrast, protein-incorporated Arg residues can be methylated with subsequent proteolysis giving rise to methylarginine compounds, such as asymmetric dimethylarginine (ADMA) that competes with Arg for binding to NOS. Most ADMA is degraded by dimethylarginine dimethyaminohydrolase (DDAH), distributed widely throughout the body and regulates ADMA levels and, therefore, NO synthesis. In recent years, several studies have suggested that increased ADMA levels are a marker of atherosclerotic change, and can be used to assess cardiovascular risk, consistent with ADMA being predominantly absorbed by endothelial cells. NO is an important messenger molecule involved in numerous biological processes, and its activity is essential to understand both pathogenic and therapeutic mechanisms in kidney disease and renal transplantation. NO production is reduced in renal patients because of their elevated ADMA levels with associated reduced DDAH activity. These factors contribute to endothelial dysfunction, oxidative stress and the progression of renal damage, but there are treatments that may effectively reduce ADMA levels in patients with kidney disease. Available data on ADMA levels in controls and renal patients, both in adults and children, also are summarized in this review. PMID:23109853

  9. Method for in vitro differentiation of bone marrow mesenchymal stem cells into endothelial progenitor cells and vascular endothelial cells

    PubMed Central

    Wang, Qihong; Zhang, Weifeng; He, Guifen; Sha, Huifang; Quan, Zhe

    2016-01-01

    Vascular development is a regulated process and is dependent on the participation and differentiation of many cell types including the proliferation and migration of vascular endothelial cells and differentiation of endothelial progenitor cells (EPCs) to mesodermal precursor cells. Thus, reconstitution of this process in vitro necessitates providing ambient conditions for generating and culturing EPCs in vitro and differentiating them to vascular endothelial cells. In the present study, we developed methods to differentiate bone marrow mesenchymal stem cells (MSC) into EPCs and to vascular endothelial cells. Bone marrow MSC from canines and human sources were differentiated in vitro in to EPCs. These EPCs were able to express a variety of endothelial markers following 7 days in culture. Further culturing led to the appearance of an increased number and proportion of endothelial cells. These cells were stable even after 30 generations in culture. There was a gradual loss of CD31 and increased expression of factor VIII, VEGFR and CD133. VEGF being highly angiogenic, helps in the vascular development. These results provide the basis for the possible development of vasculature in vitro conditions for biomedical applications and in vivo for organ/tissue reconstruction therapies. PMID:27878275

  10. [Transplantation of corneal endothelial cells].

    PubMed

    Amano, Shiro

    2002-12-01

    endothelial cells. Cultured rabbit corneal endothelial cells that endocytosed iron were injected into the anterior chamber of rabbits whose corneal endothelium was cryo-injured, and were pulled to Descemet's membrane by putting a magnet on the eyelid. In these rabbits, corneal edema decreased more quickly than in the control group and no intraocular pressure rise was observed during 8 weeks after the operation, suggesting that the direct delivery of cultured HCECs into the anterior chamber can be an alternative method of choice. The following obstacles should be addressed to make the transplantation of cultured corneal endothelial cells clinically applicable. 1. To reconstruct a cornea that is the same as or superior to the normal cornea, more innovation is necessary in the method of culturing and seeding HCECs. We should consider utilizing HCECs obtained from fetuses after clearing ethical issues. Moreover, we need to develop a method to enhance the cell density and the cell functions. 2. Porcine corneal stroma is promising as a carrier of HCECs instead of human corneal stroma, which is in very limited supply. The usefulness of porcine corneal stroma acellularized to prevent retrovirus infection should be evaluated. 3. To make the self immature cells applicable to corneal transplantation, we should elucidate the corneal endothelial cell specific markers and the factors that are necessary to induce self immature cells to become corneal endothelial cells. 4. The direct delivery of cultured HCECs into the anterior chamber can be an alternative method of choice when its long-term safety is confirmed.

  11. Increased circulating inflammatory endothelial cells in blacks with essential hypertension.

    PubMed

    Eirin, Alfonso; Zhu, Xiang-Yang; Woollard, John R; Herrmann, Sandra M; Gloviczki, Monika L; Saad, Ahmed; Juncos, Luis A; Calhoun, David A; Rule, Andrew D; Lerman, Amir; Textor, Stephen C; Lerman, Lilach O

    2013-09-01

    Morbidity and mortality attributable to hypertension are higher in black essential hypertensive (EH) compared with white EH patients, possibly related to differential effects on vascular injury and repair. Although circulating endothelial progenitor cells (EPCs) preserve endothelial integrity, inflammatory endothelial cells (IECs) detach from sites of injury and represent markers of vascular damage. We hypothesized that blood levels of IECs and inflammatory markers would be higher in black EH compared with white EH patients. Inferior vena cava and renal vein levels of CD34+/KDR+ (EPC) and VAP-1+ (IEC) cells were measured by fluorescence-activated cell sorting in white EH and black EH patients under fixed sodium intake and blockade of the renin-angiotensin system, and compared with systemic levels in normotensive control subjects (n=19 each). Renal vein and inferior vena cava levels of inflammatory cytokines and EPC homing factors were measured by Luminex. Blood pressure, serum creatinine, lipids, and antihypertensive medications did not differ between white and black EH patients, and EPC levels were decreased in both. Circulating IEC levels were elevated in black EH patients, and inversely correlated with EPC levels (R(2)=0.58; P=0.0001). Systemic levels of inflammatory cytokines and EPC homing factors were higher in black EH compared with white EH patients, and correlated directly with IECs. Renal vein inflammatory cytokines, EPCs, and IECs did not differ from their circulating levels. Most IECs expressed endothelial markers, fewer expressed progenitor cell markers, but none showed lymphocyte or phagocytic cell markers. Thus, increased release of cytokines and IECs in black EH patients may impair EPC reparative capacity and aggravate vascular damage, and accelerate hypertension-related complications.

  12. Methylglyoxal promotes oxidative stress and endothelial dysfunction.

    PubMed

    Sena, Cristina M; Matafome, Paulo; Crisóstomo, Joana; Rodrigues, Lisa; Fernandes, Rosa; Pereira, Paulo; Seiça, Raquel M

    2012-05-01

    Modern diets can cause modern diseases. Research has linked a metabolite of sugar, methylglyoxal (MG), to the development of diabetic complications, but the exact mechanism has not been fully elucidated. The present study was designed to investigate whether MG could directly influence endothelial function, oxidative stress and inflammation in Wistar and Goto-Kakizaki (GK) rats, an animal model of type 2 diabetes. Wistar and GK rats treated with MG in the drinking water for 3 months were compared with the respective control rats. The effects of MG were investigated on NO-dependent vasorelaxation in isolated rat aortic arteries from the different groups. Insulin resistance, NO bioavailability, glycation, a pro-inflammatory biomarker monocyte chemoattractant protein-1 (MCP-1) and vascular oxidative stress were also evaluated. Methylglyoxal treated Wistar rats significantly reduced the efficacy of NO-dependent vasorelaxation (p<0.001). This impairment was accompanied by a three fold increase in the oxidative stress marker nitrotyrosine. Advanced glycation endproducts (AGEs) formation was significantly increased as well as MCP-1 and the expression of the receptor for AGEs (RAGE). NO bioavailability was significantly attenuated and accompanied by an increase in superoxide anion immunofluorescence. Methylglyoxal treated GK rats significantly aggravated endothelial dysfunction, oxidative stress, AGEs accumulation and diminished NO bioavailability when compared with control GK rats. These results indicate that methylglyoxal induced endothelial dysfunction in normal Wistar rats and aggravated the endothelial dysfunction present in GK rats. The mechanism is at least in part by increasing oxidative stress and/or AGEs formation with a concomitant increment of inflammation and a decrement in NO bioavailability. The present study provides further evidence for methylglyoxal as one of the causative factors in the pathogenesis of atherosclerosis and development of macrovascular

  13. Inflammation induces neuro-lymphatic protein expression in multiple sclerosis brain neurovasculature

    PubMed Central

    2013-01-01

    Background Multiple sclerosis (MS) is associated with ectopic lymphoid follicle formation. Podoplanin+ (lymphatic marker) T helper17 (Th17) cells and B cell aggregates have been implicated in the formation of tertiary lymphoid organs (TLOs) in MS and experimental autoimmune encephalitis (EAE). Since podoplanin expressed by Th17 cells in MS brains is also expressed by lymphatic endothelium, we investigated whether the pathophysiology of MS involves inductions of lymphatic proteins in the inflamed neurovasculature. Methods We assessed the protein levels of lymphatic vessel endothelial hyaluronan receptor and podoplanin, which are specific to the lymphatic system and prospero-homeobox protein-1, angiopoietin-2, vascular endothelial growth factor-D, vascular endothelial growth factor receptor-3, which are expressed by both lymphatic endothelium and neurons. Levels of these proteins were measured in postmortem brains and sera from MS patients, in the myelin proteolipid protein (PLP)-induced EAE and Theiler’s murine encephalomyelitis virus (TMEV) induced demyelinating disease (TMEV-IDD) mouse models and in cell culture models of inflamed neurovasculature. Results and conclusions Intense staining for LYVE-1 was found in neurons of a subset of MS patients using immunohistochemical approaches. The lymphatic protein, podoplanin, was highly expressed in perivascular inflammatory lesions indicating signaling cross-talks between inflamed brain vasculature and lymphatic proteins in MS. The profiles of these proteins in MS patient sera discriminated between relapsing remitting MS from secondary progressive MS and normal patients. The in vivo findings were confirmed in the in vitro cell culture models of neuroinflammation. PMID:24124909

  14. Tumor endothelial cells express high pentraxin 3 levels.

    PubMed

    Hida, Kyoko; Maishi, Nako; Kawamoto, Taisuke; Akiyama, Kosuke; Ohga, Noritaka; Hida, Yasuhiro; Yamada, Kenji; Hojo, Takayuki; Kikuchi, Hiroshi; Sato, Masumi; Torii, Chisaho; Shinohara, Nobuo; Shindoh, Masanobu

    2016-12-01

    It has been described that tumor progression has many similarities to inflammation and wound healing in terms of the signaling processes involved. Among biological responses, angiogenesis, which is necessary for tumor progression and metastasis, is a common hallmark; therefore, tumor blood vessels have been considered as important therapeutic targets in anticancer therapy. We focused on pentraxin 3 (PTX3), which is a marker of cancer-related inflammation, but we found no reports on its expression and function in tumor blood vessels. Here we showed that PTX3 is expressed in mouse and human tumor blood vessels based on immunohistochemical analysis. We found that PTX3 is upregulated in primary mouse and human tumor endothelial cells compared to normal endothelial cells. We also showed that PTX3 plays an important role in the proliferation of the tumor endothelial cells. These results suggest that PTX3 is an important target for antiangiogenic therapy.

  15. Cell trafficking of endothelial progenitor cells in tumor progression.

    PubMed

    de la Puente, Pilar; Muz, Barbara; Azab, Feda; Azab, Abdel Kareem

    2013-07-01

    Blood vessel formation plays an essential role in many physiologic and pathologic processes, including normal tissue growth and healing, as well as tumor progression. Endothelial progenitor cells (EPC) are a subtype of stem cells with high proliferative potential that are capable of differentiating into mature endothelial cells, thus contributing to neovascularization in tumors. In response to tumor-secreted cytokines, EPCs mobilize from the bone marrow to the peripheral blood, home to the tumor site, and differentiate to mature endothelial cells and secrete proangiogenic factors to facilitate vascularization of tumors. In this review, we summarize the expression of surface markers, cytokines, receptors, adhesion molecules, proteases, and cell signaling mechanisms involved in the different steps (mobilization, homing, and differentiation) of EPC trafficking from the bone marrow to the tumor site. Understanding the biologic mechanisms of EPC cell trafficking opens a window for new therapeutic targets in cancer.

  16. Obesity-induced Endoplasmic Reticulum Stress Causes Lung Endothelial Dysfunction and Promotes Acute Lung Injury.

    PubMed

    Shah, Dilip; Romero, Freddy; Guo, Zhi; Sun, Jianxin; Li, Jonathan; Kallen, Caleb B; Naik, Ulhas P; Summer, Ross

    2017-03-09

    Obesity is a significant risk factor for the acute respiratory distress syndrome (ARDS). The mechanisms underlying this association are unknown. We recently showed that diet-induced obese (DIO) mice exhibit pulmonary vascular endothelial dysfunction which is associated with enhanced susceptibility to lipopolysaccharide (LPS)-induced lung injury. Here, we demonstrate that lung endothelial dysfunction in DIO mice coincides with increased endoplasmic reticulum (ER) stress. Specifically, we observed enhanced expression of the major sensors of misfolded proteins including PERK, IREα and ATF6, in whole lung and in lung endothelial cells isolated from DIO mice. Further, we found that lung endothelial cells exposed to serum from obese mice, or to saturated fatty acids that mimic obese serum, resulted in enhanced expression of markers of ER stress and the induction of other biological responses that typify the lung endothelium of DIO mice. Similar changes were observed in lung endothelial cells and in whole lung tissue after exposure to tunicamycin, a compound that causes ER stress by blocking N-linked glycosylation; indicating that ER stress causes endothelial dysfunction in the lung. Treatment with 4-PBA, a chemical protein chaperone that reduces ER stress, restored vascular endothelial cell expression of adhesion molecules and protected against LPS-induced acute lung injury in DIO mice. Our work indicates that fatty acids in obese serum induce ER stress in the pulmonary endothelium leading to pulmonary endothelial cell dysfunction. Our work suggests that reducing protein load in the endoplasmic reticulum of pulmonary endothelial cells might protect against ARDS in obese individuals.

  17. Endothelial cells and human cerebral small vessel disease.

    PubMed

    Hainsworth, Atticus H; Oommen, Asho T; Bridges, Leslie R

    2015-01-01

    Brain endothelial cells have unique properties in terms of barrier function, local molecular signaling, regulation of local cerebral blood flow (CBF) and interactions with other members of the neurovascular unit. In cerebral small vessel disease (arteriolosclerosis; SVD), the endothelial cells in small arteries survive, even when mural pathology is advanced and myocytes are severely depleted. Here, we review aspects of altered endothelial functions that have been implicated in SVD: local CBF dysregulation, endothelial activation and blood-brain barrier (BBB) dysfunction. Reduced CBF is reported in the diffuse white matter lesions that are a neuroradiological signature of SVD. This may reflect an underlying deficit in local CBF regulation (possibly via the nitric oxide/cGMP signaling pathway). While many laboratories have observed an association of symptomatic SVD with serum markers of endothelial activation, it is apparent that the origin of these circulating markers need not be brain endothelium. Our own neuropathology studies did not confirm local endothelial activation in small vessels exhibiting SVD. Local BBB failure has been proposed as a cause of SVD and associated parenchymal lesions. Some groups find that computational analyses of magnetic resonance imaging (MRI) scans, following systemic injection of a gadolinium-based contrast agent, suggest that extravasation into brain parenchyma is heightened in people with SVD. Our recent histochemical studies of donated brain tissue, using immunolabeling for large plasma proteins [fibrinogen, immunoglobulin G (IgG)], do not support an association of SVD with recent plasma protein extravasation. It is possible that a trigger leakage episode, or a size-selective loosening of the BBB, participates in SVD pathology.

  18. Lymphatic Endothelial Cells Produce M-CSF, Causing Massive Bone Loss in Mice.

    PubMed

    Wang, Wensheng; Wang, Hua; Zhou, Xichao; Li, Xing; Sun, Wen; Dellinger, Michael; Boyce, Brendan F; Xing, Lianping

    2017-01-04

    Gorham-Stout disease (GSD) is a rare bone disorder characterized by aggressive osteolysis associated with lymphatic vessel invasion within bone marrow cavities. The etiology of GSD is not known, and there is no effective therapy or animal model for the disease. Here, we investigated if lymphatic endothelial cells (LECs) affect osteoclasts (OCs) to cause a GSD osteolytic phenotype in mice. We examined the effect of a mouse LEC line on osteoclastogenesis in co-cultures. LECs significantly increased receptor activator of NF-κB ligand (RANKL)-mediated OC formation and bone resorption. LECs expressed high levels of macrophage colony-stimulating factor (M-CSF), but not RANKL, interleukin-6 (IL-6), and tumor necrosis factor (TNF). LEC-mediated OC formation and bone resorption were blocked by an M-CSF neutralizing antibody or Ki20227, an inhibitor of the M-CSF receptor, c-Fms. We injected LECs into the tibias of wild-type (WT) mice and observed massive osteolysis on X-ray and micro-CT scans. Histology showed that LEC-injected tibias had significant trabecular and cortical bone loss and increased OC numbers. M-CSF protein levels were significantly higher in serum and bone marrow plasma of mice given intra-tibial LEC injections. Immunofluorescence staining showed extensive replacement of bone and marrow by podoplanin+ LECs. Treatment of LEC-injected mice with Ki20227 significantly decreased tibial bone destruction. In addition, lymphatic vessels in a GSD bone sample were stained positively for M-CSF. Thus, LECs cause bone destruction in vivo in mice by secreting M-CSF, which promotes OC formation and activation. Blocking M-CSF signaling may represent a new therapeutic approach for treatment of patients with GSD. Furthermore, tibial injection of LECs is a useful mouse model to study GSD. © 2017 American Society for Bone and Mineral Research.

  19. Lung endothelial cells strengthen, but brain endothelial cells weaken barrier properties of a human alveolar epithelium cell culture model.

    PubMed

    Neuhaus, Winfried; Samwer, Fabian; Kunzmann, Steffen; Muellenbach, Ralf M; Wirth, Michael; Speer, Christian P; Roewer, Norbert; Förster, Carola Y

    2012-11-01

    The blood-air barrier in the lung consists of the alveolar epithelium, the underlying capillary endothelium, their basement membranes and the interstitial space between the cell layers. Little is known about the interactions between the alveolar and the blood compartment. The aim of the present study was to gain first insights into the possible interplay between these two neighbored cell layers. We established an in vitro Transwell model of the alveolar epithelium based on human cell line H441 and investigated the influence of conditioned medium obtained from human lung endothelial cell line HPMEC-ST1.6R on the barrier properties of the H441 layers. As control for tissue specificity H441 layers were exposed to conditioned medium from human brain endothelial cell line hCMEC/D3. Addition of dexamethasone was necessary to obtain stable H441 cell layers. Moreover, dexamethasone increased expression of cell type I markers (caveolin-1, RAGE) and cell type II marker SP-B, whereas decreased the transepithelial electrical resistance (TEER) in a concentration dependent manner. Soluble factors obtained from the lung endothelial cell line increased the barrier significantly proven by TEER values and fluorescein permeability on the functional level and by the differential expression of tight junctional proteins on the molecular level. In contrast to this, soluble factors derived from brain endothelial cells weakened the barrier significantly. In conclusion, soluble factors from lung endothelial cells can strengthen the alveolar epithelium barrier in vitro, which suggests communication between endothelial and epithelial cells regulating the integrity of the blood-air barrier.

  20. Endothelial RIG-I activation impairs endothelial function

    SciTech Connect

    Asdonk, Tobias; Nickenig, Georg; Zimmer, Sebastian

    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.

  1. The Endothelial Glycocalyx: New Diagnostic and Therapeutic Approaches in Sepsis

    PubMed Central

    Koczera, Patrick

    2016-01-01

    Sepsis is defined as a life-threatening organ dysfunction caused by a dysregulated host response to infection. The endothelial glycocalyx is one of the earliest sites involved during sepsis. This fragile layer is a complex network of cell-bound proteoglycans, glycosaminoglycan side chains, and sialoproteins lining the luminal side of endothelial cells with a thickness of about 1 to 3 μm. Sepsis-associated alterations of its structure affect endothelial permeability and result in the liberation of endogenous damage-associated molecular patterns (DAMPs). Once liberated in the circulatory system, DAMPs trigger the devastating consequences of the proinflammatory cascades in sepsis and septic shock. In this way, the injury to the glycocalyx with the consecutive release of DAMPs contributes to a number of specific clinical effects of sepsis, including acute kidney injury, respiratory failure, and septic cardiomyopathy. Moreover, the extent of glycocalyx degradation serves as a marker of endothelial dysfunction and sepsis severity. In this review, we highlight the crucial role of the glycocalyx in sepsis as a diagnostic tool and discuss the potential of members of the endothelial glycocalyx serving as hopeful therapeutic targets in sepsis-associated multiple organ failures. PMID:27699168

  2. Endothelial dysfunction and tendinopathy: how far have we come?

    PubMed

    Papalia, R; Moro, L; Franceschi, F; Albo, E; D'Adamio, S; Di Martino, A; Vadalà, G; Faldini, C; Denaro, V

    2013-12-01

    Symptomatic tendon tears are one of the most important causes of pain and joint dysfunction. Among the intrinsic causes, vascularization recently gained a major role. Endothelial function is indeed a key factor, as well as vascular tone and thrombotic factors, in the regulation of vascular homeostasis and the composition of vascular wall. In this review, we studied systematically whether there is a relationship between endothelial dysfunction and tendinopathy. A literature search was performed using the isolated or combined keywords endothelial dysfunction and tendon,' 'nitric oxide (NO) and tendinopathy,' and 'endothelial dysfunction in tendon healing.' We identified 21 published studies. Of the selected studies, 9 were in vivo studies, 2 focusing on animals and 7 on humans, while 12 reported about in vitro evaluations, where 7 were carried out on humans and 5 on animals. The evidence about a direct relationship between tendinopathy and endothelial dysfunction is still poor. As recent studies have shown, there is no significant improvement in clinical and functional assessments after treatment with NO in patients suffering from tendinopathy in different locations. No significant differences were identified in the outcomes reported for experiment group when compared with controls treated with conventional surgical procedures or rehabilitation programs. Nitric oxide could be a marker to quantify the response of the endothelium to mechanical stress or hypoxia indicating the final balance between vasodilatating and vasoconstricting factors and their effects, but more ad stronger evidence is still needed to fully support this practice.

  3. Phenotypic and Functional Changes of Endothelial and Smooth Muscle Cells in Thoracic Aortic Aneurysms

    PubMed Central

    Malashicheva, Anna; Kostina, Daria; Kostina, Aleksandra; Irtyuga, Olga; Voronkina, Irina; Smagina, Larisa; Ignatieva, Elena; Gavriliuk, Natalia; Uspensky, Vladimir; Moiseeva, Olga; Vaage, Jarle; Kostareva, Anna

    2016-01-01

    Thoracic aortic aneurysm develops as a result of complex series of events that alter the cellular structure and the composition of the extracellular matrix of the aortic wall. The purpose of the present work was to study the cellular functions of endothelial and smooth muscle cells from the patients with aneurysms of the thoracic aorta. We studied endothelial and smooth muscle cells from aneurysms in patients with bicuspid aortic valve and with tricuspid aortic valve. The expression of key markers of endothelial (CD31, vWF, and VE-cadherin) and smooth muscle (SMA, SM22α, calponin, and vimentin) cells as well extracellular matrix and MMP activity was studied as well as and apoptosis and cell proliferation. Expression of functional markers of endothelial and smooth muscle cells was reduced in patient cells. Cellular proliferation, migration, and synthesis of extracellular matrix proteins are attenuated in the cells of the patients. We show for the first time that aortic endothelial cell phenotype is changed in the thoracic aortic aneurysms compared to normal aortic wall. In conclusion both endothelial and smooth muscle cells from aneurysms of the ascending aorta have downregulated specific cellular markers and altered functional properties, such as growth rate, apoptosis induction, and extracellular matrix synthesis. PMID:26904289

  4. Phenotypic and Functional Changes of Endothelial and Smooth Muscle Cells in Thoracic Aortic Aneurysms.

    PubMed

    Malashicheva, Anna; Kostina, Daria; Kostina, Aleksandra; Irtyuga, Olga; Voronkina, Irina; Smagina, Larisa; Ignatieva, Elena; Gavriliuk, Natalia; Uspensky, Vladimir; Moiseeva, Olga; Vaage, Jarle; Kostareva, Anna

    2016-01-01

    Thoracic aortic aneurysm develops as a result of complex series of events that alter the cellular structure and the composition of the extracellular matrix of the aortic wall. The purpose of the present work was to study the cellular functions of endothelial and smooth muscle cells from the patients with aneurysms of the thoracic aorta. We studied endothelial and smooth muscle cells from aneurysms in patients with bicuspid aortic valve and with tricuspid aortic valve. The expression of key markers of endothelial (CD31, vWF, and VE-cadherin) and smooth muscle (SMA, SM22α, calponin, and vimentin) cells as well extracellular matrix and MMP activity was studied as well as and apoptosis and cell proliferation. Expression of functional markers of endothelial and smooth muscle cells was reduced in patient cells. Cellular proliferation, migration, and synthesis of extracellular matrix proteins are attenuated in the cells of the patients. We show for the first time that aortic endothelial cell phenotype is changed in the thoracic aortic aneurysms compared to normal aortic wall. In conclusion both endothelial and smooth muscle cells from aneurysms of the ascending aorta have downregulated specific cellular markers and altered functional properties, such as growth rate, apoptosis induction, and extracellular matrix synthesis.

  5. Methodological issues in the assessment of skin microvascular endothelial function in humans.

    PubMed

    Cracowski, Jean-Luc; Minson, Christopher T; Salvat-Melis, Muriel; Halliwill, John R

    2006-09-01

    The study of microvascular function can be performed in humans using laser Doppler flowmetry of the skin. This technology lends itself to a wide range of applications for studying the endothelial function of skin blood vessels. We review the advantages and limitations of postocclusive hyperemia, local thermal hyperemia, acetylcholine iontophoresis, flowmotion and association with microdialysis as tools with which to investigate skin microvascular endothelial function in humans. Postocclusive hyperemia, thermal hyperemia and acetylcholine iontophoresis provide integrated indexes of microvascular function rather than specific endothelial markers. However, they are valuable tools and can be used as surrogate endpoints in clinical trials in which the assessment of microvascular function in humans is required.

  6. Origins of circulating endothelial cells and endothelial outgrowth from blood

    PubMed Central

    Lin, Yi; Weisdorf, Daniel J.; Solovey, Anna; Hebbel, Robert P.

    2000-01-01

    Normal adults have a small number of circulating endothelial cells (CEC) in peripheral blood, and endothelial outgrowth has been observed from cultures of blood. In this study we seek insight into the origins of CEC and endothelial outgrowth from cultures of blood. Fluorescence in situ hybridization analysis of blood samples from bone marrow transplant recipients who had received gender-mismatched transplants 5–20 months earlier showed that most CEC in fresh blood had recipient genotype. Endothelial outgrowth from the same blood samples after 9 days in culture (5-fold expansion) was still predominantly of the recipient genotype. In contrast, endothelial outgrowth after ∼1 month (102-fold expansion) was mostly of donor genotype. Thus, recipient-genotype endothelial cells expanded only ∼20-fold over this period, whereas donor-genotype endothelial cells expanded ∼1000-fold. These data suggest that most CEC in fresh blood originate from vessel walls and have limited growth capability. Conversely, the data indicate that outgrowth of endothelial cells from cultures of blood is mostly derived from transplantable marrow-derived cells. Because these cells have more delayed outgrowth but a greater proliferative rate, our data suggest that they are derived from circulating angioblasts. PMID:10619863

  7. Efficient Generation of Human Embryonic Stem Cell-Derived Corneal Endothelial Cells by Directed Differentiation

    PubMed Central

    McCabe, Kathryn L.; Kunzevitzky, Noelia J.; Chiswell, Brian P.; Xia, Xin; Goldberg, Jeffrey L.; Lanza, Robert

    2015-01-01

    Aim To generate human embryonic stem cell derived corneal endothelial cells (hESC-CECs) for transplantation in patients with corneal endothelial dystrophies. Materials and Methods Feeder-free hESC-CECs were generated by a directed differentiation protocol. hESC-CECs were characterized by morphology, expression of corneal endothelial markers, and microarray analysis of gene expression. Results hESC-CECs were nearly identical morphologically to primary human corneal endothelial cells, expressed Zona Occludens 1 (ZO-1) and Na+/K+ATPaseα1 (ATPA1) on the apical surface in monolayer culture, and produced the key proteins of Descemet’s membrane, Collagen VIIIα1 and VIIIα2 (COL8A1 and 8A2). Quantitative PCR analysis revealed expression of all corneal endothelial pump transcripts. hESC-CECs were 96% similar to primary human adult CECs by microarray analysis. Conclusion hESC-CECs are morphologically similar, express corneal endothelial cell markers and express a nearly identical complement of genes compared to human adult corneal endothelial cells. hESC-CECs may be a suitable alternative to donor-derived corneal endothelium. PMID:26689688

  8. Characterization and comparison of embryonic stem cell-derived KDR+ cells with endothelial cells.

    PubMed

    Sun, Xuan; Cheng, Lamei; Duan, Huaxin; Lin, Ge; Lu, Guangxiu

    2012-09-01

    Growing interest in utilizing endothelial cells (ECs) for therapeutic purposes has led to the exploration of human embryonic stem cells (hESCs) as a potential source for endothelial progenitors. In this study, ECs were induced from hESC lines and their biological characteristics were analyzed and compared with both cord blood endothelial progenitor cells (CBEPCs) and human umbilical vein endothelial cells (HUVECs) in vitro. The results showed that isolated embryonic KDR+ cells (EC-KDR+) display characteristics that were similar to CBEPCs and HUVECs. EC-KDR+, CBEPCs and HUVECs all expressed CD31 and CD144, incorporated DiI-Ac-LDL, bound UEA1 lectin, and were able to form tube-like structures on Matrigel. Compared with CBEPCs and HUVECs, the expression level of endothelial progenitor cell markers such as CD133 and KDR in EC-KDR+ was significantly higher, while the mature endothelial marker vWF was lowly expressed in EC-KDR+. In summary, the study showed that EC-KDR+ are primitive endothelial-like progenitors and might be a potential source for therapeutic vascular regeneration and tissue engineering.

  9. Interaction of recombinant octameric hemoglobin with endothelial cells.

    PubMed

    Gaucher, Caroline; Domingues-Hamdi, Élisa; Prin-Mathieu, Christine; Menu, Patrick; Baudin-Creuza, Véronique

    2015-02-01

    Hemoglobin-based oxygen carriers (HBOCs) may generate oxidative stress, vasoconstriction and inflammation. To reduce these undesirable vasoactive properties, we increased hemoglobin (Hb) molecular size by genetic engineering with octameric Hb, recombinant (r) HbβG83C. We investigate the potential side effects of rHbβG83C on endothelial cells. The rHbβG83C has no impact on cell viability, and induces a huge repression of endothelial nitric oxide synthase gene transcription, a marker of vasomotion. No induction of Intermolecular-Adhesion Molecule 1 and E-selectin (inflammatory markers) transcription was seen. In the presence of rHbβG83C, the transcription of heme oxygenase-1 (oxidative stress marker) is weakly increased compared to the two other HBOCs (references) or Voluven (control). This genetically engineered octameric Hb, based on a human Hb βG83C mutant, leads to little impact at the level of endothelial cell inflammatory response and thus appears as an interesting molecule for HBOC development.

  10. Tumor Endothelial Cells

    PubMed Central

    Dudley, Andrew C.

    2012-01-01

    The vascular endothelium is a dynamic cellular “organ” that controls passage of nutrients into tissues, maintains the flow of blood, and regulates the trafficking of leukocytes. In tumors, factors such as hypoxia and chronic growth factor stimulation result in endothelial dysfunction. For example, tumor blood vessels have irregular diameters; they are fragile, leaky, and blood flow is abnormal. There is now good evidence that these abnormalities in the tumor endothelium contribute to tumor growth and metastasis. Thus, determining the biological basis underlying these abnormalities is critical for understanding the pathophysiology of tumor progression and facilitating the design and delivery of effective antiangiogenic therapies. PMID:22393533

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

    PubMed Central

    Kono, Ken; Hiruma, Hitomi; Kobayashi, Shingo; Sato, Yoji; Tanaka, Masaru; Sawada, Rumi; Niimi, Shingo

    2016-01-01

    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. PMID:27348615

  12. Endothelial to mesenchymal transition contributes to arsenic-trioxide-induced cardiac fibrosis

    PubMed Central

    Zhang, Yong; Wu, Xianxian; Li, Yang; Zhang, Haiying; Li, Zhange; Zhang, Ying; Zhang, Longyin; Ju, Jiaming; Liu, Xin; Chen, Xiaohui; Glybochko, Peter V.; Nikolenko, Vladimir; Kopylov, Philipp; Xu, Chaoqian; Yang, Baofeng

    2016-01-01

    Emerging evidence has suggested the critical role of endothelial to mesenchymal transition (EndMT) in fibrotic diseases. The present study was designed to examine whether EndMT is involved in arsenic trioxide (As2O3)-induced cardiac fibrosis and to explore the underlying mechanisms. Cardiac dysfunction was observed in rats after exposure to As2O3 for 15 days using echocardiography, and the deposition of collagen was detected by Masson’s trichrome staining and electron microscope. EndMT was indicated by the loss of endothelial cell markers (VE-cadherin and CD31) and the acquisition of mesenchymal cell markers (α-SMA and FSP1) determined by RT-PCR at the mRNA level and Western blot and immunofluorescence analysis at the protein level. In the in-vitro experiments, endothelial cells acquired a spindle-shaped morphology accompanying downregulation of the endothelial cell markers and upregulation of the mesenchymal cell markers when exposed to As2O3. As2O3 activated the AKT/GSK-3β/Snail signaling pathway, and blocking this pathway with PI3K inhibitor (LY294002) abolished EndMT in As2O3-treated endothelial cells. Our results highlight that As2O3 is an EndMT-promoting factor during cardiac fibrosis, suggesting that targeting EndMT is beneficial for preventing As2O3-induced cardiac toxicity. PMID:27671604

  13. STUDIES ON ENDOTHELIAL REACTIONS

    PubMed Central

    Foot, Nathan Chandler

    1920-01-01

    1. The injection of a colloidal suspension, or sol, of carbon into the veins of a living animal, as recommended by McJunkin, furnishes an apparently reliable means of tracing the so called epithelioid cell of the pulmonary tubercle from its origin in the vascular endothelium to the lesion. 2. Experimental tubercles are formed in the lung, as in the liver, primarily by cells originating in the capillary endothelium. These cells are probably present in small numbers in the normal lung, lying free both in the alveolar wall and the air vesicles. In response to infection they proliferate in the capillary walls in the vicinity of the invading organisms, migrate in steadily increasing numbers, and, arriving at the site of the infection, further multiply and to some extent fuse to form the syncytia known as giant cells. 3. The epithelial cell takes no active part in the process; its proliferation tends to repair denuded surfaces and is regenerative rather than combative or phagocytic in nature. This cell is free from carbon and stains only diffusely with carmine, in contradistinction to the endothelial cell which readily takes up both pigments in granular form. 4. The cells of endothelial origin not only phagocytose tubercle bacilli, but carry them into the tissues, for example into lymph nodes, by way of the lymphatics, or into other lung lobules by way of the air passages, in which they are readily demonstrable. PMID:19868459

  14. Early radiation-induced endothelial cell loss and blood-spinal cord barrier breakdown in the rat spinal cord.

    PubMed

    Li, Yu-Qing; Chen, Paul; Jain, Vipan; Reilly, Raymond M; Wong, C Shun

    2004-02-01

    Using a rat spinal cord model, this study was designed to characterize radiation-induced vascular endothelial cell loss and its relationship to early blood-brain barrier disruption in the central nervous system. Adult rats were given a single dose of 0, 2, 8, 19.5, 22, 30 or 50 Gy to the cervical spinal cord. At various times up to 2 weeks after irradiation, the spinal cord was processed for histological and immunohistochemical analysis. Radiation-induced apoptosis was assessed by morphology and TdT-mediated dUTP nick end labeling combined with immunohistochemical markers for endothelial and glial cells. Image analysis was performed to determine endothelial cell and microvessel density using immunohistochemistry with endothelial markers, namely endothelial barrier antigen, glucose transporter isoform 1, laminin and zonula occludens 1. Blood-spinal cord barrier permeability was assessed using immunohistochemistry for albumin and (99m)Tc-diethylenetriamine pentaacetic acid as a vascular tracer. Endothelial cell proliferation was assessed using in vivo BrdU labeling. During the first 24 h after irradiation, apoptotic endothelial cells were observed in the rat spinal cord. The decrease in endothelial cell density at 24 h after irradiation was associated with an increase in albumin immunostaining around microvessels. The decrease in the number of endothelial cells persisted for 7 days and recovery of endothelial density was apparent by day 14. A similar pattern of blood-spinal cord barrier disruption and recovery of permeability was observed over the 2 weeks, and an increase in BrdU-labeled endothelial cells was seen at day 3. These results are consistent with an association between endothelial cell death and acute blood-spinal cord barrier disruption in the rat spinal cord after irradiation.

  15. Human Bone Derived Collagen for the Development of an Artificial Corneal Endothelial Graft. In Vivo Results in a Rabbit Model

    PubMed Central

    Vázquez, Natalia; Chacón, Manuel; Rodríguez-Barrientos, Carlos A.; Merayo-Lloves, Jesús; Naveiras, Miguel; Baamonde, Begoña; Alfonso, Jose F.; Zambrano-Andazol, Iriana; Riestra, Ana C.; Meana, Álvaro

    2016-01-01

    Corneal keratoplasty (penetrating or lamellar) using cadaveric human tissue, is nowadays the main treatment for corneal endotelial dysfunctions. However, there is a worldwide shortage of donor corneas available for transplantation and about 53% of the world’s population have no access to corneal transplantation. Generating a complete cornea by tissue engineering is still a tough goal, but an endothelial lamellar graft might be an easier task. In this study, we developed a tissue engineered corneal endothelium by culturing human corneal endothelial cells on a human purified type I collagen membrane. Human corneal endothelial cells were cultured from corneal rims after corneal penetrating keratoplasty and type I collagen was isolated from remnant cancellous bone chips. Isolated type I collagen was analyzed by western blot, liquid chromatography -mass spectrometry and quantified using the exponentially modified protein abundance index. Later on, collagen solution was casted at room temperature obtaining an optically transparent and mechanically manageable membrane that supports the growth of human and rabbit corneal endothelial cells which expressed characteristic markers of corneal endothelium: zonula ocluddens-1 and Na+/K+ ATPase. To evaluate the therapeutic efficiency of our artificial endothelial grafts, human purified type I collagen membranes cultured with rabbit corneal endothelial cells were transplanted in New Zealand white rabbits that were kept under a minimal immunosuppression regimen. Transplanted corneas maintained transparency for as long as 6 weeks without obvious edema or immune rejection and maintaining the same endothelial markers that in a healthy cornea. In conclusion, it is possible to develop an artificial human corneal endothelial graft using remnant tissues that are not employed in transplant procedures. This artificial endothelial graft can restore the integrality of corneal endothelium in an experimental model of endothelial dysfunction

  16. Na(+), K(+)-ATPase dysfunction causes cerebrovascular endothelial cell degeneration in rat prefrontal cortex slice cultures.

    PubMed

    Kurauchi, Yuki; Hisatsune, Akinori; Seki, Takahiro; Katsuki, Hiroshi

    2016-08-01

    Cerebrovascular endothelial cell dysfunction resulting in imbalance of cerebral blood flow contributes to the onset of psychiatric disorders such as depression, schizophrenia and bipolar disorder. Although decrease in Na(+), K(+)-ATPase activity has been reported in the patients with schizophrenia and bipolar disorder, the contribution of Na(+), K(+)-ATPase to endothelial cell dysfunction remains poorly understood. Here, by using rat neonatal prefrontal cortex slice cultures, we demonstrated that pharmacological inhibition of Na(+), K(+)-ATPase by ouabain induced endothelial cell injury. Treatment with ouabain significantly decreased immunoreactive area of rat endothelial cell antigen-1 (RECA-1), a marker of endothelial cells, in a time-dependent manner. Ouabain also decreased Bcl-2/Bax ratio and phosphorylation level of glycogen synthase kinase 3β (GSK3β) (Ser9), which were prevented by lithium carbonate. On the other hand, ouabain-induced endothelial cell injury was exacerbated by concomitant treatment with LY294002, an inhibitor of phosphoinositide 3- (PI3-) kinase. We also found that xestospongin C, an inhibitor of inositol triphosphate (IP3) receptor, but not SEA0400, an inhibitor of Na(+), Ca(2+) exchanger (NCX), protected endothelial cells from cytotoxicity of ouabain. These results suggest that cerebrovascular endothelial cell degeneration induced by Na(+), K(+)-ATPase inhibition resulting in Ca(2+) release from endoplasmic reticulum (ER) and activation of GSK3β signaling underlies pathogenesis of these psychiatric disorders.

  17. Endothelial cell permeability to water and antipyrine

    SciTech Connect

    Garrick, R.A.

    1986-03-05

    The endothelium provides a structural barrier between plasma constituents and the tissues. The permeability characteristics of the the endothelial cells regulate the transcellular movement of materials across this barrier while other movement is paracellular. In this study the permeability of the endothelial cells to tritiated water (/sup 3/HHO) and /sup 14/C-labeled antipyrine (AP) was investigated. The cells were isolated non-enzymatically from calf pulmonary artery and were maintained in culture and used between the seventh and fifteenth passage. The cells were removed from the T-flasks with a rubber policeman, titurated with a 22g needle and centrifuged. The cells were mixed with an extracellular marker, drawn into polyethylene tubing and packed by centrifugation for use in the linear diffusion technique. All measurements were made at 37 C. The diffusion coefficients for /sup 3/HHO through the packed cells (D), the intracellular material (D/sub 2/), and the extracellular material (D/sub 1/) were 0.682, 0.932 and 2.45 x 10/sup -5/ cm/sup 2/ s/sup -1/ and for AP were 0.273, 0.355 and 1.13 x 10/sup -5/ cm/sup 2/ s/sup -1/ respectively. The permeability coefficient calculated by the series-parallel pathway model for /sup 3/HHO was higher than that for AP and for both /sup 3/HHO and AP were lower than those calculated for isolated lung cells and erythrocytes.

  18. Mechanisms of tubulogenesis and endothelial phenotype expression by MSCs.

    PubMed

    Rytlewski, Julie A; Alejandra Aldon, M; Lewis, Evan W; Suggs, Laura J

    2015-05-01

    Stem cell-based therapies are a promising new avenue for treating ischemic disease and chronic wounds. Mesenchymal stem cells (MSCs) have a proven ability to augment the neovascularization processes necessary for wound healing and are widely popular as an autologous source of progenitor cells. Our lab has previously reported on PEGylated fibrin as a unique hydrogel that promotes spontaneous tubulogenesis of encapsulated MSCs without exogenous factors. However, the mechanisms underlying this process have remained unknown. To better understand the therapeutic value of PEGylated fibrin delivery of MSCs, we sought to clarify the relationship between biomaterial properties and cell behavior. Here we find that fibrin PEGylation does not dramatically alter the macroscopic mechanical properties of the fibrin-based matrix (less than 10% difference). It does, however, dramatically reduce the rate of diffusion through the gel matrix. PEGylated fibrin enhances the tubulogenic growth of encapsulated MSCs demonstrating fluid-filled lumens by interconnected MSCs. Image analysis gave a value of 4320 ± 1770 μm total network length versus 618 ± 443 μm for unmodified fibrin. PEGylation promotes the endothelial phenotype of encapsulated MSCs--compared to unmodified fibrin--as evidenced by higher levels of endothelial markers (von Willebrand factor, 2.2-fold; vascular endothelial cadherin, 1.8-fold) and vascular endothelial growth factor (VEGF, up to 1.8-fold). Prospective analysis of underlying molecular pathways demonstrated that this endothelial-like MSC behavior is sensitively modulated by hypoxic stress, but not VEGF supplementation as evidenced by a significant increase in VEGF and MMP-2 secretion per cell under hypoxia. Further gain-of-function studies under hypoxic stress demonstrated that hypoxia culture of MSCs in unmodified fibrin could increase both vWF and VE-cadherin levels to values that were not significantly different than cells cultured in PEGylated fibrin. This

  19. Endothelial dysfunction in morbid obesity.

    PubMed

    Mauricio, Maria Dolores; Aldasoro, Martin; Ortega, Joaquin; Vila, José María

    2013-01-01

    Morbid obesity is a chronic multifunctional disease characterized by an accumulation of fat. Epidemiological studies have shown that obesity is associated with cardiovascular and metabolic disorders. Endothelial dysfunction, as defined by an imbalance between relaxing and contractile endothelial factors, plays a central role in the pathogenesis of these cardiometabolic diseases. Diminished bioavailability of nitric oxide (NO) contributes to endothelial dysfunction and impairs endothelium- dependent vasodilatation. But this is not the only mechanism that drives to endothelial dysfunction. Obesity has been associated with a chronic inflammatory process, atherosclerosis, and oxidative stress. Moreover levels of asymmetrical dimethyl-L-arginine (ADMA), an endogenous inhibitor of endothelial nitric oxide synthase (eNOS), are elevated in obesity. On the other hand, increasing prostanoid-dependent vasoconstriction and decreasing vasodilator prostanoids also lead to endothelial dysfunction in obesity. Other mechanisms related to endothelin-1 (ET-1) or endothelium derived hyperpolarizing factor (EDHF) have been proposed. Bariatric surgery (BS) is a safe and effective means to achieve significant weight loss, but its use is limited only to patients with severe obesity including morbid obesity. BS also proved efficient in endothelial dysfunction reduction improving cardiovascular and metabolic comorbidities associated with morbid obesity such as diabetes, coronary artery disease, nonalcoholic fatty liver disease and cancer. This review will provide a brief overview of the mechanisms that link obesity with endothelial dysfunction, and how weight loss is a cornerstone treatment for cardiovascular comorbidities obesity-related. A better understanding of the mechanisms of obesity-induced endothelial dysfunction may help develop new therapeutic strategies to reduce cardiovascular morbidity and mortality.

  20. Glucose and angiotensin II-derived endothelial extracellular vesicles regulate endothelial dysfunction via ERK1/2 activation.

    PubMed

    Taguchi, Kumiko; Hida, Mari; Narimatsu, Haruka; Matsumoto, Takayuki; Kobayashi, Tsuneo

    2017-02-01

    In various diseases, including diabetes, extracellular vesicles (EVs) have been detected in circulation and tissues. EVs are small membrane vesicles released from various cell types under varying conditions. Recently, endothelial cell-derived EVs (EEVs) were identified as a marker of endothelial dysfunction in diabetes, but the ensuing mechanisms remain poorly understood. In this study, we dissected the ensuing pathways with respect to nitric oxide (NO) production under the condition of type 2 diabetes. Human umbilical vein endothelial cells (HUVECs) were stimulated with glucose alone and with glucose in combination with angiotensin II (Ang II) for 48 h. In supernatants from glucose + Ang II-stimulated HUVECs, release of EEVs was assessed using Western blotting with an anti-CD144 antibody. EEV release was significantly increased after stimulation of HUVECs, and high glucose + Ang II-derived EEVs impaired ACh-induced vascular relaxation responses and NO production in mice aortic rings. Furthermore, high glucose + Ang II-derived EEVs induced ERK1/2 signalling and decreased endothelial NO synthase (eNOS) protein expression in mice aortas. Furthermore, in the presence of the MEK/ERK1/2 inhibitor PD98059, high glucose plus Ang II treatment stimulated EEVs in HUVECs and those EEVs prevented the impairments of ACh-induced relaxation and NO production in mice aortas. These data strongly indicate that high glucose and Ang II directly affect endothelial cells and the production of EEVs; the resultant EEVs aggravate endothelial dysfunction by regulating eNOS protein levels and ERK1/2 signalling in mice aortas.

  1. Decellularized extracellular matrix of human umbilical vein endothelial cells promotes endothelial differentiation of stem cells from exfoliated deciduous teeth.

    PubMed

    Gong, Ting; Heng, Boon Chin; Xu, Jianguang; Zhu, Shaoyue; Yuan, Changyong; Lo, Edward Chin Man; Zhang, Chengfei

    2017-04-01

    Dental stem cells can serve as a potential source of functional endothelial cells for tissue engineering applications, but the endothelial-lineage differentiation efficiency is rather low even with growth factors and mechanical stimuli, which greatly limits their clinical applications. This is partly due to the deficiency of standard two-dimensional (2-D) culture systems, which is unable to recapitulate the three-dimensional (3-D) in vivo milieu that is rich in extracellular matrix. Hence, we extracted decellularized extracellular matrix from human umbilical vein endothelial cells (HUVECs-DECM) to provide a bioactive substratum conducive to the endothelial differentiation of dental stem cells. Compared to cells plated on tissue culture polystyrene (TCP), stem cells from exfoliated deciduous teeth (SHED) cultured on the HUVECs-DECM demonstrated more regular arrangement and elongated morphology. HUVECs-DECM significantly enhanced the rapid adhesion and proliferation rates of SHED, as demonstrated by WST-8 assay and immunocytochemistry indicating higher expression levels of vinculin by newly adherent SHED on HUVECs-DECM versus TCP. In addition, there was twofold to fivefold higher mRNA expression levels of endothelial-specific markers CD31 and VEGFR-2 in SHED after seven days of culture on DECM versus TCP. Functional testing with in vitro matrigel angiogenesis assay identified more capillary-like structure formation with significantly higher tubule length in SHED induced by DECM versus TCP. Hence, the results of this study provide a better understanding of the unique characteristics of cell-specific ECM and demonstrated the potential use of HUVECs-DECM as a culture substratum conducive for stimulating the endothelial differentiation of SHED for therapeutic angiogenic applications. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 1083-1093, 2017.

  2. A Bilayer Construct Controls Adipose-Derived Stem Cell Differentiation into Endothelial Cells and Pericytes without Growth Factor Stimulation

    DTIC Science & Technology

    2011-01-01

    cells and pericytes. Gene expression analysis revealed strong upregulation of endothelial markers, CD31, and von Willebrand factor, up to day 11 in...phenotype, but a subset of the ASC expressed pericyte markers. The NG2 gene was upregulated over 11 days with corresponding evidence for the cell...surface marker. Platelet- derived growth factor receptor beta gene expression decreased as the multipotent ASC differentiated up to day 7. Increased

  3. Human microvascular lymphatic and blood endothelial cells produce fibrillin: deposition patterns and quantitative analysis.

    PubMed

    Rossi, Antonella; Gabbrielli, Erica; Villano, Marilisa; Messina, Mario; Ferrara, Francesco; Weber, Elisabetta

    2010-12-01

    Fibrillin microfibrils constitute a scaffold for elastin deposition in the wall of arteries and form the anchoring filaments that connect the lymphatic endothelium to surrounding elastic fibers. We previously reported that fibrillin is deposited in a honeycomb pattern in bovine arterial endothelial cells, which also deposit microfibril-associated glycoprotein (MAGP)-1, whereas thoracic duct endothelial cells form an irregular web. The present immunohistochemical study was designed to verify whether lymphatic and blood human dermal microvascular endothelial cells (HDMECs) isolated from human foreskin by the sequential use of a pan-endothelial marker, CD31, and the lymphatic specific marker, D2-40, deposit fibrillin and MAGP-1. In both cell types, fibrillin and MAGP-1 co-localized and were deposited with different patterns of increasing complexity co-existing in the same culture. Fibrillin microfibrils formed a wide-mesh honeycomb leaving fibrillin-free spaces that were gradually filled. This modality of fibrillin deposition, similar to that of bovine large artery endothelial cells, was basically the same in blood and lymphatic HDMECs. In some lymphatic HDMECs, fibrillin was initially deposited as uniformly scattered short fibrillin strands probably as a result of anchoring filaments carried over from the vessels of origin. Our findings show that blood and lymphatic endothelial cells participate in fibrillin deposition in human skin.

  4. Examining Endothelial Function and Platelet Reactivity in Patients with Depression before and after SSRI Therapy

    PubMed Central

    Dawood, Tye; Barton, David A.; Lambert, Elisabeth A.; Eikelis, Nina; Lambert, Gavin W.

    2016-01-01

    Although it is recognized that patients with major depressive disorder (MDD) are at increased risk of developing cardiovascular disease (CVD) the mechanisms responsible remain unknown. Endothelial dysfunction is one of the first signs of CVD. Using two techniques, flow-mediated dilatation in response to reactive hyperemia and laser Doppler velocimetry with iontophoresis, we examined endothelial function in the forearm before and after serotonin-specific reuptake inhibitor (SSRI) treatment in 31 patients with MDD. Measurement of intercellular adhesion molecule-1, vascular cell adhesion molecule-1, soluble P-selectin, and noradrenaline in plasma was also performed. Prior to treatment, markers of endothelial and vascular function and platelet reactivity were within the normal range. Following SSRI therapy (95 ± 5 days) symptoms of depression were reduced (paired difference between pre- and post-treatment Hamilton rating −18 ± 1, P < 0.001) with 19 patients recovered and 4 remitted. There occurred no significant change in markers of endothelial or vascular function following SSRI therapy. The improvement in Hamilton depression rating in response to therapy could be independently predicted by the baseline arterial plasma noradrenaline concentration (r2 = 0.36, P = 0.003). In this cohort of patients with MDD, SSRI therapy did not influence endothelial function or markers of vascular or platelet reactivity. Patient response to SSRI therapy could be predicted by the initial circulating level of noradrenaline, with noradrenaline levels being lower in responders. PMID:26924994

  5. Selective capture of endothelial and perivascular cells from brain microvessels using laser capture microdissection.

    PubMed

    Kinnecom, Katie; Pachter, Joel S

    2005-12-01

    Laser capture microdissection (LCM) of the major cell types comprising brain microvessels offers a powerful technology to explore the molecular basis of the blood-brain barrier in health and disease. However, the ability to selectively retrieve endothelial or perivascular cells, without cross-contamination from the other, has proven difficult. Additionally, histochemical methods previously described for use with LCM have not allowed for identification of all the different size branches of the microvascular tree. Here, we describe a double immunostaining method, combining bright-field and fluorescence microscopy, and using an extensive dehydration with xylene, to clearly identify and spatially resolve endothelial from perivascular cells within all size microvascular branches in frozen brain sections. LCM of these sections, coupled with RNA analysis by reverse-transcription polymerase chain reaction, revealed that captured endothelial cells show endothelial markers but no detectable markers for astrocytes or smooth muscle cells/pericytes. Conversely, captured astrocytes or smooth muscle cells/pericytes demonstrate their respective markers, but not those of endothelial cells. This approach has applicability to microarray analysis, thereby enabling global gene profiling of the different cell types along the entirety of the brain microvascular tree.

  6. Isolated tumor endothelial cells maintain specific character during long-term culture

    SciTech Connect

    Matsuda, Kohei; Ohga, Noritaka; Hida, Yasuhiro; Muraki, Chikara; Tsuchiya, Kunihiko; Kurosu, Takuro; Akino, Tomoshige; Shih, Shou-Ching; and others

    2010-04-16

    Tumor angiogenesis is necessary for solid tumor progression and metastasis. Increasing evidence indicates that tumor endothelial cells (TECs) are more relevant to the study of tumor angiogenesis than normal endothelial cells (NECs) because their morphologies and gene expression are different from NECs. However, it is challenging to isolate and culture large numbers of pure ECs from tumor tissue since the percentage of ECs is only about 1-2% and tumor cells and fibroblasts easily overgrow them. In addition, there has been concern that isolated TECs may lose their special phenotype once they are dissociated from tumor cells. In this study, we have successfully purified murine TECs from four different human tumor xenografts and NECs from murine dermal tissue. Isolated ECs expressed endothelial markers, such as CD31, VE-cadherin (CD144), and endoglin (CD105), for more than 3 months after isolation. TECs maintained tumor endothelial-specific markers, such as tumor endothelial marker 8 (TEM8) and aminopeptidase N (APN), as in tumor blood vessels in vivo. In addition, TECs were more proliferative and motile than NECs. TECs showed a higher response to VEGF and higher expression of VEGF receptors-1 and -2 than NECs did. Stem cell antigen-1 was up-regulated in all four TECs, suggesting that they have a kind of stemness. Cultured TECs maintain distinct biological differences from NECs as in vivo. In conclusion, it was suggested that TECs are relevant material for tumor angiogenesis research.

  7. Endotoxin-induced endothelial fibrosis is dependent on expression of transforming growth factors β1 and β2.

    PubMed

    Echeverría, César; Montorfano, Ignacio; Tapia, Pablo; Riedel, Claudia; Cabello-Verrugio, Claudio; Simon, Felipe

    2014-09-01

    During endotoxemia-induced inflammatory disease, bacterial endotoxins circulate in the bloodstream and interact with endothelial cells (ECs), inducing dysfunction of the ECs. We previously reported that endotoxins induce the conversion of ECs into activated fibroblasts. Through endotoxin-induced endothelial fibrosis, ECs change their morphology and their protein expression pattern, thereby suppressing endothelial markers and upregulating fibrotic proteins. The most commonly used fibrotic inducers are transforming growth factor β1 (TGF-β1) and TGF-β2. However, whether TGF-β1 and TGF-β2 participate in endotoxin-induced endothelial fibrosis remains unknown. We have shown that the endotoxin-induced endothelial fibrosis process is dependent on the TGF-β receptor, ALK5, and the activation of Smad3, a protein that is activated by ALK5 activation, thus suggesting that endotoxin elicits TGF-β production to mediate endotoxin-induced endothelial fibrosis. Therefore, we investigated the dependence of endotoxin-induced endothelial fibrosis on the expression of TGF-β1 and TGF-β2. Endotoxin-treated ECs induced the expression and secretion of TGF-β1 and TGF-β2. TGF-β1 and TGF-β2 downregulation inhibited the endotoxin-induced changes in the endothelial marker VE-cadherin and in the fibrotic proteins α-SMA and fibronectin. Thus, endotoxin induces the production of TGF-β1 and TGF-β2 as a mechanism to promote endotoxin-induced endothelial fibrosis. To the best of our knowledge, this is the first report showing that endotoxin induces endothelial fibrosis via TGF-β secretion, which represents an emerging source of vascular dysfunction. These findings contribute to understanding the molecular mechanism of endotoxin-induced endothelial fibrosis, which could be useful in the treatment of inflammatory diseases.

  8. Solid tumor therapy by selectively targeting stromal endothelial cells

    PubMed Central

    Liu, Shihui; Liu, Jie; Ma, Qian; Cao, Liu; Fattah, Rasem J.; Yu, Zuxi; Bugge, Thomas H.; Finkel, Toren; Leppla, Stephen H.

    2016-01-01

    Engineered tumor-targeted anthrax lethal toxin proteins have been shown to strongly suppress growth of solid tumors in mice. These toxins work through the native toxin receptors tumor endothelium marker-8 and capillary morphogenesis protein-2 (CMG2), which, in other contexts, have been described as markers of tumor endothelium. We found that neither receptor is required for tumor growth. We further demonstrate that tumor cells, which are resistant to the toxin when grown in vitro, become highly sensitive when implanted in mice. Using a range of tissue-specific loss-of-function and gain-of-function genetic models, we determined that this in vivo toxin sensitivity requires CMG2 expression on host-derived tumor endothelial cells. Notably, engineered toxins were shown to suppress the proliferation of isolated tumor endothelial cells. Finally, we demonstrate that administering an immunosuppressive regimen allows animals to receive multiple toxin dosages and thereby produces a strong and durable antitumor effect. The ability to give repeated doses of toxins, coupled with the specific targeting of tumor endothelial cells, suggests that our strategy should be efficacious for a wide range of solid tumors. PMID:27357689

  9. [The impact of detecting endothelial dysfunction in atherosclerosis: Role of positron emission tomography (PET)].

    PubMed

    Alexánderson-Rosas, Erick; Calleja-Torres, Rodrigo; Martínez-García, Alfonso; Lamothe-Molina, Pedro Alberto; Ochoa-López, Juan Manuel; Meléndez, Gabriela; Kimura-Hayama, Eric; Meave-González, Aloha

    2010-01-01

    The endothelium plays an important role in the regulation of the intracellular fluid, vascular permeability, and modulation of vascular focal tone and angiogenesis. Endothelial dysfunction is manifested by the loss of the endothelium ability to modulate physiology changes in its vascular bed, and actually it is considered a prognostic marker of coronary artery disease. The relevance of assessing endothelial dysfunction relies in that it has been observed in different pathologies like DM, dyslipidemia, hypertension, tabaquism and in immunologic diseases like antiphospholipid syndrome and systemic lupus. PET is a non invasive method that allows the absolute quantification of myocardial blood flow during rest, stress and adrenergic stimulation, which allows to asses endothelial function. Therefore PET is a useful diagnostic technique to identify patients with endothelial dysfunction, and in the assessment of its response to administered therapy, allowing an optimal control and prevention of secondary adverse events of these diseases.

  10. The effect of moesin overexpression on ageing of human dermal microvascular endothelial cells.

    PubMed

    Lee, Ju Hee; Hong, In Ae; Oh, Sang Ho; Kwon, Yeon Sook; Cho, Soo Hyun; Lee, Kwang Hoon

    2009-11-01

    Senescence of microvascular endothelial cells is known to play an important role in the pathophysiology of vascular diseases related to ageing, but the accurate mechanism or related genes are not known. Moesin, a cytoskeletal protein and the most potent candidate as an ageing-related protein, showed obvious changes in expression when compared before and after ageing. In this study, a lentivirus was used to overexpress moesin in endothelial cells. The expression of cell cycle mediators such as p16, cyclin D1 and cdk4, which can be the markers of ageing, was compared by RNA and was shown to be suppressed in moesin overexpressed endothelial cells. In conclusion, it can be said that the expression of moesin delays senescence of human dermal microvascular endothelial cells and this fundamental discovery can be used as a basis for understanding the mechanism of ageing and age-related diseases.

  11. Endothelial function in youth: A biomarker modulated by adiposity-related insulin resistance

    Technology Transfer Automated Retrieval System (TEKTRAN)

    To investigate the physical and metabolic determinants of endothelial dysfunction, an early marker of subclinical atherosclerosis, in normal weight and overweight adolescents with and without type 2 diabetes mellitus. A cross-sectional study of 81 adolescents: 21 normal weight, 25 overweight with no...

  12. Expression of parathyroid-specific genes in vascular endothelial progenitors of normal and tumoral parathyroid glands.

    PubMed

    Corbetta, Sabrina; Belicchi, Marzia; Pisati, Federica; Meregalli, Mirella; Eller-Vainicher, Cristina; Vicentini, Leonardo; Beck-Peccoz, Paolo; Spada, Anna; Torrente, Yvan

    2009-09-01

    Parathyroid tissue is able to spontaneously induce angiogenesis, proliferate, and secrete parathyroid hormone when autotransplanted in patients undergoing total parathyroidectomy. Angiogenesis is also involved in parathyroid tumorigenesis. Here we investigated the anatomical and molecular relationship between endothelial and parathyroid cells within human parathyroid glands. Immunohistochemistry for CD34 antigen identified two subpopulations in normal and tumoral parathyroid glands: one constituted by cells lining small vessels that displayed endothelial antigens (factor VIII, isolectin, laminin, CD146) and the other constituted of single cells scattered throughout the parenchyma that did not express endothelial markers. These parathyroid-derived CD34(+) cells were negative for the hematopoietic and mesenchymal markers CD45, Thy-1/CD90, CD105, and CD117/c-kit; however, a subset of CD34(+) cells co-expressed the parathyroid specific genes glial cell missing B, parathyroid hormone, and calcium sensing receptor. When cultured, these cells released significant amount of parathyroid hormone. Parathyroid-derived CD34(+) cells, but not CD34(-) cells, proliferated slowly and differentiated into mature endothelial cells. CD34(+) cells from parathyroid tumors differed from those derived from normal parathyroid glands as: 1) they were more abundant and mainly scattered throughout the parenchyma; 2) they rarely co-expressed CD146; and 3) a fraction co-expressed nestin. In conclusion, we identified cells expressing endothelial and parathyroid markers in human adult parathyroid glands. These parathyroid/endothelial cells were more abundant and less committed in parathyroid tumors compared with normal glands, showing features of endothelial progenitors, which suggests that they might be involved in parathyroid tumorigenesis.

  13. Endothelial Differentiation of Adipose-Derived Stem Cells from Elderly Patients with Cardiovascular Disease

    PubMed Central

    Zhang, Ping; Moudgill, Neil; Hager, Eric; Tarola, Nicolas; DiMatteo, Christopher; McIlhenny, Stephen; Tulenko, Thomas

    2011-01-01

    Adipose-derived stem cells (ASCs) possess significant therapeutic potential for tissue engineering and regeneration. This study investigates the endothelial differentiation and functional capacity of ASCs isolated from elderly patients. Isolation of ASCs from 53 patients (50–89 years) revealed that advanced age or comorbidity did not negatively impact stem cell harvest; rather, higher numbers were observed in older donors (>70 years) than in younger. ASCs cultured in endothelial growth medium-2 for up to 3 weeks formed cords upon Matrigel and demonstrated acetylated-low-density lipoprotein and lectin uptake. Further stimulation with vascular endothelial growth factor and shear stress upregulated endothelial cell-specific markers (CD31, von Willebrand factor, endothelial nitric oxide synthase, and VE-cadherin). Inhibition of the PI3K but not mitogen-activated protein kinase pathway blocked the observed endothelial differentiation. Shear stress promoted an anti-thrombogenic phenotype as demonstrated by production of tissue-plasminogen activator and nitric oxide, and inhibition of plasminogen activator inhibitor-1. Shear stress augmented integrin α5β1 expression and subsequently increased attachment of differentiated ASCs to basement membrane components. Finally, ASCs seeded onto a decellularized vein graft resisted detachment despite application of shear force up to 9 dynes. These results suggest that (1) advanced age and comorbidity do not negatively impact isolation of ASCs, and (2) these stem cells retain significant capacity to acquire key endothelial cell traits throughout life. As such, adipose tissue is a practical source of autologous stem cells for vascular tissue engineering. PMID:20879833

  14. A Methodology for Concomitant Isolation of Intimal and Adventitial Endothelial Cells from the Human Thoracic Aorta

    PubMed Central

    Leclercq, Anne; Veillat, Véronique; Loriot, Sandrine; Spuul, Pirjo; Madonna, Francesco; Roques, Xavier; Génot, Elisabeth

    2015-01-01

    Background Aortic diseases are diverse and involve a multiplicity of biological systems in the vascular wall. Aortic dissection, which is usually preceded by aortic aneurysm, is a leading cause of morbidity and mortality in modern societies. Although the endothelium is now known to play an important role in vascular diseases, its contribution to aneurysmal aortic lesions remains largely unknown. The aim of this study was to define a reliable methodology for the isolation of aortic intimal and adventitial endothelial cells in order to throw light on issues relevant to endothelial cell biology in aneurysmal diseases. Methodology/Principal Findings We set up protocols to isolate endothelial cells from both the intima and the adventitia of human aneurysmal aortic vessel segments. Throughout the procedure, analysis of cell morphology and endothelial markers allowed us to select an endothelial fraction which after two rounds of expansion yielded a population of >90% pure endothelial cells. These cells have the features and functionalities of freshly isolated cells and can be used for biochemical studies. The technique was successfully used for aortic vessel segments of 20 patients and 3 healthy donors. Conclusions/Significance This simple and highly reproducible method allows the simultaneous preparation of reasonably pure primary cultures of intimal and adventitial human endothelial cells, thus providing a reliable source for investigating their biology and involvement in both thoracic aneurysms and other aortic diseases. PMID:26599408

  15. Endothelial dysfunction as a predictor of cardiovascular disease in type 1 diabetes

    PubMed Central

    Bertoluci, Marcello C; Cé, Gislaine V; da Silva, Antônio MV; Wainstein, Marco V; Boff, Winston; Puñales, Marcia

    2015-01-01

    Macro and microvascular disease are the main cause of morbi-mortality in type 1 diabetes (T1DM). Although there is a clear association between endothelial dysfunction and atherosclerosis in type 2 diabetes, a cause-effect relationship is less clear in T1DM. Although endothelial dysfunction (ED) precedes atherosclerosis, it is not clear weather, in recent onset T1DM, it may progress to clinical macrovascular disease. Moreover, endothelial dysfunction may either be reversed spontaneously or in response to intensive glycemic control, long-term exercise training and use of statins. Acute, long-term and post-prandial hyperglycemia as well as duration of diabetes and microalbuminuria are all conditions associated with ED in T1DM. The pathogenesis of endothelial dysfunction is closely related to oxidative-stress. NAD(P)H oxidase over activity induces excessive superoxide production inside the mitochondrial oxidative chain of endothelial cells, thus reducing nitric oxide bioavailability and resulting in peroxynitrite formation, a potent oxidant agent. Moreover, oxidative stress also uncouples endothelial nitric oxide synthase, which becomes dysfunctional, inducing formation of superoxide. Other important mechanisms are the activation of both the polyol and protein kinase C pathways as well as the presence of advanced glycation end-products. Future studies are needed to evaluate the potential clinical applicability of endothelial dysfunction as a marker for early vascular complications in T1DM. PMID:26069717

  16. Endothelial Cells from Bovine Adrenal Medulla Develop Capillary-Like Growth Patterns in Culture

    NASA Astrophysics Data System (ADS)

    Banerjee, Dipak K.; Ornberg, Richard L.; Youdim, Moussa B. H.; Heldman, Eli; Pollard, Harvey B.

    1985-07-01

    The endocrine barrier between chromaffin cells and the blood stream in the adrenal medulla is made of capillary endothelial cells. We have now succeeded in isolating endothelial cells from adrenal medullary tissue, which are probably derived from this barrier. These cells grow on plastic surfaces in the absence of special growth factors or collagen overlays and differentiate into organized structures quite similar to true capillaries. The cells contain factor VIII:R, a marker for endothelial cells, and form intercellular junctions characteristic of capillary endothelial cells. They also synthesize and secrete basal lamina structures and engage in transcytosis, a characteristic ultrastructural and functional combination of exocytosis and endocytosis across the thin endothelial cell processes. These endothelial cells can take up and deaminate catecholamines by A-type monoamine oxidase, an enzyme functionally distinct from the B-type monoamine oxidase found in chromaffin cells. These data indicate that the chromaffin cell and its endothelial cell neighbor may constitute the functional unit of catecholamine metabolism in the adrenal medulla.

  17. Endothelial-regenerating cells: an expanding universe.

    PubMed

    Steinmetz, Martin; Nickenig, Georg; Werner, Nikos

    2010-03-01

    Atherosclerosis is the most common cause for cardiovascular diseases and is based on endothelial dysfunction. A growing body of evidence suggests the contribution of bone marrow-derived endothelial progenitor cells, monocytic cells, and mature endothelial cells to vessel formation and endothelial rejuvenation. To this day, various subsets of these endothelial-regenerating cells have been identified according to cellular origin, phenotype, and properties in vivo and in vitro. However, the definition and biology, especially of endothelial progenitor cells, is complex and under heavy debate. In this review, we focus on current definitions of endothelial progenitor cells, highlight the clinical relevance of endothelial-regenerating cells, and provide new insights into cell-cell interactions involved in endothelial cell rejuvenation.

  18. WIPP marker development

    SciTech Connect

    1994-04-01

    This article discusses the development of permanent, passive markers for the Waste Isolation Pilot Plant (WIPP) and presents some preliminary concepts in drawings and a table of components for the markers. The panel, convened by Sandia National Laboratories, was charged with developing design characteristics for permanent markers and judging the efficacy of markers in deterring inadvertent human intrusion. 6 figs., 2 tabs.

  19. Proliferation status defines functional properties of endothelial cells.

    PubMed

    Lipps, Christoph; Badar, Muhammad; Butueva, Milada; Dubich, Tatyana; Singh, Vivek Vikram; Rau, Sophie; Weber, Axel; Kracht, Michael; Köster, Mario; May, Tobias; Schulz, Thomas F; Hauser, Hansjörg; Wirth, Dagmar

    2017-04-01

    Homeostasis of solid tissue is characterized by a low proliferative activity of differentiated cells while special conditions like tissue damage induce regeneration and proliferation. For some cell types it has been shown that various tissue-specific functions are missing in the proliferating state, raising the possibility that their proliferation is not compatible with a fully differentiated state. While endothelial cells are important players in regenerating tissue as well as in the vascularization of tumors, the impact of proliferation on their features remains elusive. To examine cell features in dependence of proliferation, we established human endothelial cell lines in which proliferation is tightly controlled by a doxycycline-dependent, synthetic regulatory unit. We observed that uptake of macromolecules and establishment of cell-cell contacts was more pronounced in the growth-arrested state. Tube-like structures were formed in vitro in both proliferating and non-proliferating conditions. However, functional vessel formation upon transplantation into immune-compromised mice was restricted to the proliferative state. Kaposi's sarcoma-associated herpes virus (KSHV) infection resulted in reduced expression of endothelial markers. Upon transplantation of infected cells, drastic differences were observed: proliferation arrested cells acquired a high migratory activity while the proliferating counterparts established a tumor-like phenotype, similar to Kaposi Sarcoma lesions. The study gives evidence that proliferation governs endothelial functions. This suggests that several endothelial functions are differentially expressed during angiogenesis. Moreover, since proliferation defines the functional properties of cells upon infection with KSHV, this process crucially affects the fate of virus-infected cells.

  20. Endothelial GATA-6 deficiency promotes pulmonary arterial hypertension.

    PubMed

    Ghatnekar, Angela; Chrobak, Izabela; Reese, Charlie; Stawski, Lukasz; Seta, Francesca; Wirrig, Elaine; Paez-Cortez, Jesus; Markiewicz, Margaret; Asano, Yoshihide; Harley, Russell; Silver, Richard; Feghali-Bostwick, Carol; Trojanowska, Maria

    2013-06-01

    Pulmonary arterial hypertension (PAH) is a chronic and progressive disease characterized by pulmonary vasculopathy with elevation of pulmonary artery pressure, often culminating in right ventricular failure. GATA-6, a member of the GATA family of zinc-finger transcription factors, is highly expressed in quiescent vasculature and is frequently lost during vascular injury. We hypothesized that endothelial GATA-6 may play a critical role in the molecular mechanisms underlying endothelial cell (EC) dysfunction in PAH. Here we report that GATA-6 is markedly reduced in pulmonary ECs lining both occluded and nonoccluded vessels in patients with idiopathic and systemic sclerosis-associated PAH. GATA-6 transcripts are also rapidly decreased in rodent PAH models. Endothelial GATA-6 is a direct transcriptional regulator of genes controlling vascular tone [endothelin-1, endothelin-1 receptor type A, and endothelial nitric oxide synthase (eNOS)], pro-inflammatory genes, CX3CL1 (fractalkine), 5-lipoxygenease-activating protein, and markers of vascular remodeling, including PAI-1 and RhoB. Mice with the genetic deletion of GATA-6 in ECs (Gata6-KO) spontaneously develop elevated pulmonary artery pressure and increased vessel muscularization, and these features are further exacerbated in response to hypoxia. Furthermore, innate immune cells including macrophages (CD11b(+)/F4/80(+)), granulocytes (Ly6G(+)/CD45(+)), and dendritic cells (CD11b(+)/CD11c(+)) are significantly increased in normoxic Gata6-KO mice. Together, our findings suggest a critical role of endothelial GATA-6 deficiency in development and disease progression in PAH.

  1. Identification of novel targets for antiangiogenic therapy by comparing the gene expressions of tumor and normal endothelial cells

    PubMed Central

    Otsubo, Tsuguteru; Hida, Yasuhiro; Ohga, Noritaka; Sato, Hideshi; Kai, Toshihiro; Matsuki, Yasushi; Takasu, Hideo; Akiyama, Kosuke; Maishi, Nako; Kawamoto, Taisuke; Shinohara, Nobuo; Nonomura, Katsuya; Hida, Kyoko

    2014-01-01

    Targeting tumor angiogenesis is an established strategy for cancer therapy. Because angiogenesis is not limited to pathological conditions such as cancer, molecular markers that can distinguish between physiological and pathological angiogenesis are required to develop more effective and safer approaches for cancer treatment. To identify such molecules, we determined the gene expression profiles of murine tumor endothelial cells (mTEC) and murine normal endothelial cells using DNA microarray analysis followed by quantitative reverse transcription–polymerase chain reaction analysis. We identified 131 genes that were differentially upregulated in mTEC. Functional analysis using siRNA-mediated gene silencing revealed five novel tumor endothelial cell markers that were involved in the proliferation or migration of mTEC. The expression of DEF6 and TMEM176B was upregulated in tumor vessels of human renal cell carcinoma specimens, suggesting that they are potential targets for antiangiogenic intervention for renal cell carcinoma. Comparative gene expression analysis revealed molecular differences between tumor endothelial cells and normal endothelial cells and identified novel tumor endothelial cell markers that may be exploited to target tumor angiogenesis for cancer treatment. PMID:24602018

  2. Obesity, inflammation and endothelial dysfunction.

    PubMed

    Iantorno, M; Campia, U; Di Daniele, N; Nistico, S; Forleo, G B; Cardillo, C; Tesauro, M

    2014-01-01

    Cardiovascular disease is the leading cause of morbidity and mortality in obese individuals. Obesity dramatically increases the risk of development of metabolic and cardiovascular disease. This risk appears to originate from disruption in adipose tissue function leading to a chronic inflammatory state and to dysregulation of the endocrine and paracrine actions of adipocyte-derived factors. These, in turn, impair vascular homeostasis and lead to endothelial dysfunction. An altered endothelial cell phenotype and endothelial dysfunction are common among all obesity-related complications. A crucial aspect of endothelial dysfunction is reduced nitric oxide (NO) bioavailability. A systemic pro-inflammatory state in combination with hyperglycemia, insulin resistance, oxidative stress and activation of the renin angiotensin system are systemic disturbances in obese individuals that contribute independently and synergistically to decreasing NO bioavailability. On the other hand, pro-inflammatory cytokines are locally produced by perivascular fat and act through a paracrine mechanism to independently contribute to endothelial dysfunction and smooth muscle cell dysfunction and to the pathogenesis of vascular disease in obese individuals. The promising discovery that obesity-induced vascular dysfunction is, at least in part, reversible, with weight loss strategies and drugs that promote vascular health, has not been sufficiently proved to prevent the cardiovascular complication of obesity on a large scale. In this review we discuss the pathophysiological mechanisms underlying inflammation and vascular damage in obese patients.

  3. Coculture with endothelial cells reduces the population of cycling LeX neural precursors but increases that of quiescent cells with a side population phenotype

    SciTech Connect

    Mathieu, Celine . E-mail: marc-andre.mouthon@cea.fr

    2006-04-01

    Neural stem cell proliferation and differentiation are regulated by external cues from their microenvironment. As endothelial cells are closely associated with neural stem cell in brain germinal zones, we investigated whether endothelial cells may interfere with neurogenesis. Neural precursor cells (NPC) from telencephalon of EGFP mouse embryos were cocultured in direct contact with endothelial cells. Endothelial cells did not modify the overall proliferation and apoptosis of neural cells, albeit they transiently delayed spontaneous apoptosis. These effects appeared to be specific to endothelial cells since a decrease in proliferation and a raise in apoptosis were observed in cocultures with fibroblasts. Endothelial cells stimulated the differentiation of NPC into astrocytes and into neurons, whereas they reduced differentiation into oligodendrocytes in comparison to adherent cultures on polyornithine. Determination of NPC clonogenicity and quantification of LeX expression, a marker for NPC, showed that endothelial cells decreased the number of cycling NPC. On the other hand, the presence of endothelial cells increased the number of neural cells having 'side population' phenotype, another marker reported on NPC, which we have shown to contain quiescent cells. Thus, we show that endothelial cells may regulate neurogenesis by acting at different level of NPC differentiation, proliferation and quiescence.

  4. VEGF treatment promotes bone marrow-derived CXCR4+ mesenchymal stromal stem cell differentiation into vessel endothelial cells

    PubMed Central

    Li, Qiming; Xia, Shudong; Fang, Hanyun; Pan, Jiansheng; Jia, Yinfeng; Deng, Gang

    2017-01-01

    Stem/progenitor cells serve an important role in the process of blood vessel repair. However, the mechanism of vascular repair mediated by C-X-C chemokine receptor type 4-positive (CXCR4+) bone marrow-derived mesenchymal stem cells (BMSCs) following myocardial infarction remains unclear. The aim of the present study was to investigate the effects of vascular endothelial growth factor (VEGF) on vessel endothelial differentiation from BMSCs. CXCR4+ BMSCs were isolated from the femoral bone marrow of 2-month-old mice and the cells were treated with VEGF. Expression of endothelial cell markers and the functional properties were assessed by reverse transcription-quantitative polymerase chain reaction, flow cytometry and vascular formation analyses. The results indicated that the CXCR4+ BMSCs from femoral bone marrow cells expressed putative cell surface markers of mesenchymal stem cells. Treatment with VEGF induced platelet/endothelial cell adhesion molecule-1 (PECAM-1) and von Willebrand factor (vWF) expression at the transcriptional and translational levels, compared with untreated controls. Moreover, VEGF treatment induced CXCR4+ BMSCs to form hollow tube-like structures on Matrigel, suggesting that the differentiated endothelial cells had the functional properties of blood vessels. The results demonstrate that the CXCR4+ BMSCs were able to differentiate into vessel endothelial cells following VEGF treatment. For cell transplantation in vascular disease, it may be concluded that CXCR4+ BMSCs are a novel source of endothelial progenitor cells with high potential for application in vascular repair. PMID:28352314

  5. Instruction of Circulating Endothelial Progenitors In Vitro towards Specialized Blood-Brain Barrier and Arterial Phenotypes

    PubMed Central

    Ponio, Julie Boyer-Di; El-Ayoubi, Fida; Glacial, Fabienne; Ganeshamoorthy, Kayathiri; Driancourt, Catherine; Godet, Maeva; Perrière, Nicolas; Guillevic, Oriane; Couraud, Pierre Olivier; Uzan, Georges

    2014-01-01

    Objective The vascular system is adapted to specific functions in different tissues and organs. Vascular endothelial cells are important elements of this adaptation, leading to the concept of ‘specialized endothelial cells’. The phenotype of these cells is highly dependent on their specific microenvironment and when isolated and cultured, they lose their specific features after few passages, making models using such cells poorly predictive and irreproducible. We propose a new source of specialized endothelial cells based on cord blood circulating endothelial progenitors (EPCs). As prototype examples, we evaluated the capacity of EPCs to acquire properties characteristic of cerebral microvascular endothelial cells (blood-brain barrier (BBB)) or of arterial endothelial cells, in specific inducing culture conditions. Approach and Results First, we demonstrated that EPC-derived endothelial cells (EPDCs) co-cultured with astrocytes acquired several BBB phenotypic characteristics, such as restricted paracellular diffusion of hydrophilic solutes and the expression of tight junction proteins. Second, we observed that culture of the same EPDCs in a high concentration of VEGF resulted, through activation of Notch signaling, in an increase of expression of most arterial endothelial markers. Conclusions We have thus demonstrated that in vitro culture of early passage human cord blood EPDCs under specific conditions can induce phenotypic changes towards BBB or arterial phenotypes, indicating that these EPDCs maintain enough plasticity to acquire characteristics of a variety of specialized phenotypes. We propose that this property of EPDCs might be exploited for producing specialized endothelial cells in culture to be used for drug testing and predictive in vitro assays. PMID:24392113

  6. Acute effects of hyperinsulinemia and hyperglycemia on vascular inflammatory biomarkers and endothelial function in overweight and obese humans

    PubMed Central

    Perkins, Jennifer M.; Joy, Nino G.; Tate, Donna B.

    2015-01-01

    We investigated the separate and combined effects of hyperglycemia and hyperinsulinemia on markers of endothelial function, proinflammatory and proatherothrombotic responses in overweight/obese nondiabetic humans. Twenty-two individuals (13 F/9 M, BMI 30.1 ± 4.1 kg/m2) were studied during four randomized, single-blind protocols. The pancreatic clamp technique was combined with 4-h glucose clamps consisting of either 1) euinsulinemia-euglycemia, 2) euinsulinemia-hyperglycemia, 3) hyperinsulinemia-hyperglycemia, or 4) hyperinsulinemia-euglycemia. Insulin levels were higher (998 ± 66 vs. 194 ± 22 pmol/l) during hyperinsulinemia compared with euinsulinemia. Glucose levels were 11.1 mmol/l during hyperinsulinemia compared with 5.1 ± 0.1 mmol/l during euglycemia. VCAM, ICAM, P-selectin, E-selectin, IL-6, adiponectin, and PAI-1 responses were all increased (P < 0.01-0.0001), and endothelial function was decreased (P < 0.0005) during euinsulinemia-hyperglycemia compared with other protocols. Hyperinsulinemia in the presence of hyperglycemia prevented the increase in proinflammatory and proatherothrombotic markers while also normalizing vascular endothelial function. We conclude that 4 h of moderate hyperglycemia can result in increases of proinflammatory markers (ICAM, VCAM, IL-6, E-selectin), platelet activation (P-selectin), reduced fibrinolytic balance (increased PAI-1), and disordered endothelial function in a group of obese and overweight individuals. Hyperinsulinemia prevents the actions of moderate hyperglycemia to reduce endothelial function and increase proinflammatory and proatherothrombotic markers. PMID:26015434

  7. Genetic Regulation of Endothelial Vasomotor Function

    PubMed Central

    Kim, Seung Kyum; Massett, Michael P.

    2016-01-01

    The endothelium plays an important role in the regulation of vasomotor tone and the maintenance of vascular integrity. Endothelial dysfunction, i.e., impaired endothelial dependent dilation, is a fundamental component of the pathogenesis of cardiovascular disease. Although endothelial dysfunction is associated with a number of cardiovascular disease risk factors, those risk factors are not the only determinants of endothelial dysfunction. Despite knowing many molecules involved in endothelial signaling pathways, the genetic contribution to endothelial function has yet to be fully elucidated. This mini-review summarizes current evidence supporting the genetic contribution to endothelial vasomotor function. Findings from population-based studies, association studies for candidate genes, and unbiased large genomic scale studies in humans and rodent models are discussed. A brief synopsis of the current studies addressing the genetic regulation of endothelial responses to exercise training is also included. PMID:27932996

  8. The potential lymphangiogenic effects of hepatocyte growth factor/scatter factor in vitro and in vivo.

    PubMed

    Jiang, Wen G; Davies, Gaynor; Martin, Tracey A; Parr, Christian; Watkins, Gareth; Mansel, Robert E; Mason, Malcolm D

    2005-10-01

    Lymphangiogenesis is key to the lymphatic spread of cancer cells. The current study examined the potential effect of hepatocyte growth factor (HGF), a factor known to have strong biological effects on endothelial cells, on the lymphangiogenic function of endothelial cells and the formation of lymphatic vessels using both in vitro and in vivo models. Human endothelial cells that have lymphatic characteristics, human prostate and breast cancer cells PC-3 and MDA MB 231, were used. Expression of lymphatic markers, podoplanin, Prox-1, vascular endothelial growth factor receptor 3 (VEGF-R3) and LYVE-1 was determined using reverse transcription polymerase reaction and quantitative PCR. In nude mice prostate and breast xenograft tumour models, either HGF or an HGF-producing fibroblast cell line MRC-5 was given with or without the HGF antagonist, NK4. The lymphangiogenic marker and lymphatic vessels in tumour tissues were also assessed using quantitative PCR and immunohistochemistry, respectively. In the mice tumour models, infusion of rhHGF significantly increased the levels of podoplanin and LYVE-1 in the tumour (p=0.05 for podoplanin and p<0.05 for LYVE-1 vs. without HGF in the prostate tumour model, p<0.05 for podoplanin and p<0.01 for LYVE-1 vs. without HGF for the breast tumour model; p<0.05 for podoplanin and p<0.01 for LYVE-1 vs. without HGF in the breast tumour model). The increased level of LYVE-1 transcript was supported by an increase in the number of LYVE-1-positive lymphatic vessels in tumours, using immunohistochemical analysis. Co-injection of MRC5 cells also increased the levels of LYVE-1 and number of LYVE-1-positive vessels in tumour tissues. The effects of HGF and MRC5 were significantly reduced by the HGF antagonist, NK4. In the in vitro models, rhHGF significantly increased the level of both podoplanin and LYVE-1, as shown by quantitative PCR analysis. Hepatocyte growth factor has potential lymphangiogenic activities, and this may have important

  9. Efavirenz Causes Oxidative Stress, Endoplasmic Reticulum Stress, and Autophagy in Endothelial Cells.

    PubMed

    Weiß, Marlene; Kost, Bernd; Renner-Müller, Ingrid; Wolf, Eckhard; Mylonas, Ioannis; Brüning, Ansgar

    2016-01-01

    The non-nucleoside reverse transcriptase inhibitor efavirenz is a widely prescribed antiretroviral drug used in combined antiretroviral therapy. Despite being an essential and life-saving medication, the required lifelong use of HIV drugs has been associated with a variety of adverse effects, including disturbances in lipid metabolism and increased cardiovascular risk. Efavirenz belongs to those HIV drugs for which cardiovascular and endothelial dysfunctions have been reported. It is here shown that elevated concentrations of efavirenz can inhibit endothelial meshwork formation on extracellular matrix gels by normal and immortalized human umbilical vein cells. This inhibition was associated with an increase in oxidative stress markers, endoplasmic reticulum (ER) stress markers, and autophagy. Induction of ER stress occurred at pharmacologically relevant concentrations of efavirenz and resulted in reduced proliferation and cell viability of endothelial cells, which worsened in the presence of elevated efavirenz concentrations. In combination with the HIV protease inhibitor nelfinavir, both oxidative stress and ER stress became elevated in endothelial cells. These data indicate that pharmacologically relevant concentrations of efavirenz can impair cell viability of endothelial cells and that these effects may be aggravated by either elevated concentrations of efavirenz or by a combined use of efavirenz with other oxidative stress-inducing medications.

  10. Endothelial keratoplasty versus penetrating keratoplasty for Fuchs endothelial dystrophy

    PubMed Central

    Nanavaty, Mayank A; Wang, Xue; Shortt, Alex J

    2014-01-01

    Background Fuchs endothelial dystrophy (FED) is a condition in which there is premature degeneration of corneal endothelial cells. When the number of endothelial cells is reduced to a significant degree, fluid begins to accumulate within the cornea. As a result, the cornea loses its transparency and the individual suffers a reduction in vision. The only successful surgical treatment for this condition is replacement of part or all of the cornea with healthy tissue from a donor. The established procedure, penetrating keratoplasty (PKP), has been used for many years and its safety and efficacy are well known. Endothelial keratoplasty (EK) techniques are relatively new surgical procedures and their safety and efficacy relative to PKP are uncertain. Objectives The objective of this review was to compare the benefits and complications related to two surgical methods (EK and PKP) of replacing the diseased endothelial layer of the cornea with a healthy layer in people with FED. Search methods We searched CENTRAL (which contains the Cochrane Eyes and Vision Group Trials Register) (The Cochrane Library 2014, Issue 1), MEDLINE (January 1950 to January 2014), EMBASE (January 1980 to January 2014), Latin American and Caribbean Health Sciences Literature Database (LILACS) (January 1982 to January 2014), the metaRegister of Controlled Trials (mRCT) (www.controlled-trials.com) and ClinicalTrials.gov (www.clinicaltrials.gov). There were no date or language restrictions in the electronic searches for trials. The electronic databases were last searched on 27 January 2014. Selection criteria We included all randomised controlled trials (RCTs) comparing EK versus PKP for people (of any age and gender) who had been clinically diagnosed with FED. Data collection and analysis Two authors independently screened the search results, assessed trial quality and extracted data using the standard methodological procedures expected by The Cochrane Collaboration. Main results We included three

  11. Ceramic subsurface marker prototypes

    SciTech Connect

    Lukens, C.E.

    1985-05-02

    The client submitted 5 sets of porcelain and stoneware subsurface (radioactive site) marker prototypes (31 markers each set). The following were determined: compressive strength, thermal shock resistance, thermal crazing resistance, alkali resistance, color retention, and chemical resistance.

  12. TNF-α stimulates endothelial palmitic acid transcytosis and promotes insulin resistance

    PubMed Central

    Li, Wenjing; Yang, Xiaoyan; Zheng, Tao; Xing, Shasha; Wu, Yaogong; Bian, Fang; Wu, Guangjie; Li, Ye; Li, Juyi; Bai, Xiangli; Wu, Dan; Jia, Xiong; Wang, Ling; Zhu, Lin; Jin, Si

    2017-01-01

    Persistent elevation of plasma TNF-α is a marker of low grade systemic inflammation. Palmitic acid (PA) is the most abundant type of saturated fatty acid in human body. PA is bound with albumin in plasma and could not pass through endothelial barrier freely. Albumin-bound PA has to be transported across monolayer endothelial cells through intracellular transcytosis, but not intercellular diffusion. In the present study, we discovered that TNF-α might stimulate PA transcytosis across cardiac microvascular endothelial cells, which further impaired the insulin-stimulated glucose uptake by cardiomyocytes and promoted insulin resistance. In this process, TNF-α-stimulated endothelial autophagy and NF-κB signaling crosstalk with each other and orchestrate the whole event, ultimately result in increased expression of fatty acid transporter protein 4 (FATP4) in endothelial cells and mediate the increased PA transcytosis across microvascular endothelial cells. Hopefully the present study discovered a novel missing link between low grade systemic inflammation and insulin resistance. PMID:28304381

  13. Therapeutically targeting mitochondrial redox signalling alleviates endothelial dysfunction in preeclampsia

    PubMed Central

    McCarthy, Cathal; Kenny, Louise C.

    2016-01-01

    Aberrant placentation generating placental oxidative stress is proposed to play a critical role in the pathophysiology of preeclampsia. Unfortunately, therapeutic trials of antioxidants have been uniformly disappointing. There is provisional evidence implicating mitochondrial dysfunction as a source of oxidative stress in preeclampsia. Here we provide evidence that mitochondrial reactive oxygen species mediates endothelial dysfunction and establish that directly targeting mitochondrial scavenging may provide a protective role. Human umbilical vein endothelial cells exposed to 3% plasma from women with pregnancies complicated by preeclampsia resulted in a significant decrease in mitochondrial function with a subsequent significant increase in mitochondrial superoxide generation compared to cells exposed to plasma from women with uncomplicated pregnancies. Real-time PCR analysis showed increased expression of inflammatory markers TNF-α, TLR-9 and ICAM-1 respectively in endothelial cells treated with preeclampsia plasma. MitoTempo is a mitochondrial-targeted antioxidant, pre-treatment of cells with MitoTempo protected against hydrogen peroxide-induced cell death. Furthermore MitoTempo significantly reduced mitochondrial superoxide production in cells exposed to preeclampsia plasma by normalising mitochondrial metabolism. MitoTempo significantly altered the inflammatory profile of plasma treated cells. These novel data support a functional role for mitochondrial redox signaling in modulating the pathogenesis of preeclampsia and identifies mitochondrial-targeted antioxidants as potential therapeutic candidates. PMID:27604418

  14. Sitting and endothelial dysfunction: The role of shear stress

    PubMed Central

    Thosar, Saurabh S.; Johnson, Blair D.; Johnston, Jeanne D.; Wallace, Janet P.

    2012-01-01

    Summary Sedentary activity is a modifiable life-style behavior and a key component in the etiology of atherosclerotic cardiovascular disease (ACVD). US adults and children spend more than half their waking time in sedentary pursuits. Sedentary activity has been shown to result in impaired insulin sensitivity, impaired metabolic function and attenuated endothelial function, which are classic markers of ACVD. Sedentary activity is defined as ‘sitting without otherwise being active.’ This behavior promotes reduced muscular activity of the lower extremities which decreases leg blood flow, increases blood pooling in the calf, augments mean arterial pressure, and deforms arterial segments resulting in low mean shear stress (SS). SS activates distinct physiological mechanisms which have been proposed to be protective against ACVD; specifically through a SS-induced endothelium-derived nitric oxide mechanism. Reduced bioavailability of nitric oxide creates a pro-oxidant milieu resulting in increased oxidative stress. There is sufficient evidence which demonstrates that endothelial function is attenuated in the presence of oxidative stress. Sedentary activity results in low SS in the lower extremities which may result in increased oxidative stress and impaired endothelial function. This review furthers the use of sitting as model to study the effects of inactivity, discusses possible physiological mechanisms and suggests future directions. PMID:23197245

  15. Microenvironmental Regulation of the Sinusoidal Endothelial Cell Phenotype In Vitro

    PubMed Central

    March, Sandra; Hui, Elliot E.; Underhill, Gregory H.; Khetani, Salman; Bhatia, Sangeeta N.

    2010-01-01

    Liver Sinusoidal Endothelial Cells (LSEC) differ, both structurally and functionally, from endothelial cells (EC) lining blood vessels of other tissues. For example, in contrast to other EC, LSEC posses fenestrations, have low detectable levels of PECAM-1 expression, and in rat tissue, they distinctively express a cell surface marker recognized by the SE-1 antibody. These unique phenotypic characteristics seen in hepatic tissue are lost over time upon culture in vitro; therefore, this study sought to systematically examine the effects of microenvironmental stimuli, namely, extracellular matrix (ECM) and neighboring cells, on the LSEC phenotype in vitro. In probing the role of the underlying extracellular matrix, we identified collagen I and collagen III as well as mixtures of collagen I/collagen IV/fibronectin as having a positive effect on LSEC survival. Furthermore, using a stable hepatocellular model (hepatocyte-fibroblast) we were able to prolong the expression of both SE-1 and phenotypic functions of LSEC such as Factor VIII activity in co-cultured LSECs through the production of short-range paracrine signals. In the course of these experiments, we identified the antigen recognized by SE-1 as CD32b. Collectively, this study has identified several microenvironmental regulators of liver sinusoidal endothelial cells that prolong their phenotypic functions for up to 2 weeks in culture, enabling the development of better in vitro models of liver physiology and disease. PMID:19585615

  16. Activation of the TGFβ pathway impairs endothelial to haematopoietic transition.

    PubMed

    Vargel, Özge; Zhang, Yang; Kosim, Kinga; Ganter, Kerstin; Foehr, Sophia; Mardenborough, Yannicka; Shvartsman, Maya; Enright, Anton J; Krijgsveld, Jeroen; Lancrin, Christophe

    2016-02-19

    The endothelial to haematopoietic transition (EHT) is a key developmental process where a drastic change of endothelial cell morphology leads to the formation of blood stem and progenitor cells during embryogenesis. As TGFβ signalling triggers a similar event during embryonic development called epithelial to mesenchymal transition (EMT), we hypothesised that TGFβ activity could play a similar role in EHT as well. We used the mouse embryonic stem cell differentiation system for in vitro recapitulation of EHT and performed gain and loss of function analyses of the TGFβ pathway. Quantitative proteomics analysis showed that TGFβ treatment during EHT increased the secretion of several proteins linked to the vascular lineage. Live cell imaging showed that TGFβ blocked the formation of round blood cells. Using gene expression profiling we demonstrated that the TGFβ signalling activation decreased haematopoietic genes expression and increased the transcription of endothelial and extracellular matrix genes as well as EMT markers. Finally we found that the expression of the transcription factor Sox17 was up-regulated upon TGFβ signalling activation and showed that its overexpression was enough to block blood cell formation. In conclusion we showed that triggering the TGFβ pathway does not enhance EHT as we hypothesised but instead impairs it.

  17. Endothelial Plasticity: Shifting Phenotypes through Force Feedback

    PubMed Central

    Krenning, Guido; Barauna, Valerio G.; Krieger, José E.; Harmsen, Martin C.; Moonen, Jan-Renier A. J.

    2016-01-01

    The endothelial lining of the vasculature is exposed to a large variety of biochemical and hemodynamic stimuli with different gradients throughout the vascular network. Adequate adaptation requires endothelial cells to be highly plastic, which is reflected by the remarkable heterogeneity of endothelial cells in tissues and organs. Hemodynamic forces such as fluid shear stress and cyclic strain are strong modulators of the endothelial phenotype and function. Although endothelial plasticity is essential during development and adult physiology, proatherogenic stimuli can induce adverse plasticity which contributes to disease. Endothelial-to-mesenchymal transition (EndMT), the hallmark of endothelial plasticity, was long thought to be restricted to embryonic development but has emerged as a pathologic process in a plethora of diseases. In this perspective we argue how shear stress and cyclic strain can modulate EndMT and discuss how this is reflected in atherosclerosis and pulmonary arterial hypertension. PMID:26904133

  18. Cellular glutathione peroxidase deficiency and endothelial dysfunction.

    PubMed

    Forgione, Marc A; Weiss, Norbert; Heydrick, Stanley; Cap, André; Klings, Elizabeth S; Bierl, Charlene; Eberhardt, Robert T; Farber, Harrison W; Loscalzo, Joseph

    2002-04-01

    Cellular glutathione peroxidase (GPx-1) is the most abundant intracellular isoform of the GPx antioxidant enzyme family. In this study, we hypothesized that GPx-1 deficiency directly induces an increase in vascular oxidant stress, with resulting endothelial dysfunction. We studied vascular function in a murine model of homozygous deficiency of GPx-1 (GPx-1(-/-)). Mesenteric arterioles of GPx-1(-/-) mice demonstrated paradoxical vasoconstriction to beta-methacholine and bradykinin, whereas wild-type (WT) mice showed dose-dependent vasodilation in response to both agonists. One week of treatment of GPx-1(-/-) mice with L-2-oxothiazolidine-4-carboxylic acid (OTC), which increases intracellular thiol pools, resulted in restoration of normal vascular reactivity in the mesenteric bed of GPx-1(-/-) mice. We observed an increase of the isoprostane iPF(2alpha)-III, a marker of oxidant stress, in the plasma and aortas of GPx-1(-/-) mice compared with WT mice, which returned toward normal after OTC treatment. Aortic sections from GPx-1(-/-) mice showed increased binding of an anti-3-nitrotyrosine antibody in the absence of frank vascular lesions. These findings demonstrate that homozygous deficiency of GPx-1 leads to impaired endothelium-dependent vasodilator function presumably due to a decrease in bioavailable nitric oxide and to increased vascular oxidant stress. These vascular abnormalities can be attenuated by increasing bioavailable intracellular thiol pools.

  19. Impact of diabetic serum on endothelial cells: An in-vitro-analysis of endothelial dysfunction in diabetes mellitus type 2

    SciTech Connect

    Muenzel, Daniela; Lehle, Karla Haubner, Frank; Schmid, Christof; Birnbaum, Dietrich E.; Preuner, Juergen G.

    2007-10-19

    Diabetic endothelial dysfunction was characterized by altered levels of adhesion molecules and cytokines. Aim of our study was to evaluate the effects of diabetic serum on cell-growth and proinflammatory markers in human saphenous vein endothelial cells (HSVEC) from diabetic and non-diabetic patients. Diabetic serum showed (1) complementary proliferative activity for non-diabetic and diabetic HSVEC, (2) unchanged surface expression of adhesion molecules, and (3) elevated levels of sICAM-1 in HSVEC of all donors. The concentration of sVCAM-1 was increased only in diabetic cells. The proinflammatory state of diabetic HSVEC characterized by increased levels of cytokines was compensated. We concluded that even under normoglycemic conditions the serum itself contains critical factors leading to abnormal regulation of inflammation in diabetics. We introduced an in vitro model of diabetes representing the endothelial situation at the beginning of diabetes (non-diabetic cells/diabetic serum) as well as the diabetic chronic state (diabetic cells/diabetic serum)

  20. Glyoxalase 1-knockdown in human aortic endothelial cells – effect on the proteome and endothelial function estimates

    PubMed Central

    Stratmann, Bernd; Engelbrecht, Britta; Espelage, Britta C.; Klusmeier, Nadine; Tiemann, Janina; Gawlowski, Thomas; Mattern, Yvonne; Eisenacher, Martin; Meyer, Helmut E.; Rabbani, Naila; Thornalley, Paul J.; Tschoepe, Diethelm; Poschmann, Gereon; Stühler, Kai

    2016-01-01

    Methylglyoxal (MG), an arginine-directed glycating agent, is implicated in diabetic late complications. MG is detoxified by glyoxalase 1 (GLO1) of the cytosolic glyoxalase system. The aim was to investigate the effects of MG accumulation by GLO1-knockdown under hyperglycaemic conditions in human aortic endothelial cells (HAECs) hypothesizing that the accumulation of MG accounts for the deleterious effects on vascular function. SiRNA-mediated knockdown of GLO1 was performed and MG concentrations were determined. The impact of MG on the cell proteome and targets of MG glycation was analysed, and confirmed by Western blotting. Markers of endothelial function and apoptosis were assessed. Collagen content was assayed in cell culture supernatant. GLO1-knockdown increased MG concentration in cells and culture medium. This was associated with a differential abundance of cytoskeleton stabilisation proteins, intermediate filaments and proteins involved in posttranslational modification of collagen. An increase in fibrillar collagens 1 and 5 was detected. The extracellular concentration of endothelin-1 was increased following GLO1-knockdown, whereas the phosphorylation and amount of eNOS was not influenced by GLO1-knockdown. The expression of ICAM-1, VCAM-1 and of MCP-1 was elevated and apoptosis was increased. MG accumulation by GLO1-knockdown provoked collagen expression, endothelial inflammation and dysfunction and apoptosis which might contribute to vascular damage. PMID:27898103

  1. Obesity indices and metabolic markers are related to hs-CRP and adiponectin levels in overweight and obese females.

    PubMed

    Sanip, Zulkefli; Ariffin, Farah Diana; Al-Tahami, Belqes Abdullah Mohammed; Sulaiman, Wan Azman Wan; Rasool, Aida Hanum Ghulam

    2013-01-01

    Obese subjects had increased serum high sensitivity C-reactive protein (hs-CRP), decreased adiponectin levels, and impaired microvascular endothelial function compared to lean subjects. We investigated the relationships of serum hs-CRP, adiponectin and microvascular endothelial function with obesity indices and metabolic markers in overweight and obese female subjects. Anthropometric profile, body fat composition, biochemical analysis, serum hs-CRP and adiponectin levels, and microvascular endothelial function were measured in 91 female subjects. Microvascular endothelial function was determined using laser Doppler fluximetry and the process of iontophoresis. Mean age and body mass index (BMI) of subjects were 34.88 (7.87) years and 32.93 (4.82) kg/m(2). hs-CRP levels were positively correlated with weight, BMI, waist circumference, hip circumference, body fat and visceral fat. Adiponectin levels were positively correlated with insulin sensitivity index (HOMA-%S), and inversely correlated with waist hip ratio, triglyceride, fasting insulin and insulin resistance index (HOMA-IR). No relationship was seen between microvascular endothelial function and obesity indices, and metabolic markers. In overweight and obese female subjects, hs-CRP levels were correlated with obesity indices while adiponectin levels were inversely correlated with obesity indices and metabolic markers. No significant relationship was seen between microvascular endothelial function with obesity indices and metabolic markers including hs-CRP and adiponectin in female overweight and obese subjects.

  2. Endothelial dysfunction: a comprehensive appraisal

    PubMed Central

    Esper, Ricardo J; Nordaby, Roberto A; Vilariño, Jorge O; Paragano, Antonio; Cacharrón, José L; Machado, Rogelio A

    2006-01-01

    The endothelium is a thin monocelular layer that covers all the inner surface of the blood vessels, separating the circulating blood from the tissues. It is not an inactive organ, quite the opposite. It works as a receptor-efector organ and responds to each physical or chemical stimulus with the release of the correct substance with which it may maintain vasomotor balance and vascular-tissue homeostasis. It has the property of producing, independently, both agonistic and antagonistic substances that help to keep homeostasis and its function is not only autocrine, but also paracrine and endocrine. In this way it modulates the vascular smooth muscle cells producing relaxation or contraction, and therefore vasodilatation or vasoconstriction. The endothelium regulating homeostasis by controlling the production of prothrombotic and antithrombotic components, and fibrynolitics and antifibrynolitics. Also intervenes in cell proliferation and migration, in leukocyte adhesion and activation and in immunological and inflammatory processes. Cardiovascular risk factors cause oxidative stress that alters the endothelial cells capacity and leads to the so called endothelial "dysfunction" reducing its capacity to maintain homeostasis and leads to the development of pathological inflammatory processes and vascular disease. There are different techniques to evaluate the endothelium functional capacity, that depend on the amount of NO produced and the vasodilatation effect. The percentage of vasodilatation with respect to the basal value represents the endothelial functional capacity. Taking into account that shear stress is one of the most important stimulants for the synthesis and release of NO, the non-invasive technique most often used is the transient flow-modulate "endothelium-dependent" post-ischemic vasodilatation, performed on conductance arteries such as the brachial, radial or femoral arteries. This vasodilatation is compared with the vasodilatation produced by drugs that

  3. A coculture system of cavernous endothelial and smooth muscle cells.

    PubMed

    Ning, H; Lin, G; Lue, T F; Lin, C-S

    2013-01-01

    In erectile dysfunction (ED) research, monocultures of cavernous endothelial cells (CECs) and smooth muscle cells (CSMCs) have been reported, but a CEC-CSMC coculture system is still lacking. In the present study, we wished to investigate the feasibility of setting up such a system and test whether it can be used for diabetic ED research. Cavernous tissues were obtained from patients undergoing surgery for penile prosthesis. CSMCs were isolated by explant culture and verified by calponin staining. CECs were isolated by binding to CD31 antibody, followed by magnetic capture. These CECs were nearly 100% pure endothelial cells as determined by flow cytometric analysis for endothelial markers CD31, vWF and eNOS. Functional analyses, that is, low-density lipoprotein (LDL) uptake and capillary tube formation, also confirmed their endothelial phenotype. When cocultured with CSMCs, CECs formed capillary-like structures, and based on the extent of this capillary-like network, it was determined that a ratio of 1:4 in cell number between CECs and CSMCs was better than ratios of 1:1 and 1:9. It was also found that direct contact between CECs and CSMCs was necessary and a coculture period of 3 weeks was optimal. Autologous CSMCs were better than allogeneic CSMCs, and fibroblasts were completely incompetent. When treated with high glucose (25 mM), the CEC-CSMC coculture expressed significantly lower level of CD31 but significantly higher level of collagen-IV (Col-IV), and the diameter of the capillaries increased significantly, when compared with normal glucose (5 mM)-treated cocultures. These data are consistent with previously observed changes in the cavernous tissues of diabetic patients and thus suggest that the coculture system could be utilized for diabetic ED research.

  4. Arterial identity of endothelial cells is controlled by local cues.

    PubMed

    Othman-Hassan, K; Patel, K; Papoutsi, M; Rodriguez-Niedenführ, M; Christ, B; Wilting, J

    2001-09-15

    The ephrins and their Eph receptors comprise the largest family of receptor tyrosine kinases. Studies on mice have revealed an important function of ephrin-B2 and Eph-B4 for the development of the arterial and venous vasculature, respectively, but the mechanisms regulating their expression have not been studied yet. We have cloned a chick ephrin-B2 cDNA probe. Expression was observed in endothelial cells of extra- and intraembryonic arteries and arterioles in all embryos studied from day 2 (stage 10 HH, before perfusion of the vessels) to day 16. Additionally, expression was found in the somites and neural tube in early stages, and later also in the smooth muscle cells of the aorta, parts of the Müllerian duct, dosal neural tube, and joints of the limbs. We isolated endothelial cells from the internal carotid artery and the vena cava of 14-day-old quail embryos and grafted them separately into day-3 chick embryos. Reincubation was performed until day 6 and the quail endothelial cells were identified with the QH1 antibody. The grafted arterial and venous endothelial cells expressed ephrin-B2 when they integrated into the lining of arteries. Cells that were not integrated into vessels, or into vessels other than arteries, were ephrin-B2-negative. The studies show that the expression of the arterial marker ephrin-B2 is controlled by local cues in arterial vessels of older embryos. Physical forces or the media smooth muscle cells may be involved in this process.

  5. Therapeutic transdifferentiation of human fibroblasts into endothelial cells using forced expression of lineage-specific transcription factors.

    PubMed

    Wong, Wing Tak; Cooke, John P

    2016-01-01

    Transdifferentiation is the direct conversion from one somatic cell type into another desired somatic cell type. This reprogramming method offers an attractive approach for regenerative medicine. Here, we demonstrate that neonatal fibroblasts can be transdifferentiated into endothelial cells using only four endothelial transcription factors, namely, ETV2, FLI1, GATA2, and KLF4. We observed a significant up-regulation of endothelial genes including KDR, CD31, CD144, and vWF in human neonatal foreskin (BJ) fibroblasts infected with the lentiviral construct encoding the open reading frame of the four transcription factors. We observed morphological changes in BJ fibroblasts from the fibroblastic spindle shape into a more endothelial-like cobblestone structures. Fluorescence-activated cell sorting analysis revealed that ~16% of the infected cells with the lentiviral constructs encoding 4F expressed CD31. The sorted cells were allowed to expand for 2 weeks and these cells were immunostained and found to express endothelial markers CD31. The induced endothelial cells also incorporated fluorescence-labeled acetylated low-density lipoprotein and efficiently formed capillary-like networks when seeded on Matrigel. These results suggested that the induced endothelial cells were functional in vitro. Taken together, we successfully demonstrated the direct conversion of human neonatal fibroblasts into endothelial cells by transduction of lentiviral constructs encoding endothelial lineage-specific transcription factors ETV2, FLI1, GATA2, and KLF4. The directed differentiation of fibroblasts into endothelial cells may have significant utility in diseases characterized by fibrosis and loss of microvasculature.

  6. Ischemia-induced endothelial cell dysfunction.

    PubMed

    Keep, R F; Andjelkovic, A V; Stamatovic, S M; Shakui, P; Ennis, S R

    2005-01-01

    Hemorrhagic transformation upon reperfusion therapy has focused attention on ischemia-induced endothelial dysfunction. This study examined whether hyperglycemia may induce hemorrhagic transformation by enhancing endothelial mitochondrial damage during ischemia and whether preconditioning (PC) stimuli may limit ischemia-induced endothelial damage. In vivo, rats received 2.8 M D-glucose or arabinose (1 ml/100 g; i.p.) prior to undergoing two hours of middle cerebral artery occlusion and transcardiac fixation for electron microscopy. In vitro, brain endothelial cells were exposed to a PC impulse (short-term oxygen glucose deprivation; OGD) prior to an injurious event (5 hours OGD). Endothelial injury was assessed by measuring lactate dehydrogenase release. Hyperglycemia during cerebral ischemia resulted in marked changes in endothelial morphology and mitochondrial swelling. Thus, in the ischemic hemisphere, there was no evidence of endothelial mitochondrial swelling in normoglycemic rats (mean profile width 0.22 +/- 0.04 vs. 0.17 +/- 0.01 microm in contralateral hemisphere) but there was marked swelling in hyperglycemic rats (0.44 +/- 0.02 microm). In vitro, cells preconditioned with one hour of OGD one day prior to 5 hours of OGD, showed reduced lactate dehydrogenase release (p < 0.05). In conclusion, hyperglycemia may have specific adverse effects on endothelial cell mitochondria during ischemia. Preventing those effects may help to ameliorate blood-brain barrier disruption on reperfusion. Insights into how to prevent endothelial injury may come from determining the mechanisms involved in endothelial preconditioning.

  7. Beta-lactam antibiotic-mediated changes in platelet reactivity and vascular endothelial functions.

    PubMed

    Togna, G I; Togna, A R; Caprino, L

    2001-05-01

    To evaluate vascular and platelet compatibility of intravenous administration of beta-lactam antibiotics, we assessed the effects of therapeutic concentrations of ceftriaxone, aztreonam, and ceftazidime on platelet reactivity to different agonists (sodium arachidonate, collagen and adenosine diphosphate) and on selected vascular endothelial functions (adenosine diphosphatase activity, prostacyclin production and t-PA release). Ceftriaxone and, to a lesser degree, aztreonam, enhanced platelet reactivity, evaluated as onset of platelet aggregating response, and increased thromboxane production to subthreshold concentrations of arachidonate. There was no modification in platelet reactivity after ceftazidime treatment. Ceftriaxone and ceftazidime, but not aztreonam, inhibited endothelial adenosine diphosphatase activity. Prostacyclin production and t-PA release were inhibited only by ceftriaxone at high concentrations. While it is difficult to establish which marker (platelet or endothelial functions) has more clinical reference in human vascular compatibility, it seems feasible to consider aztreonam the most compatible of the beta-lactams studied.

  8. Benidipine improves oxidized LDL-dependent monocyte and endothelial dysfunction in hypertensive patients with type 2 diabetes mellitus.

    PubMed

    Nomura, S; Shouzu, A; Omoto, S; Nishikawa, M; Iwasaka, T

    2005-07-01

    We investigated the effects of long-term benidipine treatment on levels of monocyte and endothelial cell activation markers in hypertensive patients with (n = 28) and without (n = 10) type 2 diabetes mellitus. Benidipine, 4 mg/day, was administered for 6 months; there were no other changes in any of the patients pharmacologic regimens during benidipine treatment. Clinical and biochemical data obtained before and after benidipine administration were compared; all markers were measured by ELISA. The levels of platelet activation markers (CD62P, CD63, and PAC-1), microparticles (monocyte-derived microparticles: MDMPs, and endothelial cell-derived microparticles: EDMPs), chemokines (monocyte chemotactic peptide 1: MCP-1, regulated on activation normally T-cell expressed and secreted: RANTES) and soluble adhesion markers (soluble E-selectin and soluble ICAM-1) differed in the control and hypertension groups. In addition, levels of platelet, monocyte, and endothelial cell activation markers, microparticles, chemokines, and soluble adhesion molecules were higher in hypertensive patients than in those without type 2 diabetes. Furthermore, benidipine administration decreased the concentrations of all these markers. The effect of this drug was significant in diabetes patients with high levels of antioxidized low-density lipoprotein (LDL) antibody. These results suggest that benidipine is effective for the treatment of oxLDL-dependent vascular disorders in hypertensive patients with type 2 diabetes.

  9. Polyphenols in preventing endothelial dysfunction.

    PubMed

    Biegańska-Hensoldt, Sylwia; Rosołowska-Huszcz, Danuta

    2017-03-27

    One of the main causes of mortality in developed countries is atherosclerosis. The pathogenesis of atherosclerosis is associated with endothelial dysfunction. Consumption of food rich in natural antioxidants including polyphenols significantly improves endothelial cells functions. Polyphenols have a beneficial effect on the human body and play an important part in protecting the cardiovascular system. Polyphenols present in food have antioxidant, anti-inflammatory, antihypertensive, antithrombotic and antiproliferative properties. Catechins cause an increase in the activity of endothelial nitric oxide synthase (eNOS) and increased production of nitric oxide (NO) and decrease in blood pressure. Catechins also reduce platelet adhesion, lower the concentration of C-reactive protein and tumor necrosis factor alpha and interleukin-6. Resveratrol inhibits NADPH oxidase expression, increases the expression of eNOS and NO production as well as decreases the expression of proinflammatory cytokines, and also lowers the concentration of the soluble forms of adhesion molecules - sICAM-1 and sVCAM-1 in blood. Quercetin reduces the blood level of low density lipoprotein cholesterol, lowers blood pressure, reduces the concentration of C-reactive protein and F2-isoprostane level. Curcumin has antagonistic activity to homocysteine. Curcumin increases the expression of eNOS and reduces oxidative DNA damage in rat cardiomyocytes. Numerous attempts are taken for improving the bioavailability of polyphenols in order to increase their use in the body.

  10. Mussel-inspired immobilization of vascular endothelial growth factor (VEGF) for enhanced endothelialization of vascular grafts.

    PubMed

    Shin, Young Min; Lee, Yu Bin; Kim, Seok Joo; Kang, Jae Kyeong; Park, Jong-Chul; Jang, Wonhee; Shin, Heungsoo

    2012-07-09

    Most polymeric vascular prosthetic materials have low patency rate for replacement of small diameter vessels (<5 mm), mainly due to failure to generate healthy endothelium. In this study, we present polydopamine-mediated immobilization of growth factors on the surface of polymeric materials as a versatile tool to modify surface characteristics of vascular grafts potentially for accelerated endothelialization. Polydopamine was deposited on the surface of biocompatible poly(L-lactide-co-ε-caprolactone) (PLCL) elastomer, on which vascular endothelial growth factor (VEGF) was subsequently immobilized by simple dipping. Surface characteristics and composition were investigated by using scanning electron microscopy, atomic force microscopy, and X-ray photoelectron spectroscopy. Immobilization of VEGF on the polydopamine-deposited PLCL films was effective (19.8 ± 0.4 and 197.4 ± 19.7 ng/cm(2) for DPv20 and DPv200 films, respectively), and biotin-mediated labeling of immobilized VEGF revealed that the fluorescence intensity increased as a function of the concentration of VEGF solution. The effect of VEGF on adhesion of HUVECs was marginal, which may have been masked by polydopamine layer that also enhanced cell adhesion. However, VEGF-immobilized substrate significantly enhanced proliferation of HUVECs for over 7 days of in vitro culture and also improved their migration. In addition, immobilized VEGF supported robust cell to cell interactions with strong expression of CD 31 marker. The same process was effective for immobilization of basic fibroblast growth factor, demonstrating the robustness of polydopamine layer for secondary ligation of growth factors as a simple and novel surface modification strategy for vascular graft materials.

  11. Differentiation of human endometrial stem cells into endothelial-like cells on gelatin/chitosan/bioglass nanofibrous scaffolds.

    PubMed

    Shamosi, Atefeh; Mehrabani, Davood; Azami, Mahmoud; Ebrahimi-Barough, Somayeh; Siavashi, Vahid; Ghanbari, Hossein; Sharifi, Esmaeel; Roozafzoon, Reza; Ai, Jafar

    2017-02-01

    The capacity of gelatin/chitosan/bioactive glass nanopowders (GEL/CS/BGNPs) scaffolds was investigated for increasing human endometrial stem cells (hEnSCs) differentiation into the endothelial cells in the presence of angiogenic factors. GEL/CS nanofibrous scaffold with different contents of BGNPs were fabricated and assessed. Expression of endothelial markers (CD31, vascular endothelial cadherin (VE-cadherin), and KDR) in differentiated cells was evaluated. Results showed the diameter of nanofiber increases with decreasing the BG content in GEL/CS scaffolds. Moreover, in vitro study indicated that the GEL/CS/BGNPs scaffold with 1.5% BGNPs content provided a suitable three-dimensional structure for endothelial cells differentiation. Thus, the GEL/CS/BGNPs scaffold can be recommended for blood vessels repair.

  12. [Endothelial dysfunction and nonspecific immune reactions in development and progression of osteoarthrosis in women engaged into manual work].

    PubMed

    Maliutina, N N; Nevzorova, M S

    2015-01-01

    The article considers mechanisms of development and progression of osteoarthrosis as an occupationally conditioned disease in women of manual work. Women working in physical overstrain conditions are under occupational risk with dysfunction of many body systems. The authors set a hypothesis on association of endothelial dysfunction markers dysbalance and structural remodelling of cartilage matrix as a proof of degenerative changes.

  13. Purification and long-term expansion of multipotent endothelial-like cells with potential cardiovascular regeneration.

    PubMed

    Marchal, Juan A; Picón, Manuel; Perán, Macarena; Bueno, Clara; Jiménez-Navarro, Manuel; Carrillo, Esmeralda; Boulaiz, Houria; Rodríguez, Noela; Álvarez, Pablo; Menendez, Pablo; de Teresa, Eduardo; Aránega, Antonia

    2012-03-01

    Endothelial progenitor cells (EPC) represent a relatively rare cell population, and expansion of sufficient cell numbers remains a challenge. Nevertheless, human adipose-derived stem cells (hASC) can be easily isolated and possess the ability to differentiate into endothelial cells. Here, we propose the isolation and characterization of multipotent endothelial-like cells (ME-LC) with the capacity to maintain their vascular progenitor properties for long periods. hASC were isolated from lipoaspirates and cultured through distinct consecutive culture stages for 2 months to enrich ME-LC: first in Dulbecco's modified Eagle's medium-fetal bovine serum (stage I), followed by a stage of culture in absent of fetal bovine serum (stage II), a culture in SFO3 medium (stage III), and, finally, the culture of ME-LC into collagen IV-coated flasks in endothelial growth medium (EGM-2) (stage IV). ME-LC display increased expression levels of endothelial and hematopoietic lineage markers (CD45, KDR, and CXCR4) and EPC markers (CD34 and CD133), whereas the expression of CD31 was barely detectable. Reverse transcription (RT)-polymerase chain reaction assays showed expression of genes involved in early stages of EPC differentiation and decreased expression of genes associated to differentiated EPC (TIE-2, DLL4, and FLT-1). ME-LC formed capillary-like structures when grown on Matrigel, secreted increased levels of stromal cell-derived factor-1 (SDF-1), and showed the ability to migrate attracted by SDF-1, vascular endothelial growth factor, and hematopoietic growth factor cytokines. Importantly, ME-LC retained the capacity to differentiate into cardiomyocyte-like cells. We present a simplified and efficient method to generate large numbers of autologous ME-LC from lipoaspirates-derived hASC, opening up potential cell-based therapies for cardiovascular regenerative medicine.

  14. Metabolism of vasoactive peptides by human endothelial cells in culture. Angiotensin I converting enzyme (kininase II) and angiotensinase.

    PubMed

    Johnson, A R; Erdös, E G

    1977-04-01

    Cultured endothelial cells provide a model for the study of interactions of vasoactive peptides with endothelium. Endothelial cell cultured from veins of human umbilical cords contain both angiotensin I converting enzyme (kininase II) and angiotensinase activities. Intact monolayers of cells can both activate angiotensin I and inactivate bradykinin when the peptides are added to culture flasks in protein-free medium. Intact suspended cells or lysed cells convert angiotensin I to angiotensin II, inactivate bradykinin, and hydrolyze hippuryldiglycine to hippuric acid and diglycine. These actions are inhibited by SQ 20881, the specific inhibitor of converting enzyme. The kininase activity of endothelial cells was partially inhibited by antibody to human lung converting enzyme. Endothelial cells also inactivate longer analogs of bradykinin, such as kallidin, methionyl-lysyl bradykinin, and bradykinin coupled covalently to 500,000 mol wt dextran. The endothelial cells retained converting enzyme activity through four successive subcultures, indicating that the enzyme is synthesized by the cells surface, and it is apparently a marker for endothelial cells, since cultured human fibroblasts, smooth muscle cells, and baby hamster kidney cells do not have it. Endothelial cells also contain an aminopheptidase which hydrolyzes both angiotensin II and the synthetic substrate, alpha-L-aspartyl beta-naphthylamide. The angiotensinase activity increased when the cells were lysed, which suggests that the enzyme is localized within the cells, Hydrolysis of both alpha-L-aspartyl beta-naphthylamide and angiotensin II was inhibited by omicron-phenanthroline, indicating that the enzyme is an A-tipe anigotensinase.

  15. Long-Term Exposure to AZT, but not d4T, Increases Endothelial Cell Oxidative Stress and Mitochondrial Dysfunction

    PubMed Central

    Kline, Erik R.; Bassit, Leda; Hernandez-Santiago, Brenda I.; Detorio, Mervi A.; Liang, Bill; Kleinhenz, Dean J.; Walp, Erik R.; Dikalov, Sergey; Jones, Dean P.; Schinazi, Raymond F.

    2009-01-01

    Nucleoside reverse transcriptase inhibitors (NRTIs), such as zidovudine (AZT) and stavudine (d4T), cause toxicities to numerous tissues, including the liver and vasculature. While much is known about hepatic NRTI toxicity, the mechanism of toxicity in endothelial cells is incompletely understood. Human aortic endothelial and HepG2 liver cells were exposed to 1 μM AZT or d4T for up to 5 weeks. Markers of oxidative stress, mitochondrial function, NRTI phosphorylation, mitochondrial DNA (mtDNA) levels, and cytotoxicity were monitored over time. In endothelial cells, AZT significantly oxidized glutathione redox potential, increased total cellular and mitochondrial-specific superoxide, decreased mitochondrial membrane potential, increased lactate release, and caused cell death from weeks 3 through 5. Toxicity occurred in the absence of di- and tri-phosphorylated AZT and mtDNA depletion. These data show that oxidative stress and mitochondrial dysfunction in endothelial cells occur with a physiologically relevant concentration of AZT, and require long-term exposure to develop. In contrast, d4T did not induce endothelial oxidative stress, mitochondrial dysfunction, or cytotoxicity despite the presence of d4T-triphosphate. Both drugs depleted mtDNA in HepG2 cells without causing cell death. Endothelial cells are more susceptible to AZT-induced toxicity than HepG2 cells, and AZT caused greater endothelial dysfunction than d4T because of its pro-oxidative effects. PMID:19067249

  16. Nucleoside reverse transcriptase inhibitors induce a mitophagy-associated endothelial cytotoxicity that is reversed by coenzyme Q10 cotreatment.

    PubMed

    Xue, Stephen Y; Hebert, Valeria Y; Hayes, Danicia M; Robinson, Corie N; Glover, Mitzi; Dugas, Tammy R

    2013-08-01

    Cardiovascular complications have been documented in HIV-1 infected populations, and antiretroviral therapy may play a role. Nucleoside reverse transcriptase inhibitors (NRTIs) are antiretrovirals known to induce mitochondrial damage in endothelial cells, culminating in endothelial dysfunction, an initiating event in atherogenesis. Though the mechanism for NRTI-induced endothelial toxicity is not yet clear, our prior work suggested that a mitochondrial oxidative stress may be involved. To further delineate the mechanism of toxicity, endothelial cells were treated with NRTIs of varying subclasses, and the level of reactive oxygen species (ROS) and mitochondrial function were assessed. To test whether rescue of mitochondrial electron transport attenuated NRTI-induced endothelial cytotoxicity, in some cases, cells were cotreated with the electron transport cofactor coenzyme Q10 (Q10). At 4-6h, NRTIs increased levels of ROS but decreased the activities of electron transport chain complexes I-IV, levels of ATP and the NAD/NADH ratio. Moreover, nitric oxide levels were decreased, whereas endothelin-1 release was increased. Q10 abolished NRTI-induced mitochondria injury and effects on endothelial agonist production. Interestingly, in cells treated with NRTIs only, markers for mitochondrial toxicity returned to baseline levels by 18-24h, suggesting a compensatory mechanism for clearing damaged mitochondria. Using confocal microscopy, with confirmation utilizing the autophagy and mitophagy markers LC-3 and Nix, respectively, we observed autophagy of mitochondria at 8-10h after treatment. Q10 prevented NRTI-mediated increase in LC-3. These findings suggest that NRTI-induced mitophagy may be involved in NRTI-induced endothelial dysfunction and that this damage likely results from oxidant injury. Further, Q10 supplementation could potentially prevent NRTI-induced endothelial dysfunction.

  17. BMP-mediated specification of the erythroid lineage suppresses endothelial development in blood island precursors

    PubMed Central

    Myers, Candace T.

    2013-01-01

    The developmental relationship between the blood and endothelial cell (EC) lineages remains unclear. In the extra-embryonic blood islands of birds and mammals, ECs and blood cells are closely intermixed, and blood island precursor cells in the primitive streak express many of the same molecular markers, leading to the suggestion that both lineages arise from a common precursor, called the hemangioblast. Cells within the blood island of Xenopus also coexpress predifferentiation markers of the blood and EC lineages. However, using multiple assays, we find that precursor cells in the Xenopus blood island do not normally differentiate into ECs, suggesting that classic hemangioblasts are rare or nonexistent in Xenopus. What prevents these precursor cells from developing into mature ECs? We have found that bone morphogenetic protein (BMP) signaling is essential for erythroid differentiation, and in the absence of BMP signaling, precursor cells adopt an EC fate. Furthermore, inhibition of the erythroid transcription pathway leads to endothelial differentiation. Our results indicate that bipotential endothelial/erythroid precursor cells do indeed exist in the Xenopus blood island, but BMP signaling normally acts to constrain EC fate. More generally, these results provide evidence that commitment to the erythroid lineage limits development of bipotential precursors toward an endothelial fate. PMID:24100450

  18. Immunofluorescence identifies distinct subsets of endothelial cells in the human liver

    PubMed Central

    Strauss, Otto; Phillips, Anthony; Ruggiero, Katya; Bartlett, Adam; Dunbar, P. Rod

    2017-01-01

    As well as systemic vascular endothelial cells, the liver has specialised sinusoidal endothelial cells (LSEC). LSEC dysfunction has been documented in many diseased states yet their phenotype in normal human liver has not been comprehensively assessed. Our aim was to improve characterisation of subsets of endothelial cells and associated pericytes in the human liver. Immunofluorescence microscopy was performed on normal human liver tissue samples to assess endothelial and structural proteins in a minimum of three donors. LSEC are distributed in an acinar pattern and universally express CD36, but two distinctive subsets of LSEC can be identified in different acinar zones. Type 1 LSEC are CD36hiCD32−CD14−LYVE-1− and are located in acinar zone 1 of the lobule, while Type 2 LSEC are LYVE-1+CD32hiCD14+CD54+CD36mid-lo and are located in acinar zones 2 and 3 of the lobule. Portal tracts and central veins can be identified using markers for systemic vascular endothelia and pericytes, none of which are expressed by LSEC. In areas of low hydrostatic pressure LSEC are lined by stellate cells that express the pericyte marker CD146. Our findings identify distinctive populations of LSEC and distinguish these cells from adjacent stellate cells, systemic vasculature and pericytes in different zones of the liver acinus. PMID:28287163

  19. Innovative Flow Cytometry Allows Accurate Identification of Rare Circulating Cells Involved in Endothelial Dysfunction

    PubMed Central

    Boraldi, Federica; Bartolomeo, Angelica; De Biasi, Sara; Orlando, Stefania; Costa, Sonia; Cossarizza, Andrea; Quaglino, Daniela

    2016-01-01

    Introduction Although rare, circulating endothelial and progenitor cells could be considered as markers of endothelial damage and repair potential, possibly predicting the severity of cardiovascular manifestations. A number of studies highlighted the role of these cells in age-related diseases, including those characterized by ectopic calcification. Nevertheless, their use in clinical practice is still controversial, mainly due to difficulties in finding reproducible and accurate methods for their determination. Methods Circulating mature cells (CMC, CD45-, CD34+, CD133-) and circulating progenitor cells (CPC, CD45dim, CD34bright, CD133+) were investigated by polychromatic high-speed flow cytometry to detect the expression of endothelial (CD309+) or osteogenic (BAP+) differentiation markers in healthy subjects and in patients affected by peripheral vascular manifestations associated with ectopic calcification. Results This study shows that: 1) polychromatic flow cytometry represents a valuable tool to accurately identify rare cells; 2) the balance of CD309+ on CMC/CD309+ on CPC is altered in patients affected by peripheral vascular manifestations, suggesting the occurrence of vascular damage and low repair potential; 3) the increase of circulating cells exhibiting a shift towards an osteoblast-like phenotype (BAP+) is observed in the presence of ectopic calcification. Conclusion Differences between healthy subjects and patients with ectopic calcification indicate that this approach may be useful to better evaluate endothelial dysfunction in a clinical context. PMID:27560136

  20. eNOS-Dependent Antisenscence Effect of a Calcium Channel Blocker in Human Endothelial Cells

    PubMed Central

    Hayashi, Toshio; Yamaguchi, Tomoe; Sakakibara, Yasufumi; Taguchi, Kumiko; Maeda, Morihiko; Kuzuya, Masafumi; Hattori, Yuichi

    2014-01-01

    Senescence of vascular endothelial cells is an important contributor to the pathogenesis of age-associated vascular disorders such as atherosclerosis. We investigated the effects of antihypertensive agents on high glucose-induced cellular senescence in human umbilical venous endothelial cells (HUVECs). Exposure of HUVECs to high glucose (22 mM) for 3 days increased senescence-associated- β-galactosidase (SA-β-gal) activity, a senescence marker, and decreased telomerase activity, a replicative senescence marker. The calcium channel blocker nifedipine, but not the β1-adrenergic blocking agent atenolol or the angiotensin-converting enzyme inhibitor perindopril, reduced SA-β-gal positive cells and prevented a decrease in telomerase activity in a high-glucose environment. This beneficial effect of nifedipine was associated with reduced reactive oxygen species (ROS) and increased endothelial nitric oxide synthase (eNOS) activity. Thus, nifedipine prevented high glucose-induced ROS generation and increased basal eNOS phosphorylation level at Ser-1177. Treatment with NG-nitro-L-arginine (L-NAME) and transfection of small interfering RNA (siRNA) targeting eNOS eliminated the anti-senscence effect of nifedipine. These results demonstrate that nifedipine can prevent endothelial cell senescence in an eNOS-dependent manner. The anti-senescence action of nifedipine may represent a novel mechanism by which it protects against atherosclerosis. PMID:24520379

  1. Biomimetic, ultrathin and elastic hydrogels regulate human neutrophil extravasation across endothelial-pericyte bilayers

    PubMed Central

    Lauridsen, Holly M.

    2017-01-01

    The vascular basement membrane—a thin, elastic layer of extracellular matrix separating and encasing vascular cells—provides biological and mechanical cues to endothelial cells, pericytes, and migrating leukocytes. In contrast, experimental scaffolds typically used to replicate basement membranes are stiff and bio-inert. Here, we present thin, porated polyethylene glycol hydrogels to replicate human vascular basement membranes. Like commercial transwells, our hydrogels are approximately 10μm thick, but like basement membranes, the hydrogels presented here are elastic (E: 50-80kPa) and contain a dense network of small pores. Moreover, the inclusion of bioactive domains introduces receptor-mediated biochemical signaling. We compare elastic hydrogels to common culture substrates (E: >2GPa) for human endothelial cell and pericyte monolayers and bilayers to replicate postcapillary venules in vitro. Our data demonstrate that substrate elasticity facilitates differences in vascular phenotype, supporting expression of vascular markers that are increasingly replicative of venules. Endothelial cells differentially express vascular markers, like EphB4, and leukocyte adhesion molecules, such as ICAM-1, with decreased mechanical stiffness. With porated PEG hydrogels we demonstrate the ability to evaluate and observe leukocyte recruitment across endothelial cell and pericyte monolayers and bilayers, reporting that basement membrane scaffolds can significantly alter the rate of vascular migration in experimental systems. Overall, this study demonstrates the creation and utility of a new and accessible method to recapture the mechanical and biological complexity of human basement membranes in vitro. PMID:28234918

  2. New insights into Shiga toxin-mediated endothelial dysfunction in hemolytic uremic syndrome

    PubMed Central

    Petruzziello-Pellegrini, Tania N; Moslemi-Naeini, Mozhgan; Marsden, Philip A

    2013-01-01

    Shiga toxin-producing E. coli represents a significant global health concern, especially as hypervirulent pathogens surface amidst outbreaks of hemolytic uremic syndrome (HUS). Shiga toxin (Stx) is key in the microangiopathic events underlying the disease and its central role is underscored by the unprecedented HUS outbreak in Germany in 2011. The mechanisms of Stx-mediated endothelial dysfunction have been a major focus of research that has contributed to the current understanding of the pathogenic changes in endothelial phenotype leading to HUS. Among the newer concepts are Stx-mediated gene regulation in the absence of protein synthesis inhibition, a potential role for complement activation, and accumulating evidence for detectable serum markers before the onset of the classic clinical features of HUS. Further investigation of newer therapeutic targets and potential prognostic markers is essential to assess their utility in mitigating disease and/or predicting outcomes and will provide an improved overall understanding of HUS pathogenesis. PMID:23955166

  3. Vascular progenitor cells isolated from human embryonic stem cells give rise to endothelial and smooth muscle like cells and form vascular networks in vivo.

    PubMed

    Ferreira, Lino S; Gerecht, Sharon; Shieh, Hester F; Watson, Nicki; Rupnick, Maria A; Dallabrida, Susan M; Vunjak-Novakovic, Gordana; Langer, Robert

    2007-08-03

    We report that human embryonic stem cells contain a population of vascular progenitor cells that have the ability to differentiate into endothelial-like and smooth muscle (SM)-like cells. Vascular progenitor cells were isolated from EBs grown in suspension for 10 days and were characterized by expression of the endothelial/hematopoietic marker CD34 (CD34+ cells). When these cells are subsequently cultured in EGM-2 (endothelial growth medium) supplemented with vascular endothelial growth factor-165 (50 ng/mL), they give rise to endothelial-like cells characterized by a cobblestone cell morphology, expression of endothelial markers (platelet endothelial cell-adhesion molecule-1, CD34, KDR/Flk-1, vascular endothelial cadherin, von Willebrand factor), incorporation of acetylated low-density lipoprotein, and formation of capillary-like structures when placed in Matrigel. In contrast, when CD34+ cells are cultured in EGM-2 supplemented with platelet-derived growth factor-BB (50 ng/mL), they give rise to SM-like cells characterized by spindle-shape morphology, expression of SM cell markers (alpha-SM actin, SM myosin heavy chain, calponin, caldesmon, SM alpha-22), and the ability to contract and relax in response to common pharmacological agents such as carbachol and atropine but rarely form capillary-like structures when placed in Matrigel. Implantation studies in nude mice show that both cell types contribute to the formation of human microvasculature. Some microvessels contained mouse blood cells, which indicates functional integration with host vasculature. Therefore, the vascular progenitors isolated from human embryonic stem cells using methods established in the present study could provide a means to examine the mechanisms of endothelial and SM cell development, and they could also provide a potential source of cells for vascular tissue engineering.

  4. Progenitor endothelial cell involvement in Alzheimer's disease

    SciTech Connect

    Budinger, Thomas F.

    2003-05-01

    There is compelling evidence that endothelial cells of the brain and periphery are dysfunctional in Alzheimer's Disease. There is evidence for a fundamental defect in, or abnormal aging of, endothelial progenitor cells in atherosclerosis. The possibility that endothelial cell defects are a primary cause for Alzheimer's Disease or other dementias can be researched by molecular and cell biology studies as well as cell trafficking studies using recently demonstrated molecular imaging methods. The evidence for abnormal endothelial function and the methods to explore this hypothesis are presented.

  5. Cytokines as biochemical markers for knee osteoarthritis

    PubMed Central

    Mabey, Thomas; Honsawek, Sittisak

    2015-01-01

    Osteoarthritis (OA) is a debilitating degenerative joint disease particularly affecting weightbearing joints within the body, principally the hips and knees. Current radiographic techniques are insufficient to show biochemical changes within joint tissue which can occur many years before symptoms become apparent. The need for better diagnostic and prognostic tools is heightened with the prevalence of OA set to increase in aging and obese populations. As inflammation is increasingly being considered an important part of OAs pathophysiology, cytokines are being assessed as possible candidates for biochemical markers. Cytokines, both pro- and anti-inflammatory, as well as angiogenic and chemotactic, have in recent years been studied for relevant characteristics. Biochemical markers show promise in determination of the severity of disease in addition to monitoring of the efficacy and safety of disease-modifying OA drugs, with the potential to act as diagnostic and prognostic tools. Currently, the diagnostic power of interleukin (IL)-6 and the relationship to disease burden of IL-1β, IL-15, tumor necrosis factor-α, and vascular endothelial growth factor make these the best candidates for assessment. Grouping appropriate cytokine markers together and assessing them collectively alongside other bone and cartilage degradation products will yield a more statistically powerful tool in research and clinical applications, and additionally aid in distinguishing between OA and a number of other diseases in which cytokines are known to have an involvement. Further large scale studies are needed to assess the validity and efficacy of current biomarkers, and to discover other potential biomarker candidates. PMID:25621214

  6. Proliferation, angiogenesis and differentiation related markers in compact and follicular-compact thyroid carcinomas in dogs

    PubMed Central

    Pessina, P.; Castillo, V.A.; César, D.; Sartore, I.; Meikle, A.

    2016-01-01

    Immunohistochemical markers (IGF-1, IGF-1R, VEGF, FGF-2, RARα and RXR) were evaluated in healthy canine thyroid glands (n=8) and in follicular-compact (n=8) and compact thyroid carcinomas (n=8). IGF-1, IGF-1R and VEGF expression was higher in fibroblasts and endothelial cells of compact carcinoma than in healthy glands (P < 0.05). Compared to follicular-compact carcinoma, compact carcinoma had higher IGF-1R expression in fibroblasts, and higher FGF-2 expression in endothelial cells (P < 0.05). RARα expression was higher in endothelial cells of compact carcinoma than in those of other groups (P < 0.05). The upregulation of these proliferation- and angiogenesis-related factors in endothelial cells and/or fibroblasts and not in follicular cells of compact carcinoma compared to healthy glands supports the relevance of stromal cells in cancer progression. PMID:28116249

  7. Monoclonal endothelial cell proliferation is present in primary but not secondary pulmonary hypertension.

    PubMed Central

    Lee, S D; Shroyer, K R; Markham, N E; Cool, C D; Voelkel, N F; Tuder, R M

    1998-01-01

    The etiology and pathogenesis of the vascular lesions characterizing primary pulmonary hypertension (PPH), an often fatal pulmonary vascular disease, are largely unknown. Plexiform lesions composed of proliferating endothelial cells occur in between 20 and 80% of the cases of this irreversible pulmonary vascular disease. Recently, technology to assess monoclonality has allowed the distinction between cellular proliferation present in neoplasms from that in reactive nonneoplastic tissue. To determine whether the endothelial cell proliferation in plexiform lesions in PPH is monoclonal or polyclonal, we assessed the methylation pattern of the human androgen receptor gene by PCR (HUMARA) in proliferated endothelial cells in plexiform lesions from female PPH patients (n = 4) compared with secondary pulmonary hypertension (PH) patients (n = 4). In PPH, 17 of 22 lesions (77%) were monoclonal. However, in secondary PH, all 19 lesions examined were polyclonal. Smooth muscle cell hyperplasia in pulmonary vessels (n = 11) in PPH and secondary PH was polyclonal in all but one of the examined vessels. The monoclonal expansion of endothelial cells provides the first marker that allows the distinction between primary and secondary PH. Our data of a frequent monoclonal endothelial cell proliferation in PPH suggests that a somatic genetic alteration similar to that present in neoplastic processes may be responsible for the pathogenesis of PPH. PMID:9486960

  8. Intravascularly Administered RGD-Displaying Measles Viruses Bind to and Infect Neovessel Endothelial Cells In Vivo

    PubMed Central

    Ong, Hooi Tin; Trejo, Theodore R; Pham, Linh D; Oberg, Ann L; Russell, Stephen J; Peng, Kah-Whye

    2009-01-01

    Systemically administered vectors must cross the endothelial lining of tumor blood vessels to access cancer cells. Vectors that interact with markers on the lumenal surface of these endothelial cells might have enhanced tumor localization. Here, we generated oncolytic measles viruses (MVs) displaying αvβ3 integrin-binding peptides, cyclic arginine-glycine-aspartate (RGD) or echistatin, on the measles hemagglutinin protein. Both viruses had expanded tropisms, and efficiently entered target cells via binding to integrins, but also retained their native tropisms for CD46 and signaling lymphocyte activation molecule (SLAM). When fluorescently labeled and injected intravascularly into chick chorioallantoic membranes (CAMs), in contrast to unmodified viruses, the integrin-binding viral particles bound to the lumenal surface of the developing chick neovessels and infected the CAM vascular endothelial cells. In a mouse model of VEGF-induced angiogenesis in the ear pinna, the integrin-binding viruses, but not the parental virus, infected cells at sites of new blood vessel formation. When given intravenously to mice bearing tumor xenografts, the integrin-binding virus infected endothelial cells of tumor neovessels in addition to tumor parenchyma. To our knowledge, this is the first report demonstrating that oncolytic MVs can be engineered to target the lumenal endothelial surface of newly formed blood vessels when administered intravenously in living animals. PMID:19277014

  9. Calcineurin inhibitors cyclosporine A and tacrolimus induce vascular inflammation and endothelial activation through TLR4 signaling

    PubMed Central

    Rodrigues-Diez, Raquel; González-Guerrero, Cristian; Ocaña-Salceda, Carlos; Rodrigues-Diez, Raúl R.; Egido, Jesús; Ortiz, Alberto; Ruiz-Ortega, Marta; Ramos, Adrián M.

    2016-01-01

    The introduction of the calcineurin inhibitors (CNIs) cyclosporine and tacrolimus greatly reduced the rate of allograft rejection, although their chronic use is marred by a range of side effects, among them vascular toxicity. In transplant patients, it is proved that innate immunity promotes vascular injury triggered by ischemia-reperfusion damage, atherosclerosis and hypertension. We hypothesized that activation of the innate immunity and inflammation may contribute to CNI toxicity, therefore we investigated whether TLR4 mediates toxic responses of CNIs in the vasculature. Cyclosporine and tacrolimus increased the production of proinflammatory cytokines and endothelial activation markers in cultured murine endothelial and vascular smooth muscle cells as well as in ex vivo cultures of murine aortas. CNI-induced proinflammatory events were prevented by pharmacological inhibition of TLR4. Moreover, CNIs were unable to induce inflammation and endothelial activation in aortas from TLR4−/− mice. CNI-induced cytokine and adhesion molecules synthesis in endothelial cells occurred even in the absence of calcineurin, although its expression was required for maximal effect through upregulation of TLR4 signaling. CNI-induced TLR4 activity increased O2−/ROS production and NF-κB-regulated synthesis of proinflammatory factors in cultured as well as aortic endothelial and VSMCs. These data provide new insight into the mechanisms associated with CNI vascular inflammation. PMID:27295076

  10. Sinusoidal endotheliitis as a histological parameter for diagnosing acute liver allograft rejection

    PubMed Central

    Shi, Yu; Dong, Kun; Zhang, Yu-Guo; Michel, René P; Marcus, Victoria; Wang, Yu-Yue; Chen, Yu; Gao, Zu-Hua

    2017-01-01

    AIM To investigated the feasibility of using sinusoidal endotheliitis (SE) as a histological marker for liver allograft rejection. METHODS We compared the histological features of 88 liver allograft biopsies with acute cellular rejection (ACR) and 59 cases with no evidence of ACR. SE was scored as: (1) focal linear lifting up of the endothelial cells by lymphocytes with no obvious damage to adjacent hepatocytes; (2) focal disruption of the endothelial lining by a cluster of subendothelial lymphocytes (a group of > 3 lymphocytes); and (3) severe confluent endotheliitis with hemorrhage and adjacent hepatocyte loss. RESULTS The sensitivity and specificity of SE was 81% and 85%, respectively. Using SE as the only parameter, the positive predictive value for ACR (PPV) was 0.89, whereas the negative predictive value for ACR (NPV) was 0.75. The correlation between RAI and SE was moderate (R = 0.44, P < 0.001) (Figure 3A), whereas it became strong (R = 0.65, P < 0.001) when correlating SE with the venous endotheliitis activity index only. CONCLUSION Our data suggest that SE scoring could be a reliable and reproducible supplemental parameter to the existing Banff schema for diagnosing acute liver allograft rejection. PMID:28223723

  11. Endothelial Cell Growth and Differentiation on Collagen-Immobilized Polycaprolactone Nanowire Surfaces.

    PubMed

    Leszczak, Victoria; Baskett, Dominique A; Popat, Ketul C

    2015-06-01

    The success of cardiovascular implants is associated with the development of an endothelium on material surface, critical to the prevention of intimal hyperplasia, calcification and thrombosis. A thorough understanding of the interaction between vascular endothelial cells and the biomaterial involved is essential in order to have a successful application which promotes healing and regeneration through integration with native tissue. In this study, we have developed collagen immobilized nanostructured surfaces with controlled arrays of high aspect ratio nanowires for the growth and maintenance of human microvascular endothelial cells (HMVECs). The nanowire surfaces were fabricated from polycaprolactone using a novel nanotemplating technique, and were immobilized with collagen utilizing an aminolysis method. The collagen immobilized nanowire surfaces were characterized using contact angle measurements, scanning electron microscopy and X-ray photoelectron spectroscopy. Human microvascular endothelial cells were used to evaluate the efficacy of the collagen immobilized nanowire surfaces to promote cell adhesion, proliferation, viability and differentiation. The results presented here indicate significantly higher cellular adhesion, proliferation and viability on nanowire and collagen immobilized surfaces as compared to the control surface. Further, HMVECs have a more elongated body and low shape factor on nanostructured surfaces. The differentiation potential of collagen immobilized nanowire surfaces was also evaluated by immunostaining and western blotting for key endothelial cell markers that are expressed when human microvascular endothelial cells are differentiated. Results indicate that expression of VE-cadherin is increased on collagen immobilized surfaces while the expression of von Willebrand factor is statistically similar on all surfaces.

  12. Anti-Inflammatory Properties of Sirtuin 6 in Human Umbilical Vein Endothelial Cells

    PubMed Central

    Lappas, Martha

    2012-01-01

    A prominent feature of inflammatory diseases is endothelial dysfunction. Factors associated with endothelial dysfunction include proinflammatory cytokines, adhesion molecules, and matrix degrading enzymes. At the transcriptional level, they are regulated by the histone deacetylase sirtuin (SIRT) 1 via its actions on the proinflammatory transcription factor nuclear factor-κB (NF-κB). The role of SIRT6, also a histone deacetylase, in regulating inflammation in endothelial cells is not known. The aim of this study was to determine the effect of SIRT6 knockdown on inflammatory markers in human umbilical vein endothelial cells (HUVECs) in the presence of lipopolysaccharide (LPS). LPS decreased expression of SIRT6 in HUVECs. Knockdown of SIRT6 increased the expression of proinflammatory cytokines (IL-1β, IL-6, IL-8), COX-prostaglandin system, ECM remodelling enzymes (MMP-2, MMP-9 and PAI-1), the adhesion molecule ICAM-1, and proangiogenic growth factors VEGF and FGF-2; cell migration; cell adhesion to leukocytes. Loss of SIRT6 increased the expression of NF-κB, whereas overexpression of SIRT6 was associated with decreased NF-κB transcriptional activity. Taken together, these results demonstrate that the loss of SIRT6 in endothelial cells is associated with upregulation of genes involved in inflammation, vascular remodelling, and angiogenesis. SIRT6 may be a potential pharmacological target for inflammatory vascular diseases. PMID:23132960

  13. Peripheral Blood Endothelial Progenitors as Potential Reservoirs of Kaposi's Sarcoma-Associated Herpesvirus

    PubMed Central

    Della Bella, Silvia; Brambilla, Lucia; Bellinvia, Monica; Bergamo, Elisa; Clerici, Mario; Villa, Maria L.

    2008-01-01

    Background The cellular reservoirs of Kaposi's sarcoma-associated herpesvirus (KSHV) and the exact nature of the putative KSHV-infected circulating precursor of spindle cells of Kaposi's sarcoma (KS) still remain poorly defined. Because KS spindle cells are thought to be of endothelial origin, and because mature endothelial cells do not sustain persistent KSHV-infection, our attention was focalized on circulating hematopoietic precursors able to differentiate into endothelial lineage. Methods and Findings Late endothelial progenitor cells (late-EPCs) were cultured from the peripheral blood mononuclear cells of 16 patients with classic KS. The presence and load of KSHV genomes were analyzed by real-time polymerase chain reaction in DNA extracted from cells and supernatants of late-EPC cultures obtained from 7 patients. Endothelial colonies cultured from the peripheral blood of KS patients were found to satisfy all requisites to be defined late-EPCs: they appeared from the CD14-negative fraction of adherent cells after 11–26 days of culture, could be serially expanded in vitro, expressed high levels of endothelial antigens but lacked leukocyte markers. Late-EPC cultures were found to harbor KSHV-DNA at variable levels and to retain the virus after multiple passages in cells as well as in supernatants, suggesting that a quote of KSHV lytic infection may spontaneously occur. Lytic phase induction or hypoxia could amplify virus release in supernatants. Conclusion Our results suggest that circulating endothelial progenitors from KS patients are KSHV-infected and support viral productive replication and may therefore represent potential virus reservoirs and putative precursors of KS spindle cells. PMID:18231605

  14. In vitro induction of human adipose-derived stem cells into lymphatic endothelial-like cells.

    PubMed

    Yang, Yi; Chen, Xiao-hu; Li, Fu-gui; Chen, Yun-xian; Gu, Li-qiang; Zhu, Jia-kai; Li, Ping

    2015-02-01

    Human adipose-derived stem cells (hADSCs) may provide a suitable number of progenitors for the treatment of lymphatic edema; however, to date the protocols for inducing hADSCs into this tissue type have not been standardized. We wished to investigate the induction of hADSCs into lymphatic endothelial-like cells using vascular endothelial growth factor-C156S (VEGF-C156S) and other growth factors in vitro. hADSCs from healthy adult adipose tissue were purified using enzyme digestion. Differentiation was induced using medium containing VEGF-C156S and bovine fibroblast growth factor (bFGF). Differentiation was confirmed using immunostaining for lymphatic vessel endothelial hyaluronan receptor (LYVE-1) and fms-related tyrosine kinase 4 (FLT-4), two lymphatic endothelial cell markers. The expression levels of LYVE-1, prospero homeobox 1 (PROX-1), and FLT-4 throughout induction were assessed using reverse transcriptase quantitative polymerase chain reaction. hADSCs were successfully obtained by trypsin digest and purification. Flow cytometry showed these cells were similar to mesenchymal stem cells, with a high positive rate of CD13, CD29, CD44, and CD105, and a low positive rate of CD31, CD34, CD45, and HLA-DR. Induction to lymphatic endothelial-like cells was successful, with cells expressing high levels of LYVE-1, PROX-1, and FLT-4. Adipose-derived stem cells can be induced to differentiate into lymphatic endothelial-like cells using a medium containing VEGF-C156S, bFGF, and other growth factors. This population of lymphatic endothelial-like cells may be useful for lymphatic reconstruction in the future.

  15. Circulating Endothelial Cells and Endothelial Function predict Major Adverse Cardiac Events and Early Adverse Left Ventricular Remodeling in Patients with ST-Segment Elevation Myocardial Infarction

    PubMed Central

    Magdy, Abdel Hamid; Bakhoum, Sameh; Sharaf, Yasser; Sabry, Dina; El-Gengehe, Ahmed T; Abdel-Latif, Ahmed

    2016-01-01

    Endothelial progenitor cells (EPCs) and circulating endothelial cells (CECs) are mobilized from the bone marrow and increase in the early phase after ST-elevation myocardial infarction (STEMI). The aim of this study was to assess the prognostic significance of CECs and indices of endothelial dysfunction in patients with STEMI. In 78 patients with acute STEMI, characterization of CD34+/VEGFR2+ CECs, and indices of endothelial damage/dysfunction such as brachial artery flow mediated dilatation (FMD) were determined. Blood samples for CECs assessment and quantification were obtained within 24 hours of admission and FMD was assessed during the index hospitalization. At 30 days follow up, the primary composite end point of major cardiac adverse events (MACE) consisting of all-cause mortality, recurrent non-fatal MI, or heart failure and the secondary endpoint of early adverse left ventricular (LV) remodeling were analyzed. The 17 patients (22%) who developed MACE had significantly higher CEC level (P = 0.004), vWF level (P =0.028), and significantly lower FMD (P = 0.006) compared to the remaining patients. Logistic regression analysis showed that CECs level and LV ejection fraction were independent predictors of MACE. The areas under the receiver operating characteristic curves (ROC) for CEC level, FMD, and the logistic model with both markers were 0.73, 0.75, and 0.82 respectively for prediction of the MACE. The 16 patients who developed the secondary endpoint had significantly higher CEC level compared to remaining patients (p =0.038). In conclusion, increased circulating endothelial cells and endothelial dysfunction predicted the occurrence of major adverse cardiac events and adverse cardiac remodeling in patients with STEMI. PMID:26864952

  16. Thrombomodulin as a marker for vascular tumors. Comparative study with factor VIII and Ulex europaeus I lectin.

    PubMed

    Yonezawa, S; Maruyama, I; Sakae, K; Igata, A; Majerus, P W; Sato, E

    1987-10-01

    Thrombomodulin (TM) is a newly described endothelial cell-associated protein that functions as a potent natural anticoagulant by converting thrombin from a procoagulant protease to an anticoagulant. Various vascular tumors were characterized with immunoperoxidase staining with the use of a polyclonal anti-TM serum. The staining patterns of TM were compared with those of Factor VIII-related antigen (FVIII-RAG) and Ulex europaeus agglutinin-I (UEA-I), which have been used as markers for endothelial cells. The results showed that TM is a specific and a highly sensitive marker for angiosarcomas in comparison with FVIII-RAG or UEA-I. In contrast, UEA-I is more sensitive for benign vascular tumors than TM or FVIII-RAG. The other mesenchymal tumors of nonvascular origin showed negative staining for three endothelial markers. These results indicate that TM is a new specific and sensitive tool for the diagnosis of angiosarcomas.

  17. Endotoxin Induces Fibrosis in Vascular Endothelial Cells through a Mechanism Dependent on Transient Receptor Protein Melastatin 7 Activity

    PubMed Central

    Echeverría, Cesar; Montorfano, Ignacio; Hermosilla, Tamara; Armisén, Ricardo; Velásquez, Luis A.; Cabello-Verrugio, Claudio; Varela, Diego; Simon, Felipe

    2014-01-01

    The pathogenesis of systemic inflammatory diseases, including endotoxemia-derived sepsis syndrome, is characterized by endothelial dysfunction. It has been demonstrated that the endotoxin lipopolysaccharide (LPS) induces the conversion of endothelial cells (ECs) into activated fibroblasts through endothelial­to­mesenchymal transition mechanism. Fibrogenesis is highly dependent on intracellular Ca2+ concentration increases through the participation of calcium channels. However, the specific molecular identity of the calcium channel that mediates the Ca2+ influx during endotoxin-induced endothelial fibrosis is still unknown. Transient receptor potential melastatin 7 (TRPM7) is a calcium channel that is expressed in many cell types, including ECs. TRPM7 is involved in a number of crucial processes such as the conversion of fibroblasts into activated fibroblasts, or myofibroblasts, being responsible for the development of several characteristics of them. However, the role of the TRPM7 ion channel in endotoxin-induced endothelial fibrosis is unknown. Thus, our aim was to study whether the TRPM7 calcium channel participates in endotoxin-induced endothelial fibrosis. Using primary cultures of ECs, we demonstrated that TRPM7 is a crucial protein involved in endotoxin-induced endothelial fibrosis. Suppression of TRPM7 expression protected ECs from the fibrogenic process stimulated by endotoxin. Downregulation of TRPM7 prevented the endotoxin-induced endothelial markers decrease and fibrotic genes increase in ECs. In addition, TRPM7 downregulation abolished the endotoxin-induced increase in ECM proteins in ECs. Furthermore, we showed that intracellular Ca2+ levels were greatly increased upon LPS challenge in a mechanism dependent on TRPM7 expression. These results demonstrate that TRPM7 is a key protein involved in the mechanism underlying endotoxin-induced endothelial fibrosis. PMID:24710004

  18. Endotoxin induces fibrosis in vascular endothelial cells through a mechanism dependent on transient receptor protein melastatin 7 activity.

    PubMed

    Echeverría, Cesar; Montorfano, Ignacio; Hermosilla, Tamara; Armisén, Ricardo; Velásquez, Luis A; Cabello-Verrugio, Claudio; Varela, Diego; Simon, Felipe

    2014-01-01

    The pathogenesis of systemic inflammatory diseases, including endotoxemia-derived sepsis syndrome, is characterized by endothelial dysfunction. It has been demonstrated that the endotoxin lipopolysaccharide (LPS) induces the conversion of endothelial cells (ECs) into activated fibroblasts through endothelial-to-mesenchymal transition mechanism. Fibrogenesis is highly dependent on intracellular Ca2+ concentration increases through the participation of calcium channels. However, the specific molecular identity of the calcium channel that mediates the Ca2+ influx during endotoxin-induced endothelial fibrosis is still unknown. Transient receptor potential melastatin 7 (TRPM7) is a calcium channel that is expressed in many cell types, including ECs. TRPM7 is involved in a number of crucial processes such as the conversion of fibroblasts into activated fibroblasts, or myofibroblasts, being responsible for the development of several characteristics of them. However, the role of the TRPM7 ion channel in endotoxin-induced endothelial fibrosis is unknown. Thus, our aim was to study whether the TRPM7 calcium channel participates in endotoxin-induced endothelial fibrosis. Using primary cultures of ECs, we demonstrated that TRPM7 is a crucial protein involved in endotoxin-induced endothelial fibrosis. Suppression of TRPM7 expression protected ECs from the fibrogenic process stimulated by endotoxin. Downregulation of TRPM7 prevented the endotoxin-induced endothelial markers decrease and fibrotic genes increase in ECs. In addition, TRPM7 downregulation abolished the endotoxin-induced increase in ECM proteins in ECs. Furthermore, we showed that intracellular Ca2+ levels were greatly increased upon LPS challenge in a mechanism dependent on TRPM7 expression. These results demonstrate that TRPM7 is a key protein involved in the mechanism underlying endotoxin-induced endothelial fibrosis.

  19. PPAR Gamma and Angiogenesis: Endothelial Cells Perspective

    PubMed Central

    2016-01-01

    We summarize the current knowledge concerning PPARγ function in angiogenesis. We discuss the mechanisms of action for PPARγ and its role in vasculature development and homeostasis, focusing on endothelial cells, endothelial progenitor cells, and bone marrow-derived proangiogenic cells. PMID:28053991

  20. The endothelial protein C receptor and malaria.

    PubMed

    van der Poll, Tom

    2013-08-01

    In this issue of Blood, Moxon et al provide novel insight into the pathogenesis of cerebral malaria, linking loss of the endothelial protein C receptor (EPCR) on brain vessels, caused by cytoadherent infected erythrocytes, with localized coagulation, inflammation, and disruption of endothelial barrier function.

  1. Vascular endothelial dysfunction and pharmacological treatment

    PubMed Central

    Su, Jin Bo

    2015-01-01

    The endothelium exerts multiple actions involving regulation of vascular permeability and tone, coagulation and fibrinolysis, inflammatory and immunological reactions and cell growth. Alterations of one or more such actions may cause vascular endothelial dysfunction. Different risk factors such as hypercholesterolemia, homocystinemia, hyperglycemia, hypertension, smoking, inflammation, and aging contribute to the development of endothelial dysfunction. Mechanisms underlying endothelial dysfunction are multiple, including impaired endothelium-derived vasodilators, enhanced endothelium-derived vasoconstrictors, over production of reactive oxygen species and reactive nitrogen species, activation of inflammatory and immune reactions, and imbalance of coagulation and fibrinolysis. Endothelial dysfunction occurs in many cardiovascular diseases, which involves different mechanisms, depending on specific risk factors affecting the disease. Among these mechanisms, a reduction in nitric oxide (NO) bioavailability plays a central role in the development of endothelial dysfunction because NO exerts diverse physiological actions, including vasodilation, anti-inflammation, antiplatelet, antiproliferation and antimigration. Experimental and clinical studies have demonstrated that a variety of currently used or investigational drugs, such as angiotensin-converting enzyme inhibitors, angiotensin AT1 receptors blockers, angiotensin-(1-7), antioxidants, beta-blockers, calcium channel blockers, endothelial NO synthase enhancers, phosphodiesterase 5 inhibitors, sphingosine-1-phosphate and statins, exert endothelial protective effects. Due to the difference in mechanisms of action, these drugs need to be used according to specific mechanisms underlying endothelial dysfunction of the disease. PMID:26635921

  2. Induction of vascular endothelial phenotype and cellular proliferation from human cord blood stem cells cultured in simulated microgravity

    NASA Astrophysics Data System (ADS)

    Chiu, Brian; Z-M Wan, Jim; Abley, Doris; Akabutu, John

    2005-05-01

    Recent studies have demonstrated that stem cells derived from adult hematopoietic tissues are capable of trans-differentiation into non-hematopoietic cells, and that the culture in microgravity ( μg) may modulate the proliferation and differentiation. We investigated the application of μg to human umbilical cord blood stem cells (CBSC) in the induction of vascular endothelial phenotype expression and cellular proliferation. CD34+ mononuclear cells were isolated from waste human umbilical cord blood samples and cultured in simulated μg for 14 days. The cells were seeded in rotary wall vessels (RWV) with or without microcarrier beads (MCB) and vascular endothelial growth factor was added during culture. Controls consisted of culture in 1 G. The cell cultures in RWV were examined by inverted microscopy. Cell counts, endothelial cell and leukocyte markers performed by flow-cytometry and FACS scan were assayed at days 1, 4, 7 and at the termination of the experiments. Culture in RWV revealed significantly increased cellular proliferation with three-dimensional (3D) tissue-like aggregates. At day 4, CD34+ cells cultured in RWV bioreactor without MCB developed vascular tubular assemblies and exhibited endothelial phenotypic markers. These data suggest that CD34+ human umbilical cord blood progenitors are capable of trans-differentiation into vascular endothelial cell phenotype and assemble into 3D tissue structures. Culture of CBSC in simulated μg may be potentially beneficial in the fields of stem cell biology and somatic cell therapy.

  3. CD14(+)CD16(++) "nonclassical" monocytes are associated with endothelial dysfunction in patients with coronary artery disease.

    PubMed

    Urbanski, Karol; Ludew, Dominik; Filip, Grzegorz; Filip, Magdalena; Sagan, Agnieszka; Szczepaniak, Piotr; Grudzien, Grzegorz; Sadowski, Jerzy; Jasiewicz-Honkisz, Barbara; Sliwa, Tomasz; Kapelak, Boguslaw; McGinnigle, Eilidh; Mikolajczyk, Tomasz; Guzik, Tomasz J

    2017-02-23

    Endothelial dysfunction and inflammation are key mechanisms of vascular disease. We hypothesised that heterogeneity of monocyte subpopulations may be related to the development of vascular dysfunction in coronary artery disease (CAD). Therefore, we examined the relationships between monocyte subsets (CD14(++)CD16(-) "classical - Mon1", CD14(++)CD16(+) "intermediate - Mon2" and CD14(+)CD16(++) "nonclassical - Mon3"), endothelial function and risk factor profiles in 130 patients with CAD undergoing coronary artery bypass grafting. This allowed for direct nitric oxide (NO) bioavailability assessment using isometric tension studies ex vivo (acetylcholine; ACh- and sodium-nitropruside; SNP-dependent) in segments of internal mammary arteries. The expression of CD14 and CD16 antigens and activation markers were determined in peripheral blood mononuclear cells using flow cytometry. Patients with high CD14(+)CD16(++) "nonclassical" and low CD14(++)CD16(-) "classical" monocytes presented impaired endothelial function. High frequency of CD14(+)CD16(++) "nonclassical" monocytes was associated with increased vascular superoxide production. Furthermore, endothelial dysfunction was associated with higher expression of activation marker CD11c selectively on CD14(+)CD16(++) monocytes. Nonclassical and classical monocyte frequencies remained independent predictors of endothelial dysfunction when major risk factors for atherosclerosis were taken into account (β=0.18 p=0.04 and β=-0.19 p=0.03, respectively). In summary, our data indicate that CD14(+)CD16(++) "nonclassical" monocytes are associated with more advanced vascular dysfunction measured as NO- bioavailability and vascular reactive oxygen species production.

  4. Endothelial cell micropatterning: Methods, effects, and applications

    PubMed Central

    Anderson, Deirdre E.J.; Hinds, Monica T.

    2012-01-01

    The effects of flow on endothelial cells have been widely examined for the ability of fluid shear stress to alter cell morphology and function; however, the effects of endothelial cell morphology without flow have only recently been observed. An increase in lithographic techniques in cell culture spurred a corresponding increase in research aiming to confine cell morphology. These studies lead to a better understanding of how morphology and cytoskeletal configuration affect the structure and function of the cells. This review examines endothelial cell micropatterning research by exploring both the many alternative methods used to alter endothelial cell morphology and the resulting changes in cellular shape and phenotype. Micropatterning induced changes in endothelial cell proliferation, apoptosis, cytoskeletal organization, mechanical properties, and cell functionality. Finally, the ways these cellular manipulation techniques have been applied to biomedical engineering research, including angiogenesis, cell migration, and tissue engineering, is discussed. PMID:21761242

  5. Sphingosine-1-Phosphate Signaling in Endothelial Disorders.

    PubMed

    Sanchez, Teresa

    2016-06-01

    Numerous preclinical studies indicate that sustained endothelial activation significantly contributes to tissue edema, perpetuates the inflammatory response, and exacerbates tissue injury ultimately resulting in organ failure. However, no specific therapies aimed at restoring endothelial function are available as yet. Sphingosine-1-phosphate (S1P) is emerging as a potent modulator of endothelial function and endothelial responses to injury. Recent studies indicate that S1PR are attractive targets to treat not only disorders of the arterial endothelium but also microvascular dysfunction caused by ischemic or inflammatory injury. In this article, we will review the current knowledge of the role of S1P and its receptors in endothelial function in health and disease, and we will discuss the therapeutic potential of targeting S1PR not only for disorders of the arterial endothelium but also the microvasculature. The therapeutic targeting of S1PR in the endothelium could help to bridge the gap between biomedical research in vascular biology and clinical practice.

  6. Endothelial Dysfunction in Renal Failure: Current Update.

    PubMed

    Radenkovic, Miroslav; Stojanovic, Marko; Prostran, Milica

    2016-01-01

    Endothelial dysfunction is principally characterized by impaired endothelium- dependent transduction mechanisms related to vascular relaxation, as an outcome of decreased release of endothelium-derived relaxing factors, mainly nitric oxide, as well as augmented oxidative stress, increased inflammation and predominance of vascular action produced by endothelium-derived contracting factors. Current data strongly suggest that pathological development of different types of kidney impairment with further progression to renal failure includes notable vascular changes associated with endothelial dysfunction. In accordance, this scientific field represents an advancing area of investigation, involving different biomarkers of endothelial dysfunction linked to renal impairment, as well as clinical findings with new information that can provide a more comprehensive understanding of the role of endothelial dysfunction in kidney disease. With regards to quoted facts, the aim of this article was to review the latest data related to endothelial dysfunction and renal failure by selection of relevant articles released from 2010 to 2015.

  7. Endothelial Response to Glucocorticoids in Inflammatory Diseases

    PubMed Central

    Zielińska, Karolina A.; Van Moortel, Laura; Opdenakker, Ghislain; De Bosscher, Karolien; Van den Steen, Philippe E.

    2016-01-01

    The endothelium plays a crucial role in inflammation. A balanced control of inflammation requires the action of glucocorticoids (GCs), steroidal hormones with potent cell-specific anti-inflammatory properties. Besides the classic anti-inflammatory effects of GCs on leukocytes, recent studies confirm that endothelial cells also represent an important target for GCs. GCs regulate different aspects of endothelial physiology including expression of adhesion molecules, production of pro-inflammatory cytokines and chemokines, and maintenance of endothelial barrier integrity. However, the regulation of endothelial GC sensitivity remains incompletely understood. In this review, we specifically examine the endothelial response to GCs in various inflammatory diseases ranging from multiple sclerosis, stroke, sepsis, and vasculitis to atherosclerosis. Shedding more light on the cross talk between GCs and endothelium will help to improve existing therapeutic strategies and develop new therapies better tailored to the needs of patients. PMID:28018358

  8. Cinacalcet ameliorates aortic calcification in uremic rats via suppression of endothelial-to-mesenchymal transition

    PubMed Central

    wu, Min; Tang, Ri-ning; Liu, Hong; Pan, Ming-ming; Liu, Bi-cheng

    2016-01-01

    Aim: Experimental studies found that cinacalcet (CINA) markedly attenuated vascular calcification in uremic rats, but its underlying mechanisms are still largely unknown. Recent evidence have demonstrated that endothelial cells (ECs) participate in ectopic calcification in part by mediating endothelial-to-mesenchymal transition (EndMT). In this study, we investigated whether CINA ameliorated aortic calcification in uremic rats via suppression of EndMT. Methods: Uremia was induced in rats by feeding rats a 0.75% adenine diet for 4 weeks. After adenine withdrawal, the rats were maintained on a 1.03% phosphorus diet for next 8 weeks. At initiation of the adenine diet, rats were orally administered CINA (10mg/kg one day) for 12 weeks. The aortic expression of EndMT- and chondrocyte- markers was examined. The effect of elevated PTH on EndMT was also studied in aortic ECs. Results: In uremic rats, CINA treatment significantly decreased the serum PTH concentrations, but did not affect the elevated levels of serum calcium (Ca), phosphorus (P) and Ca×P product. Besides, CINA significantly attenuated aortic calcification, and inhibited the expression of chondrocyte markers (SOX9 and COL2A1) and chondrocyte proteoglycan in uremic aortas. Moreover, CINA treatment largely abolished the up-regulation of mesenchymal markers (FSP1 and α-SMA) and down-regulation of the endothelial marker (CD31), which accompanied aortic calcification in uremic aorta samples. In vitro, PTH increased the expression of EndMT-markers in a concentration- and time-dependent manner. Conclusion: These findings suggest that strategies aiming at reducing serum PTH might prevent uremic aortic calcification by abrogating EndMT. PMID:27593220

  9. Epigenetic mechanisms of endothelial dysfunction in type 2 diabetes.

    PubMed

    Prattichizzo, Francesco; Giuliani, Angelica; Ceka, Artan; Rippo, Maria Rita; Bonfigli, Anna Rita; Testa, Roberto; Procopio, Antonio Domenico; Olivieri, Fabiola

    2015-01-01

    The development of type-2 diabetes mellitus (T2DM) and its complications is largely due to the complex interaction between genetic factors and environmental influences, mainly dietary habits and lifestyle, which can either accelerate or slow down disease progression. Recent findings suggest the potential involvement of epigenetic mechanisms as a crucial interface between the effects of genetic predisposition and environmental factors. The common denominator of environmental factors promoting T2DM development and progression is that they trigger an inflammatory response, promoting inflammation-mediated insulin resistance and endothelial dysfunction. Proinflammatory stimuli, including hyperglycemia, oxidative stress, and other inflammatory mediators, can affect epigenetic mechanisms, altering the expression of specific genes in target cells without changes in underlying DNA sequences. DNA methylation and post-translational histone modifications (PTHMs) are the most extensively investigated epigenetic mechanisms. Over the past few years, non-coding RNA, including microRNAs (miRNAs), have also emerged as key players in gene expression modulation. MiRNAs can be actively released or shed by cells in the bloodstream and taken up in active form by receiving cells, acting as efficient systemic communication tools. The miRNAs involved in modulation of inflammatory pathways (inflammamiRs), such as miR-146a, and those highly expressed in endothelial lineages and hematopoietic progenitor cells (angiomiRs), such as miR-126, are the most extensively studied circulating miRNAs in T2DM. However, data on circulating miRNA signatures associated with specific diabetic complications are still lacking. Since immune cells and endothelial cells are primarily involved in the vascular complications of T2DM, their relative contribution to circulating miRNA signatures needs to be elucidated. An integrated approach encompassing different epigenetic mechanisms would have the potential to

  10. Vascular Endothelial Growth Factor Receptor -2 in Breast Cancer

    PubMed Central

    Guo, Shanchun; Colbert, Laronna S.; Fuller, Miles; Zhang, Yuanyuan; Gonzalez-Perez, Ruben R.

    2010-01-01

    Investigations over the last decade have established the essential role of growth factors and their receptors during angiogenesis and carcinogenesis. The vascular endothelial growth factor receptor (VEGFR) family in mammals contains three members, VEGFR-1 (Flt-1), VEGFR-2 (KDR/Flk-1) and VEGFR-3 (Flt-4), which are transmembrane tyrosine kinase receptors that regulate the formation of blood and lymphatic vessels. In the early 1990s, the above VEGFR were structurally characterized by cDNA cloning. Among these three receptors, VEGFR-2 is generally recognized to have a principal role in mediating VEGF-induced responses. VEGFR-2 is considered as the earliest marker for endothelial cell development. Importantly, VEGFR-2 directly regulates tumor angiogenesis. Therefore, several inhibitors of VEGFR-2 have been developed and many of them are now in clinical trials. In addition to targeting endothelial cells, the VEGF/VEGFR-2 system works as an essential autocrine/paracrine process for cancer cell proliferation and survival. Recent studies mark the continuous and increased interest in this related, but distinct, function of VEGF/VEGFR-2 in cancer cells: the autocrine/paracrine loop. Several mechanisms regulate VEGFR-2 levels and modulate its role in tumor angiogenesis and physiologic functions, i.e.: cellular localization/trafficking, regulation of cis-elements of promoter, epigenetic regulation and signaling from Notch, cytokines/growth factors and estrogen, etc. In this review, we will focus on updated information regarding VEGFR-2 research with respect to the molecular mechanisms of VEGFR-2 regulation in human breast cancer. Investigations in the activation, function, and regulation of VEGFR-2 in breast cancer will allow the development of new pharmacological strategies aimed at directly targeting cancer cell proliferation and survival. PMID:20462514

  11. Fiducial Marker Placement

    MedlinePlus

    ... Media Computed Tomography (CT) - Body General Ultrasound Ultrasound - Prostate Introduction to Cancer Therapy (Radiation Oncology) Proton Therapy Stereotactic Radiosurgery (SRS) and Stereotactic Body Radiotherapy (SBRT) Images related to Fiducial Marker Placement Sponsored by ...

  12. Alcoholism: Current Marker Research

    DTIC Science & Technology

    1984-03-01

    genetically determined characteristics such as color blindness and blood type . GENETIC MARKER STUDIES In 1966 Dr. Cruz-Coke and Dr. Varela reported that...and recovery from severe alcoholism symptoms. ■󈧒:584-587) Blood - typing marker studies have produced similar mixed results. One study published in...1959 showed a high correlation among 939 alcoholics and blood type A. (20:4 60-4 61) A similar study in 1973 reported no blood type distribution

  13. [Biological markers of alcoholism].

    PubMed

    Marcos Martín, M; Pastor Encinas, I; Laso Guzmán, F J

    2005-09-01

    Diagnosis of alcoholism is very important, given its high prevalence and possibility of influencing the disease course. For this reason, the so-called biological markers of alcoholism are useful. These are analytic parameters that alter in the presence of excessive alcohol consumption. The two most relevant markers are the gamma-glutamyltranspeptidase and carbohydrate deficient transferrin. With this clinical comment, we aim to contribute to the knowledge of these tests and promote its use in the clinical practice.

  14. Colon cancer: cancer stem cells markers, drug resistance and treatment.

    PubMed

    Kozovska, Zuzana; Gabrisova, Veronika; Kucerova, Lucia

    2014-10-01

    Malignant tumours consist of heterogeneous populations of tumour cells. Cancer stem cells (CSC) represent a population of cells within a tumour with highly tumorigenic and chemoresistant properties. These cells may be identified by the expression of CSC markers. There are several key stem cells markers specified for colon cancer: CD133, CD44, ALDH1, ALCAM. These days, a major obstacle to effective cancer management is development of a multidrug resistance (MDR). The principal mechanism responsible for development of MDR phenotype is the over-expression of ABC transporters. Tumours and relapsing tumours after therapy are drived by subpopulations of tumour cells with aggressive phenotype resistant to chemotherapeutics. These cells are called CSC or tumour-initiating cells (TIC). Here we outline recent information about MDR of colon cancer and CSC markers. We have focused on novel therapeutic strategies which have been developed to prevent or overcome MDR. One such strategy is a combination of chemotherapy and modulators of MDR pumps or chemotherapy and monoclonal antibodies against vascular endothelial growth factor VEGF. Colon cancer is characterized by the presence of colon CSC expressing specific stem cell markers. The divergent presence of these markers can help to adjust personalized therapy. The review provides a detailed overview of resistance of colon cancer cells and discusses how the presence of CSC markers can influence therapy and prognosis of patients.

  15. Relationship of oxidative stress and endothelial dysfunction in sleep apnoea.

    PubMed

    Jurado-Gámez, B; Fernandez-Marin, M C; Gómez-Chaparro, J L; Muñoz-Cabrera, L; Lopez-Barea, J; Perez-Jimenez, F; Lopez-Miranda, J

    2011-04-01

    The aim of the present study was to evaluate ischaemic reactive hyperaemia (IRH) in obstructive sleep apnoea (OSA) and its relationship with oxidative stress. We studied 69 consecutive patients referred to our Sleep Unit (Reina Sofia University Hospital, Cordoba, Spain). Patients with chronic diseases or those taking medication were excluded. IRH was assessed before and after polysomnography. Morning IRH and oxidative stress markers were compared between patients with (apnoea-hypopnoea index (AHI) ≥ 5) and without (AHI < 5) OSA. Measurements were repeated in 25 severe OSA patients after continuous positive airway pressure (CPAP) therapy. We included 46 OSA patients (mean ± sd AHI 49 ± 32.1) and 23 non-OSA subjects (AHI 3 ± 0.9). The OSA patients showed a significant worsening of morning IRH, and a significant increase in malondialdehyde and 8-hydroxydeoxyguanosine levels. Only the oxygen desaturation index independently explained morning IRH, while malondialdehyde levels showed a weak effect on IRH. In severe OSA patients, IRH improved significantly after CPAP treatment, as did malondialdehyde, 8-hydroxydeoxyguanosine and protein carbonyl levels. In OSA patients, endothelial dysfunction and oxidative stress were observed, and IRH worsened after sleep. The increase in oxidative stress was not associated with IRH, while intermittent hypoxia was strongly associated with IRH. In severe OSA patients, CPAP treatment improved oxidative stress and endothelial function.

  16. Hydrogen prevents corneal endothelial damage in phacoemulsification cataract surgery

    PubMed Central

    Igarashi, Tsutomu; Ohsawa, Ikuroh; Kobayashi, Maika; Igarashi, Toru; Suzuki, Hisaharu; Iketani, Masumi; Takahashi, Hiroshi

    2016-01-01

    In phacoemulsification, ultrasound induces hydroxyl radical (·OH) formation, damaging corneal endothelium. Whether H2 can prevent such oxidative damage in phacoemulsification was examined by in vitro and in vivo studies. H2 was dissolved in a commercial irrigating solution. The effects of H2 against ·OH generation were first confirmed in vitro by electron-spin resonance (ESR) and hydroxyphenyl fluorescein (HPF). ESR showed a significantly decreased signal magnitude, and fluorescence intensity by oxidized HPF was significantly less in the H2-dissolved solution. The effects of H2 in phacoemulsification were evaluated in rabbits, comparing H2-dissolved and control solutions. Five hours after the procedure, the whole cornea was excised and subjected to image analysis for corneal edema, real-time semiquantitative PCR (qPCR) for heme oxygenase (HO)-1, catalase (CAT), superoxide dismutase 1 (SOD1), and SOD2 mRNA, and immunohistochemistry. Corneal edema was significantly less and the increases in anti-oxidative HO-1, CAT and SOD2 mRNA expressions were significantly suppressed in the H2 group. In addition, corneal endothelial cell expressions of two oxidative stress markers, 4-HNE and 8-OHdG, were significantly lower in the H2 group. In conclusion, H2 dissolved in the ocular irrigating solution protected corneal endothelial cells from phacoemulsification-induced oxidative stress and damage. PMID:27498755

  17. Assessment of endothelial and neurovascular function in human skin microcirculation.

    PubMed

    Roustit, Matthieu; Cracowski, Jean-Luc

    2013-07-01

    Peripheral microvascular dysfunction has been described in many physiological and pathological conditions. Owing to its accessibility, the cutaneous microcirculation provides a unique index of microvascular function. Skin microvascular function has therefore been proposed as a prognostic marker or for evaluating the effect of drugs on the microcirculation. Various reactivity tests, coupled with techniques measuring skin blood flux, are used to non-invasively explore both endothelial and neurovascular microvascular functioning in humans. We review the advantages and limitations of the main reactivity tests, including post-occlusive reactive hyperemia, local thermal hyperemia, pressure-induced vasodilation, and iontophoresis of vasodilators, combined with measurement techniques such as laser Doppler and laser speckle contrast imaging. Recent advances in our comprehension of the physiological pathways underlying these reactivity tests, as well as technological developments in microcirculation imaging, have provided reliable and reproducible tools for studying the microcirculation.

  18. Microfluidically supported biochip design for culture of endothelial cell layers with improved perfusion conditions.

    PubMed

    Raasch, Martin; Rennert, Knut; Jahn, Tobias; Peters, Sven; Henkel, Thomas; Huber, Otmar; Schulz, Ingo; Becker, Holger; Lorkowski, Stefan; Funke, Harald; Mosig, Alexander

    2015-03-02

    Hemodynamic forces generated by the blood flow are of central importance for the function of endothelial cells (ECs), which form a biologically active cellular monolayer in blood vessels and serve as a selective barrier for macromolecular permeability. Mechanical stimulation of the endothelial monolayer induces morphological remodeling in its cytoskeleton. For in vitro studies on EC biology culture devices are desirable that simulate conditions of flow in blood vessels and allow flow-based adhesion/permeability assays under optimal perfusion conditions. With this aim we designed a biochip comprising a perfusable membrane that serves as cell culture platform multi-organ-tissue-flow (MOTiF biochip). This biochip allows an effective supply with nutrition medium, discharge of catabolic cell metabolites and defined application of shear stress to ECs under laminar flow conditions. To characterize EC layers cultured in the MOTiF biochip we investigated cell viability, expression of EC marker proteins and cell adhesion molecules of ECs dynamically cultured under low and high shear stress, and compared them with an endothelial culture in established two-dimensionally perfused flow chambers and under static conditions. We show that ECs cultured in the MOTiF biochip form a tight EC monolayer with increased cellular density, enhanced cell layer thickness, presumably as the result of a rapid and effective adaption to shear stress by remodeling of the cytoskeleton. Moreover, endothelial layers in the MOTiF biochip express higher amounts of EC marker proteins von-Willebrand-factor and PECAM-1. EC layers were highly responsive to stimulation with TNFα as detected at the level of ICAM-1, VCAM-1 and E-selectin expression and modulation of endothelial permeability in response to TNFα/IFNγ treatment under flow conditions. Compared to static and two-dimensionally perfused cell culture condition we consider MOTiF biochips as a valuable tool for studying EC biology in vitro under

  19. Oral cancer/endothelial cell fusion experiences nuclear fusion and acquisition of enhanced survival potential

    SciTech Connect

    Song, Kai; Song, Yong; Zhao, Xiao-Ping; Shen, Hui; Wang, Meng; Yan, Ting-lin; Liu, Ke; Shang, Zheng-jun

    2014-10-15

    Most previous studies have linked cancer–macrophage fusion with tumor progression and metastasis. However, the characteristics of hybrid cells derived from oral cancer and endothelial cells and their involvement in cancer remained unknown. Double-immunofluorescent staining and fluorescent in situ hybridization (FISH) were performed to confirm spontaneous cell fusion between eGFP-labeled human umbilical vein endothelial cells (HUVECs) and RFP-labeled SCC9, and to detect the expression of vementin and cytokeratin 18 in the hybrids. The property of chemo-resistance of such hybrids was examined by TUNEL assay. The hybrid cells in xenografted tumor were identified by FISH and GFP/RFP dual-immunofluoresence staining. We showed that SCC9 cells spontaneously fused with cocultured endothelial cells, and the resultant hybrid cells maintained the division and proliferation activity after re-plating and thawing. Such hybrids expressed markers of both parental cells and became more resistant to chemotherapeutic drug cisplatin as compared to the parental SCC9 cells. Our in vivo data indicated that the hybrid cells contributed to tumor composition by using of immunostaining and FISH analysis, even though the hybrid cells and SCC9 cells were mixed with 1:10,000, according to the FACS data. Our study suggested that the fusion events between oral cancer and endothelial cells undergo nuclear fusion and acquire a new property of drug resistance and consequently enhanced survival potential. These experimental findings provide further supportive evidence for the theory that cell fusion is involved in cancer progression. - Highlights: • The fusion events between oral cancer and endothelial cells undergo nuclear fusion. • The resulting hybrid cells acquire a new property of drug resistance. • The resulting hybrid cells express the markers of both parental cells (i.e. vimentin and cytokeratin 18). • The hybrid cells contribute to tumor repopulation in vivo.

  20. Vascular risk factors, endothelial function, and carotid thickness in patients with migraine: relationship to atherosclerosis.

    PubMed

    Hamed, Sherifa A; Hamed, Enas A; Ezz Eldin, Azza M; Mahmoud, Nagia M

    2010-03-01

    Recent studies indicated that migraine is associated with specific vascular risk profile. However, the functional and structural vascular abnormalities in migraine are rarely addressed. We evaluated the vascular risk factors, endothelial function, and carotid artery (CA)-intima-media thickness (IMT), segregators of preclinical atherosclerosis, in migraineurs. This preliminary study included 63 adults with headache (migraine with aura [n=14], migraine without aura [n=24], transformed migraine [n=6], and tension headache [n=19]) and 35 matched healthy subjects. The following vascular risks were assessed: body mass index (BMI), systolic blood pressure (SBP) and diastolic blood pressures (DBP), serum levels of C-reactive protein, fasting glucose, fasting insulin, total cholesterol, and triglycerides. Plasma endothelin (ET)-1, a vasoactive peptide produced by vascular smooth muscle cells and marker for endothelial injury and atherosclerosis, was measured. Endothelial-dependent vasoreactivity was assessed using brachial artery flow-mediated dilatation (FMD) in response to hyperemia. CA-IMT, structural marker of early atherosclerosis, was measured. Compared with control subjects, SBP, DBP, glucose, insulin, ET-1, and CA-IMT were elevated with migraine. FMD% was inversely correlated with SBP (P < .001), DBP (P < .01), glucose (P < .001), and insulin levels (P < .01). CA-IMT was correlated with BMI (P < .05), SBP (P < .01), total cholesterol (P < .01), triglycerides (P < .001), glucose (P < .001), insulin (P < .01), and FMD% (P < .05). In multivariate analysis, ET-1 was correlated with duration of illness, SBP, DBP, glucose, insulin, IMT, and FMD%. We conclude that endothelial injury, impaired endothelial vasoreactivity, and increased CA-IMT occur with migraine and are associated with vascular risk factors that strongly suggest that migraine could be a risk for atherosclerosis.

  1. Flow cytometric analysis of circulating endothelial cells and endothelial progenitors for clinical purposes in oncology: A critical evaluation

    PubMed Central

    DANOVA, MARCO; COMOLLI, GIUDITTA; MANZONI, MARIANGELA; TORCHIO, MARTINA; MAZZINI, GIULIANO

    2016-01-01

    Malignant tumors are characterized by uncontrolled cell growth and metastatic spread, with a pivotal importance of the phenomenon of angiogenesis. For this reason, research has focused on the development of agents targeting the vascular component of the tumor microenvironment and regulating the angiogenic switch. As a result, the therapeutic inhibition of angiogenesis has become an important component of anticancer treatment, however, its utility is partly limited by the lack of an established methodology to assess its efficacy in vivo. Circulating endothelial cells (CECs), which are rare in healthy subjects and significantly increased in different tumor types, represent a promising tool for monitoring the tumor clinical outcome and the treatment response. A cell population circulating into the blood also able to form endothelial colonies in vitro and to promote vasculogenesis is represented by endothelial progenitor cells (EPCs). The number of both of these cell types is extremely low and they cannot be identified using a single marker, therefore, in absence of a definite consensus on their phenotype, require discrimination using combinations of antigens. Multiparameter flow cytometry (FCM) is ideal for rapid processing of high numbers of cells per second and is commonly utilized to quantify CECs and EPCs, however, remains technically challenging since there is as yet no standardized protocol for the identification and enumeration of these rare events. Methodology in studies on CECs and/or EPCs as clinical biomarkers in oncology is heterogeneous and data have been obtained from different studies leading to conflicting conclusions. The present review presented a critical review of the issues that limit the comparability of results of the most significant studies employing FCM for CEC and/or EPC detection in patients with cancer. PMID:27284422

  2. Leptin promotes endothelial dysfunction in chronic kidney disease through AKT/GSK3β and β-catenin signals.

    PubMed

    Ding, Nannan; Liu, Bing; Song, Jiaguang; Bao, Shougang; Zhen, Junhui; Lv, Zhimei; Wang, Rong

    2016-11-25

    Endothelial dysfunction (ED) is a well-recognized instigator of cardiovascular diseases and develops in chronic kidney disease (CKD) with high rate. Recent studies have implicated that leptin is associated with endothelial dysfunction. We investigated the relationship between leptin and markers of ED in CKD patients and how leptin contributed to endothelial damage. 140 CKD patients and 140 healthy subjects were studied. Serum leptin levels were significantly higher in CKD than in controls and displayed significantly positive association with the increase levels of sICAM-1 and sVCAM-1 but negative correlation with flow-mediated dilatation (FMD) reduction in patients. Our in vitro study demonstrated that leptin induced overexpression of ICAM-1 and VCAM-1, led to f-actin reorganization and vinculin assembly, increased endothelial monolayer permeability for FITC-dextran, and accelerated endothelial cell migration; these changes were markedly reversed when the cells were transfected with AKT or β-catenin shRNA vectors. Notably, high leptin resulted in hyper-phosphorylation of AKT and GSK3β, along with nuclear accumulation of β-catenin. In conclusion, serum leptin was elevated in CKD patients and it might contribute to endothelial dysfunction by disarrangement of f-actin cytoskeleton via a mechanism involving the AKT/GSK3β and β-catenin pathway.

  3. Establishment of a translational endothelial cell model using directed differentiation of induced pluripotent stem cells from Cynomolgus monkey

    PubMed Central

    Thoma, Eva C.; Heckel, Tobias; Keller, David; Giroud, Nicolas; Leonard, Brian; Christensen, Klaus; Roth, Adrian; Bertinetti-Lapatki, Cristina; Graf, Martin; Patsch, Christoph

    2016-01-01

    Due to their broad differentiation potential, pluripotent stem cells (PSCs) offer a promising approach for generating relevant cellular models for various applications. While human PSC-based cellular models are already advanced, similar systems for non-human primates (NHPs) are still lacking. However, as NHPs are the most appropriate animals for evaluating the safety of many novel pharmaceuticals, the availability of in vitro systems would be extremely useful to bridge the gap between cellular and animal models. Here, we present a NHP in vitro endothelial cell system using induced pluripotent stem cells (IPSCs) from Cynomolgus monkey (Macaca fascicularis). Based on an adapted protocol for human IPSCs, we directly differentiated macaque IPSCs into endothelial cells under chemically defined conditions. The resulting endothelial cells can be enriched using immuno-magnetic cell sorting and display endothelial marker expression and function. RNA sequencing revealed that the differentiation process closely resembled vasculogenesis. Moreover, we showed that endothelial cells derived from macaque and human IPSCs are highly similar with respect to gene expression patterns and key endothelial functions, such as inflammatory responses. These data demonstrate the power of IPSC differentiation technology to generate defined cell types for use as translational in vitro models to compare cell type-specific responses across species. PMID:27779219

  4. Glycation of paraoxonase 1 by high glucose instigates endoplasmic reticulum stress to induce endothelial dysfunction in vivo

    PubMed Central

    Yu, Wei; Liu, Xiaoli; Feng, Liru; Yang, Hui; Yu, Weiye; Feng, Tiejian; Wang, Shuangxi; Wang, Jun; Liu, Ning

    2017-01-01

    High-density lipoprotein (HDL) modulates low-density lipoprotein and cell membrane oxidation through the action of paraoxonase-1 (PON1). Endoplasmic reticulum (ER) stress has been linked to a wide range of human pathologies including diabetes, obesity, and atherosclerosis. Previous studies have reported that PON1 is glycated in diabetes. The aim of this study is to investigate whether and how PON1 glycation contributes to endothelial dysfunction in diabetes. ER stress markers were monitored by western blot. Endothelial function was determined by organ bath. Incubation of recombinant PON1 proteins with high glucose increased PON1 glycation and reduced PON1 activity. Exposure of HUVECs to glycated PON1 induced prolonged ER stress and reduced SERCA activity, which were abolished by tempol, apocynin, BAPTA, and p67 and p22 siRNAs. Chronic administration of amino guanidine or 4-PBA prevented endothelial dysfunction in STZ-injected rats. Importantly, injection of glycated PON1 but not native PON1 induced aberrant ER stress and endothelial dysfunction in rats, which were attenuated by tempol, BAPTA, and 4-PBA. In conclusion, glycation of PON1 by hyperglycemia induces endothelial dysfunction through ER stress. In perspectives, PON1 glycation is a novel risk factor of hyperglycemia-induced endothelial dysfunction. Therefore, inhibition of oxidative stress, chelating intracellular Ca2+, and ER chaperone would be considered to reduce vascular complications in diabetes. PMID:28374834

  5. Neither Proteinuria Nor Albuminuria Is Associated With Endothelial Dysfunction in HIV-Infected Patients Without Diabetes or Hypertension

    PubMed Central

    Shen, Changyu; Mather, Kieren J.; Agarwal, Rajiv; Dubé, Michael P.

    2011-01-01

    It is unknown whether systemic endothelial dysfunction underlies the association between nephropathy and cardiovascular disease (CVD) in persons infected with human immunodeficiency virus (HIV). Spot urine protein to creatinine ratio, spot urine albumin to creatinine ratio, creatinine clearance, estimated glomerular filtration rate, and flow-mediated dilation (FMD) of the brachial artery were evaluated in 123 study participants infected with HIV (58 receiving antiretroviral therapy [ART] and 65 not receiving ART) with no history of diabetes or hypertension. None of the renal markers, modeled as either continuous or categorical variables, correlated with FMD. Contrary to expectations, endothelial dysfunction may not be the link between nephropathy and CVD in HIV. PMID:22013226

  6. Possible vasculoprotective role of linagliptin against sodium arsenite-induced vascular endothelial dysfunction.

    PubMed

    Jyoti, Uma; Kansal, Sunil Kumar; Kumar, Puneet; Goyal, Sandeep

    2016-02-01

    Vascular endothelial dysfunction (VED) interrupts the integrity and function of endothelial lining through enhanced markers of oxidative stress and decrease endothelial nitric oxide synthase (eNOS) expression. The main aim of the present study has been designed to investigate the possible vasculoprotective role of linagliptin against sodium arsenite-induced VED. Sodium arsenite (1.5 mg/kg, i.p., 2 weeks) abrogated the acetylcholine-induced, endothelium-dependent vasorelaxation by depicting the decrease in serum nitrite/nitrate concentration, reduced glutathione level, and simultaneously enhance the thiobarbituric acid reactive substances (TBARS) level, superoxide level, and tumor necrosis factor-alpha. These elevated markers interrupt the integrity of endothelial lining of thoracic aorta which was assessed histologically. The study elicits dose dependent effect of linagliptin (1.5 mg/kg, i.p. and 3 mg/kg, i.p.) or atorvastatin (30 mg/kg, p.o.) treatment, improved the endothelium-dependent independent relaxation, improve the integrity of endothelium lining which was assessed histologically by enhancing the serum nitrite/nitrate level, reduced glutathione level and simultaneously decreasing the TBARS level, superoxide anion level and tumor necrosis factor-alpha (TNF-α) level. L-NAME (25 mg/kg, i.p.), eNOS inhibitor, abrogated the ameliorative potential of linagliptin. However, the ameliorative potential of linagliptin has been enhanced by l-arginine (200 mg/kg, i.p.) which elicits that ameliorative potential of linagliptin was through eNOS signaling cascade and it may be concluded that linagliptin 3 mg/kg, i.p. has more significantly activated the eNOS and decreased the oxidative markers than linagliptin 1.5 mg/kg, i.p. and prevented sodium arsenite-induced VED.

  7. Early systemic sclerosis: marker autoantibodies and videocapillaroscopy patterns are each associated with distinct clinical, functional and cellular activation markers

    PubMed Central

    2013-01-01

    Introduction Early systemic sclerosis (SSc) is characterized by Raynaud's phenomenon together with scleroderma marker autoantibodies and/or a scleroderma pattern at capillaroscopy and no other distinctive feature of SSc. Patients presenting with marker autoantibodies plus a capillaroscopic scleroderma pattern seem to evolve into definite SSc more frequently than patients with either feature. Whether early SSc patients with only marker autoantibodies or capillaroscopic positivity differ in any aspect at presentation is unclear. Methods Seventy-one consecutive early SSc patients were investigated for preclinical cardiopulmonary alterations. Out of these, 44 patients and 25 controls affected by osteoarthritis or primary fibromyalgia syndrome were also investigated for serum markers of fibroblast (carboxyterminal propeptide of collagen I), endothelial (soluble E-selectin) and T-cell (soluble IL-2 receptor alpha) activation. Results Thirty-two of the 71 patients (45.1%) had both a marker autoantibody and a capillaroscopic scleroderma pattern (subset 1), 16 patients (22.5%) had only a marker autoantibody (subset 2), and 23 patients (32.4%) had only a capillaroscopic scleroderma pattern (subset 3). Patients with marker autoantibodies (n = 48, 67.6%) had a higher prevalence of impaired diffusing lung capacity for carbon monoxide (P = 0.0217) and increased serum levels of carboxyterminal propeptide of collagen I (P = 0.0037), regardless of capillaroscopic alterations. Patients with a capillaroscopic scleroderma pattern (n = 55, 77.5%) had a higher prevalence of puffy fingers (P = 0.0001) and increased serum levels of soluble E-selectin (P = 0.0003) regardless of marker autoantibodies. Conclusion These results suggest that the autoantibody and microvascular patterns in early SSc may each be related to different clinical-preclinical features and circulating activation markers at presentation. Longitudinal studies are warranted to investigate whether these subsets undergo a

  8. Blood cells and endothelial barrier function

    PubMed Central

    Rodrigues, Stephen F; Granger, D Neil

    2015-01-01

    Abstract The barrier properties of endothelial cells are critical for the maintenance of water and protein balance between the intravascular and extravascular compartments. An impairment of endothelial barrier function has been implicated in the genesis and/or progression of a variety of pathological conditions, including pulmonary edema, ischemic stroke, neurodegenerative disorders, angioedema, sepsis and cancer. The altered barrier function in these conditions is often linked to the release of soluble mediators from resident cells (e.g., mast cells, macrophages) and/or recruited blood cells. The interaction of the mediators with receptors expressed on the surface of endothelial cells diminishes barrier function either by altering the expression of adhesive proteins in the inter-endothelial junctions, by altering the organization of the cytoskeleton, or both. Reactive oxygen species (ROS), proteolytic enzymes (e.g., matrix metalloproteinase, elastase), oncostatin M, and VEGF are part of a long list of mediators that have been implicated in endothelial barrier failure. In this review, we address the role of blood borne cells, including, neutrophils, lymphocytes, monocytes, and platelets, in the regulation of endothelial barrier function in health and disease. Attention is also devoted to new targets for therapeutic intervention in disease states with morbidity and mortality related to endothelial barrier dysfunction. PMID:25838983

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

    PubMed

    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.

  10. Usefulness of retinal microvascular endothelial dysfunction as a predictor of coronary artery disease.

    PubMed

    Al-Fiadh, Ali H; Wong, Tien Y; Kawasaki, Ryo; Clark, David J; Patel, Sheila K; Freeman, Melanie; Wilson, Andrew; Burrell, Louise M; Farouque, Omar

    2015-03-01

    Endothelial dysfunction is a key feature of atherosclerosis. Retinal microvascular endothelial function can be assessed using noninvasive dynamic vessel imaging techniques. Whether it is impaired in subjects with coronary artery disease (CAD) is unknown. The aim of this study was to examine the relation of retinal microvascular endothelial function with CAD. Vascular studies were performed in 197 prospectively recruited subjects divided into 2 groups: those without CAD but ≥2 cardiovascular risk factors (non-CAD controls; n = 119) and those with stable CAD (n = 78). Retinal microvascular endothelial dysfunction was assessed by measuring retinal arteriolar and venular dilatation to flicker light, a nitric oxide-dependent phenomenon, expressed as percentage increase over baseline diameter. Fingertip pulse-volume amplitude was measured to calculate reactive hyperaemia index and brachial artery flow-mediated dilatation assessed as measures of peripheral microvascular and conduit vessel endothelial function, respectively. Mean retinal arteriolar dilatation was attenuated in patients with CAD compared with non-CAD controls (1.51 ± 1.51% vs 2.37 ± 1.95%, p = 0.001). Retinal arteriolar dilatation was independently associated with CAD after adjustment for age, gender, cardiovascular risk factors, and medication use (odds ratio 1.60, 95% confidence interval 1.14 to 2.25, p = 0.007). Reactive hyperaemia index and flow-mediated dilatation were not different. In conclusion, the capacity of retinal arterioles to dilate in response to flicker light is an independent predictor of the presence of CAD and suggests that retinal microvascular endothelial dysfunction is a marker for underlying CAD.

  11. Endothelial Dysfunction: Associations with Exposure to Ambient Fine Particles in Diabetic Individuals

    PubMed Central

    Schneider, Alexandra; Neas, Lucas; Herbst, Margaret C.; Case, Martin; Williams, Ronald W.; Cascio, Wayne; Hinderliter, Alan; Holguin, Fernando; Buse, John B.; Dungan, Kathleen; Styner, Maya; Peters, Annette; Devlin, Robert B.

    2008-01-01

    Background Exposure to fine airborne particulate matter [≤2.5 μm in aerodynamic diameter (PM2.5)] has been associated with cardiovascular and hematologic effects, especially in older people with cardiovascular disease. Some epidemiologic studies suggest that adults with diabetes also may be a particularly susceptible population. Objectives The purpose of this study was to analyze the short-term effects of ambient PM2.5 on markers of endothelial function in diabetic volunteers. Methods We conducted a prospective panel study in 22 people with type 2 diabetes mellitus in Chapel Hill, North Carolina (USA), from November 2004 to December 2005. We acquired daily measurements of PM2.5 and meteorologic data at central monitoring sites. On 4 consecutive days, we measured endothelial function by brachial artery ultrasound in all participants and by pulsewave measurements in a subgroup. Data were analyzed using additive mixed models with a random participant effect and adjusted for season, day of the week, and meteorology. Results Flow-mediated dilatation decreased in association with PM2.5 during the first 24 hr, whereas small-artery elasticity index decreased with a delay of 1 and 3 days. These PM2.5-associated decrements in endothelial function were greater among participants with a high body mass index, high glycosylated hemoglobin A1c, low adiponectin, or the null polymorphism of glutathione S-transferase M1. However, high levels of myeloperoxidase on the examination day led to strongest effects on endothelial dysfunction. Conclusions These data demonstrate that PM2.5 exposure may cause immediate endothelial dysfunction. Clinical characteristics associated with insulin resistance were associated with enhanced effects of PM on endothelial function. In addition, participants with greater oxidative potential seem to be more susceptible. PMID:19079718

  12. Markers of erectile dysfunction

    PubMed Central

    Davies, Kelvin P.; Melman, Arnold

    2008-01-01

    With the development and marketing of oral pharmacotherapy that is both noninvasive and successful in treating erectile dysfunction (ED), the quest to identify markers of organic ED lost ground. Indeed, the multi-factorial nature of ED may have led many researchers to conclude that searching for a universal marker of ED was futile. However, the realization that ED is strongly correlated with the overall health of men, and may act as a predictor for the development of cardiovascular disease (CVD) and diabetes, has stimulated interest in identifying genes that can distinguish organic ED. In addition, the potential ability to suggest to the patient that ED is reversible (i.e., psychogenic) with a simple test would be of significance to both the physician and patient, as well as for reimbursement issues for therapy by insurance companies. Such a marker may also act as a non-subjective measure of the degree of ED and the efficacy of treatment. This review discusses the importance of identifying such markers and recent work identifying potential markers in human patients. PMID:19468461

  13. Dextran induces differentiation of circulating endothelial progenitor cells

    PubMed Central

    Obi, Syotaro; Masuda, Haruchika; Akimaru, Hiroshi; Shizuno, Tomoko; Yamamoto, Kimiko; Ando, Joji; Asahara, Takayuki

    2014-01-01

    Abstract Endothelial progenitor cells (EPCs) have been demonstrated to be effective for the treatment of cardiovascular diseases. However, the differentiation process from circulation to adhesion has not been clarified because circulating EPCs rarely attached to dishes in EPC cultures previously. Here we investigated whether immature circulating EPCs differentiate into mature adhesive EPCs in response to dextran. When floating‐circulating EPCs derived from ex vivo expanded human cord blood were cultured with 5% and 10% dextran, they attached to fibronectin‐coated dishes and grew exponentially. The bioactivities of adhesion, proliferation, migration, tube formation, and differentiated type of EPC colony formation increased in EPCs exposed to dextran. The surface protein expression rate of the endothelial markers vascular endothelial growth factor (VEGF)‐R1/2, VE‐cadherin, Tie2, ICAM1, VCAM1, and integrin αv/β3 increased in EPCs exposed to dextran. The mRNA levels of VEGF‐R1/2, VE‐cadherin, Tie2, endothelial nitric oxide synthase, MMP9, and VEGF increased in EPCs treated with dextran. Those of endothelium‐related transcription factors ID1/2, FOXM1, HEY1, SMAD1, FOSL1, NFkB1, NRF2, HIF1A, EPAS1 increased in dextran‐treated EPCs; however, those of hematopoietic‐ and antiangiogenic‐related transcription factors TAL1, RUNX1, c‐MYB, GATA1/2, ERG, FOXH1, HHEX, SMAD2/3 decreased in dextran‐exposed EPCs. Inhibitor analysis showed that PI3K/Akt, ERK1/2, JNK, and p38 signal transduction pathways are involved in the differentiation in response to dextran. In conclusion, dextran induces differentiation of circulating EPCs in terms of adhesion, migration, proliferation, and vasculogenesis. The differentiation mechanism in response to dextran is regulated by multiple signal transductions including PI3K/Akt, ERK1/2, JNK, and p38. These findings indicate that dextran is an effective treatment for EPCs in regenerative medicines. PMID:24760515

  14. Amyloid β induces adhesion of erythrocytes to endothelial cells and affects endothelial viability and functionality.

    PubMed

    Nakagawa, Kiyotaka; Kiko, Takehiro; Kuriwada, Satoko; Miyazawa, Taiki; Kimura, Fumiko; Miyazawa, Teruo

    2011-01-01

    It has been suggested that amyloid β-peptide (Aβ) might mediate the adhesion of erythrocytes to the endothelium which could disrupt the properties of endothelial cells. We provide evidence here that Aβ actually induced the binding of erythrocytes to endothelial cells and decreased endothelial viability, perhaps by the generation of oxidative and inflammatory stress. These changes are likely to contribute to the pathogenesis of Alzheimer's disease.

  15. Notch as a Diagnostic Marker and Therapeutic Target in Human Breast Cancer

    DTIC Science & Technology

    2009-09-01

    endothelial cell marker that may be exploited for diagnosis and targeted therapies as outlined in this proposal. Notch1 and Jagged-1 are over-expressed in...Expression of Hey1, a direct target of Notch/CSL signaling in ECs, was decreased 2.7-fold in the Notch1 decoy–expressing tumors compared with controls...receptor activation is required for VEGF-induced up-regulation of target genes (28). One of these endothelial target genes is Dll4, which acts to reduce

  16. Asiaticoside Inhibits TNF-α-Induced Endothelial Hyperpermeability of Human Aortic Endothelial Cells.

    PubMed

    Fong, Lai Yen; Ng, Chin Theng; Zakaria, Zainul Amiruddin; Baharuldin, Mohamad Taufik Hidayat; Arifah, Abdul Kadir; Hakim, Muhammad Nazrul; Zuraini, Ahmad

    2015-10-01

    The increase in endothelial permeability often promotes edema formation in various pathological conditions. Tumor necrosis factor-alpha (TNF-α), a pro-atherogenic cytokine, impairs endothelial barrier function and causes endothelial dysfunction in early stage of atherosclerosis. Asiaticoside, one of the triterpenoids derived from Centella asiatica, is known to possess antiinflammatory activity. In order to examine the role of asiaticoside in preserving the endothelial barrier, we assessed its effects on endothelial hyperpermeability and disruption of actin filaments evoked by TNF-α in human aortic endothelial cells (HAEC). TNF-α caused an increase in endothelial permeability to fluorescein isothiocyanate (FITC)-dextran. Asiaticoside pretreatment significantly suppressed TNF-α-induced increased permeability. Asiaticoside also prevented TNF-α-induced actin redistribution by suppressing stress fiber formation. However, the increased F to G actin ratio stimulated by TNF-α was not changed by asiaticoside. Cytochalasin D, an actin depolymerizing agent, was used to correlate the anti-hyperpermeability effect of asiaticoside with actin cytoskeleton. Surprisingly, asiaticoside failed to prevent cytochalasin D-induced increased permeability. These results suggest that asiaticoside protects against the disruption of endothelial barrier and actin rearrangement triggered by TNF-α without a significant change in total actin pool. However, asiaticoside seems to work by other mechanisms to maintain the integrity of endothelial barrier rather than stabilizing the F-actin organization.

  17. Interleukin-3 greatly expands non-adherent endothelial forming cells with pro-angiogenic properties.

    PubMed

    Moldenhauer, Lachlan M; Cockshell, Michaelia P; Frost, Lachlan; Parham, Kate A; Tvorogov, Denis; Tan, Lih Y; Ebert, Lisa M; Tooley, Katie; Worthley, Stephen; Lopez, Angel F; Bonder, Claudine S

    2015-05-01

    Circulating endothelial progenitor cells (EPCs) provide revascularisation for cardiovascular disease and the expansion of these cells opens up the possibility of their use as a cell therapy. Herein we show that interleukin-3 (IL3) strongly expands a population of human non-adherent endothelial forming cells (EXnaEFCs) with low immunogenicity as well as pro-angiogenic capabilities in vivo, making their therapeutic utilisation a realistic option. Non-adherent CD133(+) EFCs isolated from human umbilical cord blood and cultured under different conditions were maximally expanded by day 12 in the presence of IL3 at which time a 350-fold increase in cell number was obtained. Cell surface marker phenotyping confirmed expression of the hematopoietic progenitor cell markers CD133, CD117 and CD34, vascular cell markers VEGFR2 and CD31, dim expression of CD45 and absence of myeloid markers CD14 and CD11b. Functional experiments revealed that EXnaEFCs exhibited classical properties of endothelial cells (ECs), namely binding of Ulex europaeus lectin, up-take of acetylated-low density lipoprotein and contribution to EC tube formation in vitro. These EXnaEFCs demonstrated a pro-angiogenic phenotype within two independent in vivo rodent models. Firstly, a Matrigel plug assay showed increased vascularisation in mice. Secondly, a rat model of acute myocardial infarction demonstrated reduced heart damage as determined by lower levels of serum creatinine and a modest increase in heart functionality. Taken together, these studies show IL3 as a potent growth factor for human CD133(+) cell expansion with clear pro-angiogenic properties (in vitro and in vivo) and thus may provide clinical utility for humans in the future.

  18. Genetics Home Reference: Fuchs endothelial dystrophy

    MedlinePlus

    ... FUCHS ENDOTHELIAL, 7 Sources for This Page Afshari NA, Li YJ, Pericak-Vance MA, Gregory S, Klintworth GK. ... Free article on PubMed Central Riazuddin SA, Zaghloul NA, Al-Saif A, Davey L, Diplas BH, Meadows ...

  19. Endothelial progenitor cells in cardiovascular diseases

    PubMed Central

    Lee, Poay Sian Sabrina; Poh, Kian Keong

    2014-01-01

    Endothelial dysfunction has been associated with the development of atherosclerosis and cardiovascular diseases. Adult endothelial progenitor cells (EPCs) are derived from hematopoietic stem cells and are capable of forming new blood vessels through a process of vasculogenesis. There are studies which report correlations between circulating EPCs and cardiovascular risk factors. There are also studies on how pharmacotherapies may influence levels of circulating EPCs. In this review, we discuss the potential role of endothelial progenitor cells as both diagnostic and prognostic biomarkers. In addition, we look at the interaction between cardiovascular pharmacotherapies and endothelial progenitor cells. We also discuss how EPCs can be used directly and indirectly as a therapeutic agent. Finally, we evaluate the challenges facing EPC research and how these may be overcome. PMID:25126384

  20. Endothelial glucocorticoid receptor suppresses atherogenesis- Brief Report

    PubMed Central

    Zhang, Xinbo; Rotllan, Noemi; Feng, Yan; Zhou, Han; Fernández-Hernando, Carlos; Yu, Jun; Sessa, William C.

    2015-01-01

    Objective The purpose of this study was to determine the role of the endothelial glucocorticoid receptor in the pathogenesis of atherosclerosis. Approach and Results Control mice and mice lacking the endothelial glucocorticoid receptor were bred onto an Apoe knockout background and subjected to high-fat diet feeding for 12 weeks. Assessment of body weight and total cholesterol and triglycerides before and after the diet revealed no differences between the two groups of mice. However, mice lacking the endothelial glucocorticoid receptor developed more severe atherosclerotic lesions in the aorta, brachiocephalic artery and aortic sinus as well as a heightened inflammatory milieu as evidence by increased macrophage recruitment in the lesions. Conclusions These data suggest the endothelial glucocorticoid receptor is important for tonic inhibition of inflammation and limitation of atherosclerosis progression in this model. PMID:25810297

  1. Mechanisms of endothelial dysfunction in obesity.

    PubMed

    Avogaro, Angelo; de Kreutzenberg, Saula Vigili

    2005-10-01

    Obesity is a chronic disease, whose incidence is alarmingly growing, affecting not only adults but also children and adolescents. It is associated with severe metabolic abnormalities and increased cardiovascular morbidity and mortality. Adipose tissue secretes a great number of hormones and cytokines that not only regulate substrate metabolism but may deeply and negatively influence endothelial physiology, a condition which may lead to the formation of the atherosclerotic plaque. In this review, the physiology of the endothelium is summarised and the mechanisms by which obesity, through the secretory products of adipose tissue, influences endothelial function are explained. A short description of methodological approaches to diagnose endothelial dysfunction is presented. The possible pathogenetic links between obesity and cardiovascular disease, mediated by oxidative stress, inflammation and endothelial dysfunction are described as well.

  2. Apicobasal polarity of brain endothelial cells

    PubMed Central

    Worzfeld, Thomas

    2015-01-01

    Normal brain homeostasis depends on the integrity of the blood–brain barrier that controls the access of nutrients, humoral factors, and immune cells to the CNS. The blood–brain barrier is composed mainly of brain endothelial cells. Forming the interface between two compartments, they are highly polarized. Apical/luminal and basolateral/abluminal membranes differ in their lipid and (glyco-)protein composition, allowing brain endothelial cells to secrete or transport soluble factors in a polarized manner and to maintain blood flow. Here, we summarize the basic concepts of apicobasal cell polarity in brain endothelial cells. To address potential molecular mechanisms underlying apicobasal polarity in brain endothelial cells, we draw on investigations in epithelial cells and discuss how polarity may go awry in neurological diseases. PMID:26661193

  3. Identification of Tumor Endothelial Cells with High Aldehyde Dehydrogenase Activity and a Highly Angiogenic Phenotype

    PubMed Central

    Maishi, Nako; Ohga, Noritaka; Hida, Yasuhiro; Kawamoto, Taisuke; Iida, Junichiro; Shindoh, Masanobu; Tsuchiya, Kunihiko; Shinohara, Nobuo; Hida, Kyoko

    2014-01-01

    Tumor blood vessels play an important role in tumor progression and metastasis. It has been reported that tumor endothelial cells (TECs) exhibit highly angiogenic phenotypes compared with those of normal endothelial cells (NECs). TECs show higher proliferative and migratory abilities than those NECs, together with upregulation of vascular endothelial growth factor (VEGF) and VEGF receptor 2 (VEGFR2). Furthermore, compared with NECs, stem cell markers such as Sca-1, CD90, and multidrug resistance 1 are upregulated in TECs, suggesting that stem-like cells exist in tumor blood vessels. In this study, to reveal the biological role of stem-like TECs, we analyzed expression of the stem cell marker aldehyde dehydrogenase (ALDH) in TECs and characterized ALDHhigh TECs. TECs and NECs were isolated from melanoma-xenografted nude mice and normal dermis, respectively. ALDH mRNA expression and activity were higher in TECs than those in NECs. Next, ALDHhigh/low TECs were isolated by fluorescence-activated cell sorting to compare their characteristics. Compared with ALDHlow TECs, ALDHhigh TECs formed more tubes on Matrigel-coated plates and sustained the tubular networks longer. Furthermore, VEGFR2 expression was higher in ALDHhigh TECs than that in ALDHlow TECs. In addition, ALDH was expressed in the tumor blood vessels of in vivo mouse models of melanoma and oral carcinoma, but not in normal blood vessels. These findings indicate that ALDHhigh TECs exhibit an angiogenic phenotype. Stem-like TECs may have an essential role in tumor angiogenesis. PMID:25437864

  4. Association of Microvascular Function and Endothelial Biomarkers With Clinical Outcome in Dengue: An Observational Study

    PubMed Central

    Yacoub, Sophie; Lam, Phung Khanh; Vu, Le Hoang Mai; Le, Thi Lien; Ha, Ngo Thanh; Toan, Tran Thi; Van, Nguyen Thu; Quyen, Nguyen Than Ha; Le Duyen, Huynh Thi; Van Kinh, Nguyen; Fox, Annette; Mongkolspaya, Juthathip; Wolbers, Marcel; Simmons, Cameron Paul; Screaton, Gavin Robert; Wertheim, Heiman; Wills, Bridget

    2016-01-01

    Background. The hallmark of severe dengue is increased microvascular permeability, but alterations in the microcirculation and their evolution over the course of dengue are unknown. Methods. We conducted a prospective observational study to evaluate the sublingual microcirculation using side-stream dark-field imaging in patients presenting early (<72 hours after fever onset) and patients hospitalized with warning signs or severe dengue in Vietnam. Clinical findings, microvascular function, global hemodynamics assessed with echocardiography, and serological markers of endothelial activation were determined at 4 time points. Results. A total of 165 patients were enrolled. No difference was found between the microcirculatory parameters comparing dengue with other febrile illnesses. The proportion of perfused vessels (PPV) and the mean flow index (MFI) were lower in patients with dengue with plasma than those without leakage (PPV, 88.1% vs 90.6% [P = .01]; MFI, 2.1 vs 2.4 [P = .007]), most markedly during the critical phase. PPV and MFI were correlated with the endothelial activation markers vascular cell adhesion molecule 1 (P < .001 for both) and angiopoietin 2 (P < .001 for both), negatively correlated. Conclusions. Modest microcirculatory alterations occur in dengue, are associated with plasma leakage, and are correlate with molecules of endothelial activation, angiopoietin 2 and vascular cell adhesion molecule 1. PMID:27230099

  5. Moderate endurance exercise in patients with sickle cell anaemia: effects on oxidative stress and endothelial activation.

    PubMed

    Faes, Camille; Balayssac-Siransy, Edwige; Connes, Philippe; Hivert, Ludovic; Danho, Clotaire; Bogui, Pascal; Martin, Cyril; Pialoux, Vincent

    2014-01-01

    Very few studies have investigated the effects of exercise on the biological parameters involved in vaso-occlusive events in sickle cell anaemia (SCA). The aim of this study was to test how a mild-moderate endurance exercise modulates oxidative stress, nitric oxide bioavailability and endothelial activation in SCA patients and healthy individuals. Eleven patients with SCA and 15 healthy subjects completed a 20-min duration submaximal cycling exercise at ≈45 Watts. Plasma markers of oxidative stress, antioxidant activity, endothelial activation and nitric oxide bioavailability were investigated before and after the exercise. Nitric oxide levels, anti-oxidant capacity, soluble (s)E-selectin and sP-selectin did not change in response to this exercise. Except for the malondialdehyde levels, which increased in the two groups, the other markers of oxidative stress remained unchanged in both groups in response to exercise. Soluble vascular cell adhesion molecule 1 levels were increased at the end of exercise in both groups. sL-selectin decreased and soluble intercellular adhesion molecule 1 increased with exercise in SCA patients only. The present data suggest that patients with SCA may undertake mild-moderate physical activities without any acute clinical complications, but care should be taken because oxidative stress and endothelial activation significantly increased in some patients.

  6. Deeper Penetration of Erythrocytes into the Endothelial Glycocalyx Is Associated with Impaired Microvascular Perfusion

    PubMed Central

    Lee, Dae Hyun; Dane, Martijn J. C.; van den Berg, Bernard M.; Boels, Margien G. S.; van Teeffelen, Jurgen W.; de Mutsert, Renée; den Heijer, Martin; Rosendaal, Frits R.; van der Vlag, Johan; van Zonneveld, Anton Jan; Vink, Hans; Rabelink, Ton J.

    2014-01-01

    Changes in endothelial glycocalyx are one of the earliest changes in development of cardiovascular disease. The endothelial glycocalyx is both an important biological modifier of interactions between flowing blood and the vessel wall, and a determinant of organ perfusion. We hypothesize that deeper penetration of erythrocytes into the glycocalyx is associated with reduced microvascular perfusion. The population-based prospective cohort study (the Netherlands Epidemiology of Obesity [NEO] study) includes 6,673 middle-aged individuals (oversampling of overweight and obese individuals). Within this cohort, we have imaged the sublingual microvasculature of 915 participants using sidestream darkfield (SDF) imaging together with a recently developed automated acquisition and analysis approach. Presence of RBC (as a marker of microvascular perfusion) and perfused boundary region (PBR), a marker for endothelial glycocalyx barrier properties for RBC accessibility, were assessed in vessels between 5 and 25 µm RBC column width. A wide range of variability in PBR measurements, with a mean PBR of 2.14 µm (range: 1.43–2.86 µm), was observed. Linear regression analysis showed a marked association between PBR and microvascular perfusion, reflected by RBC filling percentage (regression coefficient β: −0.034; 95% confidence interval: −0.037 to −0.031). We conclude that microvascular beds with a thick (“healthy”) glycocalyx (low PBR), reflects efficient perfusion of the microvascular bed. In contrast, a thin (“risk”) glycocalyx (high PBR) is associated with a less efficient and defective microvascular perfusion. PMID:24816787

  7. Evidence of endothelial progenitor cells in the human brain and spinal cord arteriovenous malformations

    PubMed Central

    Gao, Peng; Chen, Yongmei; Lawton, Michael T.; Barbaro, Nicholas M.; Yang, Guo-Yuan; Su, Hua; Ling, Feng; Young, William L.

    2010-01-01

    Objective Brain and spinal cord arteriovenous malformations (AVMs) are characterized by aberrant angiogenesis and vascular remodeling. Endothelial progenitor cells (EPCs) can be recruited by stromal cell-derived factor-1 (SDF-1), and participate in vascular remodeling in both physiological and pathological settings. We hypothesized that there was increased EPC levels in the brain and spinal cord AVM nidus. Methods Microsurgical specimens without endovascular embolization and radiosurgery from the brain (n=12) and spinal cord (n=5) AVMs were examined. Hemangioblastoma, meningioma, cerebral cortex obtained from epilepsy surgery, and the basilar artery (BA) from the autopsy were chosen for control comparisons. EPCs were identified as cells that were double-positive for the stem cell marker CD133 and the endothelial cell marker VEGFR-2 (vascular endothelial growth factor receptor-2 or KDR). In addition, SDF-1 was characterized by immunohistochemistry. Results Both brain and spinal AVM tissues displayed more CD133, SDF-1, and CD68-positive signals than epilepsy and basilar artery control tissues. The level of EPCs was increased in the brain and spinal cord AVM nidus, mainly at the edge of the vessel wall. The expression of SDF-1 was co-localized with CD31-positive and α-smooth muscle cells, and was predominantly found within the vessel wall. Conclusion Our data demonstrate that EPCs are present in the nidus of the brain and spinal cord AVMs, which may mediate pathological vascular remodeling and impact the clinical course of AVMs. PMID:20881566

  8. Black tea protects against hypertension-associated endothelial dysfunction through alleviation of endoplasmic reticulum stress

    PubMed Central

    San Cheang, Wai; Yuen Ngai, Ching; Yen Tam, Ye; Yu Tian, Xiao; Tak Wong, Wing; Zhang, Yang; Wai Lau, Chi; Chen, Zhen Yu; Bian, Zhao-Xiang; Huang, Yu; Ping Leung, Fung

    2015-01-01

    Hypertensive patients have been found to be associated with elevated levels of homocysteine, known as hyperhomocysteinemia. Homocysteine (Hcy) can induce endoplasmic reticulum (ER) stress in endothelial cells. This study aims to investigate whether black tea (BT) protects against hypertension-associated endothelial dysfunction through alleviation of ER stress. Rat aortae and cultured rat aortic endothelial cells were treated with Hcy, BT extract, and theaflavin-3,3’-digallate (TF3). Male Sprague Dawley rats were infused with angiotensin II (Ang II) to induce hypertension and orally administrated with BT extract at 15 mg/kg/day for 2 weeks. Hcy impaired endothelium-dependent relaxations of rat aortae and led to ER stress in endothelial cells, which were ameliorated by co-incubation of BT extract and TF3. The blood pressure of Ang II-infused rats and plasma Hcy level were normalized by BT consumption. Impaired endothelium-dependent relaxations in renal arteries, carotid arteries and aortae, and flow-mediated dilatations in third-order mesenteric resistance arteries were improved. Elevations of ER stress markers and ROS level, plus down-regulation of Hcy metabolic enzymes in aortae from Ang II-infused rats were prevented by BT treatment. Our data reveal the novel cardiovascular benefits of BT in ameliorating vascular dysfunctions, providing insight into developing BT into beneficial dietary supplements in hypertensive patients. PMID:25976123

  9. Cooperative control of blood compatibility and re-endothelialization by immobilized heparin and substrate topography.

    PubMed

    Ding, Yonghui; Yang, Meng; Yang, Zhilu; Luo, Rifang; Lu, Xiong; Huang, Nan; Huang, Pingbo; Leng, Yang

    2015-03-01

    A wide variety of environmental cues provided by the extracellular matrix, including biophysical and biochemical cues, are responsible for vascular cell behavior and function. In particular, substrate topography and surface chemistry have been shown to regulate blood and vascular compatibility individually. The combined impact of chemical and topographic cues on blood and vascular compatibility, and the interplay between these two types of cues, are subjects that are currently being explored. In the present study, a facile polydopamine-mediated approach is introduced for immobilization of heparin on topographically patterned substrates, and the combined effects of these cues on blood compatibility and re-endothelialization are systematically investigated. The results show that immobilized heparin and substrate topography cooperatively modulate anti-coagulation activity, endothelial cell (EC) attachment, proliferation, focal adhesion formation and endothelial marker expression. Meanwhile, the substrate topography is the primary determinant of cell alignment and elongation, driving in vivo-like endothelial organization. Importantly, combining immobilized heparin with substrate topography empowers substantially greater competitive ability of ECs over smooth muscle cells than each cue individually. Moreover, a model is proposed to elucidate the cooperative interplay between immobilized heparin and substrate topography in regulating cell behavior.

  10. Vitamin D deficiency and its relationship with endothelial dysfunction in patients with early Parkinson's disease.

    PubMed

    Yoon, Jung Han; Park, Dong Kyu; Yong, Seok Woo; Hong, Ji Man

    2015-12-01

    Increasing evidence has shown that individuals with Parkinson's disease (PD) have lower levels of 25-hydroxyvitamin D (25[OH]D) than healthy controls. Low vitamin D has been associated with endothelial dysfunction which may play a role in the pathogenesis and progression of PD. Flow-mediated dilation (FMD) is widely used as a clinical marker of overall endothelial function. We evaluated the relationship between serum 25(OH)D levels and FMD in PD. We enrolled 81 patients with early PD and 52 healthy controls, and we evaluate endothelial function based on vitamin D status and identify the association between FMD and vitamin D status in patients with early PD. The mean serum 25(OH)D levels were significantly lower in the PD patients than in the controls (21.8 ± 9.5 vs. 25.2 ± 9.3 ng/mL, p < 0.05). FMD was significantly lower in the PD patients (7.1 ± 1.8 %) than in the controls (8.1 ± 2.1 %, p < 0.05). The serum 25(OH)D was significantly associated with FMD independently of age, cardiovascular disease risk factors, body mass index, motor Unified PD Rating Scale status and homocysteine levels (adjusted R (2) = 0.331, β = 0.494, p < 0.001). These findings provide evidence of a possible association between endothelial dysfunction as assessed by FMD and low vitamin D status in patients with early PD.

  11. Inflammatory response of endothelial cells to hepatitis C virus recombinant envelope glycoprotein 2 protein exposure.

    PubMed

    Urbaczek, Ana Carolina; Ribeiro, Lívia Carolina de Abreu; Ximenes, Valdecir Farias; Afonso, Ana; Nogueira, Camila Tita; Generoso, Wesley Cardoso; Alberice, Juliana Vieira; Rudnicki, Martina; Ferrer, Renila; Fonseca, Luiz Marcos da; Costa, Paulo Inácio da

    2014-09-01

    The hepatitis C virus (HCV) encodes approximately 10 different structural and non-structural proteins, including the envelope glycoprotein 2 (E2). HCV proteins, especially the envelope proteins, bind to cell receptors and can damage tissues. Endothelial inflammation is the most important determinant of fibrosis progression and, consequently, cirrhosis. The aim of this study was to evaluate and compare the inflammatory response of endothelial cells to two recombinant forms of the HCV E2 protein produced in different expression systems (Escherichia coli and Pichia pastoris). We observed the induction of cell death and the production of nitric oxide, hydrogen peroxide, interleukin-8 and vascular endothelial growth factor A in human umbilical vein endothelial cells (HUVECs) stimulated by the two recombinant E2 proteins. The E2-induced apoptosis of HUVECs was confirmed using the molecular marker PARP. The apoptosis rescue observed when the antioxidant N-acetylcysteine was used suggests that reactive oxygen species are involved in E2-induced apoptosis. We propose that these proteins are involved in the chronic inflammation caused by HCV.

  12. Mouse stem cells seeded into decellularized rat kidney scaffolds endothelialize and remodel basement membranes

    PubMed Central

    Ross, Edward A.; Abrahamson, Dale R.; St. John, Patricia; Clapp, William L.; Williams, Matthew J.; Terada, Naohiro; Hamazaki, Takashi; Ellison, Gary W.; Batich, Christopher D.

    2012-01-01

    Introduction To address transplant organ shortage, a promising strategy is to decellularize kidneys in a manner that the scaffold retains signals for seeded pluripotent precursor cells to differentiate and recapitulate native structures: matrix-to-cell signaling followed by cell-cell and cell-matrix interactions, thereby remodeling and replacing the original matrix. This would reduce scaffold antigenicity and enable xeno-allografts. Results DAPI-labeled cells in arterial vessels and glomeruli were positive for both endothelial lineage markers, BsLB4 and VEGFR2. Rat scaffold’s basement membrane demonstrated immunolabeling with anti-mouse laminin β1. Labeling intensified over time with 14 day incubations. Conclusion We provide new evidence for matrix-to-cell signaling in acellular whole organ scaffolds that induces differentiation of pluripotent precursor cells to endothelial lineage. Production of mouse basement membrane supports remodeling of host (rat)-derived scaffolds and thereby warrants further investigation as a promising approach for xenotransplantation. Methods We previously showed that murine embryonic stem cells arterially seeded into acellular rat whole kidney scaffolds multiply and demonstrate morphologic, immunohistochemical and gene expression evidence for differentiation. Vascular cell endothelialization was now further tested by endothelial specific BsLB4 lectin and anti-VEGFR2 (Flk1) antibodies. Remodeling of the matrix basement membranes from rat to mouse (“murinization”) was assessed by a monoclonal antibody specific for mouse laminin β1 chain. PMID:22692231

  13. Circulating endothelial cells are increased in chronic myeloid leukemia blast crisis.

    PubMed

    Godoy, C R T; Levy, D; Giampaoli, V; Chamone, D A F; Bydlowski, S P; Pereira, J

    2015-06-01

    We measured circulating endothelial precursor cells (EPCs), activated circulating endothelial cells (aCECs), and mature circulating endothelial cells (mCECs) using four-color multiparametric flow cytometry in the peripheral blood of 84 chronic myeloid leukemia (CML) patients and 65 healthy controls; and vascular endothelial growth factor (VEGF) by quantitative real-time PCR in 50 CML patients and 32 healthy controls. Because of an increase in mCECs, the median percentage of CECs in CML blast crisis (0.0146%) was significantly higher than in healthy subjects (0.0059%, P<0.01) and in the accelerated phase (0.0059%, P=0.01). There were no significant differences in the percentages of CECs in chronic- or active-phase patients and healthy subjects (P>0.05). In addition, VEGF gene expression was significantly higher in all phases of CML: 0.245 in blast crisis, 0.320 in the active phase, and 0.330 in chronic phase patients than it was in healthy subjects (0.145). In conclusion, CML in blast crisis had increased levels of CECs and VEGF gene expression, which may serve as markers of disease progression and may become targets for the management of CML.

  14. Directing migration of endothelial progenitor cells with applied DC electric fields.

    PubMed

    Zhao, Zhiqiang; Qin, Lu; Reid, Brian; Pu, Jin; Hara, Takahiko; Zhao, Min

    2012-01-01

    Naturally-occurring, endogenous electric fields (EFs) have been detected at skin wounds, damaged tissue sites and vasculature. Applied EFs guide migration of many types of cells, including endothelial cells to migrate directionally. Homing of endothelial progenitor cells (EPCs) to an injury site is important for repair of vasculature and also for angiogenesis. However, it has not been reported whether EPCs respond to applied EFs. Aiming to explore the possibility to use electric stimulation to regulate the progenitor cells and angiogenesis, we tested the effects of direct-current (DC) EFs on EPCs. We first used immunofluorescence to confirm the expression of endothelial progenitor markers in three lines of EPCs. We then cultured the progenitor cells in EFs. Using time-lapse video microscopy, we demonstrated that an applied DC EF directs migration of the EPCs toward the cathode. The progenitor cells also align and elongate in an EF. Inhibition of vascular endothelial growth factor (VEGF) receptor signaling completely abolished the EF-induced directional migration of the progenitor cells. We conclude that EFs are an effective signal that guides EPC migration through VEGF receptor signaling in vitro. Applied EFs may be used to control behaviors of EPCs in tissue engineering, in homing of EPCs to wounds and to an injury site in the vasculature.

  15. Endothelial cells assemble into a 3-dimensional prevascular network in a bone tissue engineering construct.

    PubMed

    Rouwkema, Jeroen; de Boer, Jan; Van Blitterswijk, Clemens A

    2006-09-01

    To engineer tissues with clinically relevant dimensions, one must overcome the challenge of rapidly creating functional blood vessels to supply cells with oxygen and nutrients and to remove waste products. We tested the hypothesis that endothelial cells, cocultured with osteoprogenitor cells, can organize into a prevascular network in vitro. When cultured in a spheroid coculture model with human mesenchymal stem cells, human umbilical vein endothelial cells (HUVECs) form a 3-dimensional prevascular network within 10 days of in vitro culture. The formation of the prevascular network was promoted by seeding 2% or fewer HUVECs. Moreover, the addition of endothelial cells resulted in a 4-fold upregulation of the osteogenic marker alkaline phosphatase. The addition of mouse embryonic fibroblasts did not result in stabilization of the prevascular network. Upon implantation, the prevascular network developed further and structures including lumen could be seen regularly. However, anastomosis with the host vasculature was limited. We conclude that endothelial cells are able to form a 3-dimensional (3D) prevascular network in vitro in a bone tissue engineering setting. This finding is a strong indication that in vitro prevascularization is a promising strategy to improve implant vascularization in bone tissue engineering.

  16. Oral cancer/endothelial cell fusion experiences nuclear fusion and acquisition of enhanced survival potential.

    PubMed

    Song, Kai; Song, Yong; Zhao, Xiao-Ping; Shen, Hui; Wang, Meng; Yan, Ting-Lin; Liu, Ke; Shang, Zheng-Jun

    2014-10-15

    Most previous studies have linked cancer-macrophage fusion with tumor progression and metastasis. However, the characteristics of hybrid cells derived from oral cancer and endothelial cells and their involvement in cancer remained unknown. Double-immunofluorescent staining and fluorescent in situ hybridization (FISH) were performed to confirm spontaneous cell fusion between eGFP-labeled human umbilical vein endothelial cells (HUVECs) and RFP-labeled SCC9, and to detect the expression of vementin and cytokeratin 18 in the hybrids. The property of chemo-resistance of such hybrids was examined by TUNEL assay. The hybrid cells in xenografted tumor were identified by FISH and GFP/RFP dual-immunofluoresence staining. We showed that SCC9 cells spontaneously fused with cocultured endothelial cells, and the resultant hybrid cells maintained the division and proliferation activity after re-plating and thawing. Such hybrids expressed markers of both parental cells and became more resistant to chemotherapeutic drug cisplatin as compared to the parental SCC9 cells. Our in vivo data indicated that the hybrid cells contributed to tumor composition by using of immunostaining and FISH analysis, even though the hybrid cells and SCC9 cells were mixed with 1:10,000, according to the FACS data. Our study suggested that the fusion events between oral cancer and endothelial cells undergo nuclear fusion and acquire a new property of drug resistance and consequently enhanced survival potential. These experimental findings provide further supportive evidence for the theory that cell fusion is involved in cancer progression.

  17. Short-term lifestyle modification alters circulating biomarkers of endothelial health in sedentary, overweight adults.

    PubMed

    Rector, R Scott; Turk, James R; Sun, Grace Y; Guilford, Brianne L; Toedebusch, Brian W; McClanahan, Matt W; Thomas, Tom R

    2006-10-01

    Obesity and inactivity are associated with endothelial dysfunction that may contribute to the development of atherosclerosis. We examined the effects of a short-term lifestyle intervention on circulating biomarkers of endothelial health. Nineteen overweight or obese (mean body mass index (BMI): 28.9 +/- 0.7 kg/m2) men and women underwent 6 weeks of body mass reduction induced by moderate energy restriction (approximately 750 kcal/d; 1 kcal = 4.184 kJ) and aerobic training (approximately 400 kcal/d). Fasting serum samples were collected at baseline and after reduction in body mass (week 6) to assess concentrations of nitrotyrosine (NT), secretory phospholipase A2 (sPLA2), and soluble intracellular adhesion molecule-1 (sICAM-1). Body mass was significantly reduced from 81.3 +/- 2.8 to 77.3 +/- 2.6 kg (p < 0.05). Circulating concentrations of NT and sICAM-1 were significantly reduced with treatment (approximately 25% and approximately 10%, respectively), whereas sPLA2 levels were significantly elevated (approximately 45%). Elevations in sPLA2 were negatively correlated with changes in NT (r = -0.58, p = 0.047); reductions in NT did not correlate significantly with reductions in sICAM-1. It appears that circulating markers of endothelial health are susceptible to short-term exercise interventions with modest reduction in body mass, and such a lifestyle modification may improve endothelial health by reducing protein nitration products and cellular adhesion.

  18. Differentiation of lymphatic endothelial cells from bone marrow mesenchymal stem cells with VEGFs.

    PubMed

    Wei, Luwan; Liu, Yanli; Chen, Guoling; Fang, Yunhai; Song, Xianrang; Dong, Ping; Gao, Jie; Liu, Rong; Ding, Zhaoxi; Bi, Yushun; Liu, Zhiyu

    2012-12-01

    Although there have been many experimental studies demonstrating that bone marrow-derived mesenchymal stem cells (MSCs) have the potential to differentiate into mesenchymal tissues such as osteocytes, chondrocytes, and adipocytes in vivo and in vitro, little information is available regarding their potential to differentiate into lymphatic endothelial cells. Therefore, we chose to investigate differentiation of MSCs into lymphatic endothelial cells using stimulation with members of the vascular endothelial growth factor (VEGFs) family. Rat MSCs were isolated from bone marrow aspirate of Sprague-Dawley rats as previously described and characterized with flow cytometry for surface markers CD14, CD34, CD29, and CD90. Purified MSCs were plated and cultured in the presence of VEGF-A, VEGF-C, or the combination of both for 10 days. We examined the cells for Prox-1 and LYVE-1 by immunocytochemistry, RT-PCR, and Western blot analysis. Results demonstrated that compared to controls, cell differentiated with VEGF-A, VEGF-C and VEGF-A+VEGF-C expressed Prox-1 and LYVE-1. Our results indicate that MSCs induced by VEGFs are capable of differentiating into lymphatic endothelial-like cells in vitro, and this response has the potential to make them attractive candidates for the development of autologous tissue grafts for future therapy.

  19. Mobilization of endothelial progenitor cells in acute cardiovascular events in the PROCELL study: time-course after acute myocardial infarction and stroke.

    PubMed

    Regueiro, Ander; Cuadrado-Godia, Elisa; Bueno-Betí, Carlos; Diaz-Ricart, Maribel; Oliveras, Anna; Novella, Susana; Gené, Gemma González; Jung, Carole; Subirana, Isaac; Ortiz-Pérez, Jose Tomás; Roqué, Mercè; Freixa, Xavier; Núñez, Julio; Escolar, Gines; Marrugat, Jaume; Hermenegildo, Carlos; Valverde, Miguel Angel; Roquer, Jaume; Sanchis, Juan; Heras, Magda

    2015-03-01

    The mobilization pattern and functionality of endothelial progenitor cells after an acute ischemic event remain largely unknown. The aim of our study was to characterize and compare the short- and long-term mobilization of endothelial progenitor cells and circulating endothelial cells after acute myocardial infarction or atherothrombotic stroke, and to determine the relationship between these cell counts and plasma concentrations of vascular cell adhesion molecule (VCAM-1) and Von Willebrand factor (VWF) as surrogate markers of endothelial damage and inflammation. In addition, we assessed whether endothelial progenitor cells behave like functional endothelial cells. We included 150 patients with acute myocardial infarction or atherothrombotic stroke and 145 controls. Endothelial progenitor cells [CD45-, CD34+, KDR+, CD133+], circulating endothelial cells [CD45-, CD146+, CD31+], VWF, and VCAM-1 levels were measured in controls (baseline only) and in patients within 24h (baseline) and at 7, 30, and 180 days after the event. Myocardial infarction patients had higher counts of endothelial progenitor cells and circulating endothelial cells than the controls (201.0/mL vs. 57.0/mL; p<0.01 and 181.0/mL vs. 62.0/mL; p<0.01). Endothelial progenitor cells peaked at 30 days post-infarction (201.0/mL vs. 369.5/mL; p<0.01), as did VCAM-1 (573.7 ng/mL vs. 701.8 ng/mL; p<0.01). At 180 days post-infarction, circulating endothelial cells and VWF decreased, compared to baseline. In stroke patients, the number of endothelial progenitor cells - but not circulating endothelial cells - was higher than in controls (90.0/mL vs. 37.0/mL; p=0.01; 105.0/mL vs. 71.0/mL; p=0.11). At 30 days after stroke, however, VCAM-1 peaked (628.1/mL vs. 869.1/mL; p<0.01) but there was no significant change in endothelial progenitor cells (90/mL vs. 78/mL; p<0.34). At 180 days after stroke, circulating endothelial cells and VWF decreased, compared to baseline. Cultured endothelial progenitor cells from

  20. Insulin resistance and vessel endothelial function.

    PubMed Central

    van Oostrom, A J H H M; Cabezas, M Castro; Rabelink, T J

    2002-01-01

    IRS is a complex disease consisting of a clustering of metabolic disorders, of which hyperglycaemia, hyper-insulinaemia and dyslipidaemia are the most important. Endothelial dysfunction plays an important role in the pathogenesis of atherosclerosis. The effects of hyperinsulinaemia seem to depend on lipidaemia and glycaemia. Hyperglycaemia and hyperlipidaemia have detrimental effects on endothelial function in the fasting as well as the postprandial states. In both situations, the generation of ROS and vasoactive molecules plays a major role in interfering with the atheroprotective endothelium-dependent NO system. Treatment of IRS in regard to endothelial function should be focused initially on lifestyle improvement, such as stopping smoking and eating a balanced diet containing antioxidant vitamins, folic-acid, L-arginine and long-chain omega-3 unsaturated FA. Strict glucose control has shown to improve endothelial function and decrease microvascular complications. However, macrovascular complications, in line with endothelial functional improvement, have so far been reduced only when treatment was focused on other characteristics of the IRS syndrome, in particular dyslipidaemia. Other relevant treatments include ACE inhibitors and thiazolidinediones, and probably tetrahydrobiopterin and folic acid supplementation. Future studies should address the effects of therapeutic neovascularization on endothelial dysfunction. PMID:12216328

  1. HUMAN VASCULAR ENDOTHELIAL CELLS IN CULTURE

    PubMed Central

    Gimbrone, Michael A.; Cotran, Ramzi S.; Folkman, Judah

    1974-01-01

    Human endothelial cells, obtained by collagenase treatment of term umbilical cord veins, were cultured using Medium 199 supplemented with 20% fetal calf serum. Small clusters of cells initially spread on plastic or glass, coalesced and grew to form confluent monolayers of polygonal cells by 7 days. Cells in primary and subcultures were identified as endothelium by the presence of Weibel-Palade bodies by electron microscopy. A morphologically distinct subpopulation of cells contaminating some primary endothelial cultures was selectively subcultured, and identified by ultrastructural criteria as vascular smooth muscle. Autoradiography of endothelial cells after exposure to [3H]thymidine showed progressive increases in labeling in growing cultures beginning at 24 h. In recently confluent cultures, labeling indices were 2.4% in central closely packed regions, and 53.2% in peripheral growing regions. 3 days after confluence, labeling was uniform, being 3.5 and 3.9% in central and peripheral areas, respectively. When small areas of confluent cultures were experimentally "denuded," there were localized increases in [3H]thymidine labeling and eventual reconstitution of the monolayer. Liquid scintillation measurements of [3H]thymidine incorporation in primary and secondary endothelial cultures in microwell trays showed a similar correlation of DNA synthesis with cell density. These data indicate that endothelial cell cultures may provide a useful in vitro model for studying pathophysiologic factors in endothelial regeneration. PMID:4363161

  2. Endothelial cell dynamics in vascular remodelling.

    PubMed

    Barbacena, Pedro; Carvalho, Joana R; Franco, Claudio A

    2016-01-01

    In this ESCHM 2016 conference talk report, we summarise two recently published original articles Franco et al. PLoS Biology 2015 and Franco et al. eLIFE 2016. The vascular network undergoes extensive vessel remodelling to become fully functional. Is it well established that blood flow is a main driver for vascular remodelling. It has also been proposed that vessel pruning is a central process within physiological vessel remodelling. However, despite its central function, the cellular and molecular mechanisms regulating vessel regression, and their interaction with blood flow patterns, remain largely unexplained. We investigated the cellular process governing developmental vascular remodelling in mouse and zebrafish. We established that polarised reorganization of endothelial cells is at the core of vessel regression, representing vessel anastomosis in reverse. Moreover, we established for the first time an axial polarity map for all endothelial cells together with an in silico method for the computation of the haemodynamic forces in the murine retinal vasculature. Using network-level analysis and microfluidics, we showed that endothelial non-canonical Wnt signalling regulates endothelial sensitivity to shear forces. Loss of Wnt5a/11 renders endothelial cells more sensitive to shear, resulting in axial polarisation at lower shear stress levels. Collectively our data suggest that non-canonical Wnt signalling stabilizes forming vascular networks by reducing endothelial shear sensitivity, thus keeping vessels open under low flow conditions that prevail in the primitive plexus.

  3. [SERUM LEVEL OF ENDOTHELIAL MONOCYTE ACTIVATING POLYPEPTIDE-II IN CHILDHOOD-ONSET TYPE 1 DIABETIC PATIENTS AND OBESE ADOLESCENTS].

    PubMed

    Mogylnytska, L A

    2015-01-01

    The atherosclerotic process begins in adolescence, and its progression is determined by the same risk factors as in adults. Endothelial monocyte activating polypeptide-II (EMAP-II) is a multifunctional cytokine with proinflammatory and antiangiogenetic activity that may play a pathogenic role in the development of endothelial dysfunction and atherosclerosis. The aim of our study was to determine the serum level of EMAP-II in childhood-onset type 1 diabetic patients and obese adolescents. We found increased of serum level of EMAP-II in childhood-onset type 1 diabetic patients and in patients with obesity that do not suffer from diabetes. Also, the level of EMAP-II correlated with the serum level of glycosylated hemoglobin and blood glucose, and key markers of lipid metabolism, body mass index. Increased serum level of EMAP-II may be one of the pathway of endothelial dysfunction in type 1 diabetes.

  4. The Swift Turbidity Marker

    ERIC Educational Resources Information Center

    Omar, Ahmad Fairuz; MatJafri, Mohd Zubir

    2011-01-01

    The Swift Turbidity Marker is an optical instrument developed to measure the level of water turbidity. The components and configuration selected for the system are based on common turbidity meter design concepts but use a simplified methodology to produce rapid turbidity measurements. This work is aimed at high school physics students and is the…

  5. Glycosylation of serum ribonuclease 1 indicates a major endothelial origin and reveals an increase in core fucosylation in pancreatic cancer.

    PubMed

    Barrabés, Sílvia; Pagès-Pons, Lluís; Radcliffe, Catherine M; Tabarés, Glòria; Fort, Esther; Royle, Louise; Harvey, David J; Moenner, Michel; Dwek, Raymond A; Rudd, Pauline M; De Llorens, Rafael; Peracaula, Rosa

    2007-04-01

    Human pancreatic ribonuclease 1 (RNase 1) is a glycoprotein expressed mainly by the pancreas and also found in endothelial cells. The diagnosis of pancreatic cancer (PaC) remains difficult and therefore the search for sensitive and specific markers is required. Previous studies showed that RNase 1 from human healthy pancreas contained only neutral glycans, whereas RNase 1 from PaC cell lines contained sialylated structures. To determine whether these glycan tumor cell-associated changes were also characteristic of serum RNase 1 and could be used as a marker of PaC, we have analyzed the glycosylation of serum RNase 1. The origin of serum RNase 1 was also investigated. Serum RNase 1 from two PaC patients and two controls was purified and the glycans analyzed by high-performance liquid chromatography (HPLC)-based sequencing and mass spectrometry. Although normal and tumor serum RNase 1 contained the same glycan structures, there was an increase of 40% in core fucosylation in the main sialylated biantennary glycans in the PaC serum RNase 1. This change in proportion would be indicative of a subset of tumor-associated glycoforms of RNase 1, which may provide a biomarker for PaC. Two-dimensional electrophoresis of the RNase 1 from several endothelial cell lines, EA.hy926, human umbilical vein endothelial cells (HUVEC), human mammary microvessel endothelial cells (HuMMEC), and human lung microvessel endothelial cells (HuLEC), showed basically the same pattern and was also very similar to that of serum RNase 1. RNase 1 from EA.hy926 was then purified and presented a glycosylation profile very similar to that from serum RNase 1, suggesting that endothelial cells are the main source of this enzyme.

  6. Vascularization of the embryonic kidney. Detection of endothelial cells with Ulex europaeus I lectin.

    PubMed

    Holthöfer, H

    1987-01-01

    Frozen sections of human fetal kidneys were studied for the appearance of vascular elements to the developing glomeruli using Ulex europaeus (UEA-I) lectin as a marker for endothelial cells. Chains of UEA-I-positive cells, seen to extend from larger vessels, could be observed among the uninduced cells of the nephrogenic mesenchyme. During the S-shaped body stage of nephrogenesis, the cleft of the comma-shaped cluster of epithelial cells was invaded by the UEA-I-positive cells, and during further glomerular development, the UEA-I-positive cells were seen to grow in number, prior to the appearance of visible lumina to the capillaries. The present results show, in addition to revealing the vascularization pattern of developing glomeruli, that endothelial cells obtain some of their characteristic glycoconjugates during early stages of differentiation. Thus, UEA-I seems to be a valuable tool to study in detail the vascularization of various developing human tissues.

  7. Effect of cerium oxide nanoparticles on inflammation in vascular endothelial cells

    PubMed Central

    Gojova, Andrea; Lee, Jun-Tae; Jung, Heejung S.; Guo, Bing; Barakat, Abdul I.; Kennedy, Ian M.

    2010-01-01

    Because vascular endothelial cell inflammation is critical in the development of cardiovascular pathology, we hypothesized that direct exposure of human aortic endothelial cells (HAECs) to ultrafine particles induces an inflammatory response. To test the hypothesis, we incubated HAECs for 4 h with different concentrations (0.001–50 μg/ml) of CeO2 nanoparticles and subsequently measured mRNA levels of the three inflammatory markers intercellular adhesion molecule 1 (ICAM-1), interleukin (IL)-8, and monocyte chemotactic protein (MCP-1) using real-time polymerase chain reaction (PCR). Ceria nanoparticles caused very little inflammatory response in HAECs, even at the highest dose. This material is apparently rather benign in comparison with Y2O3 and ZnO nanoparticles that we have studied previously. These results suggest that inflammation in HAECs following acute exposure to metal oxide nanoparticles depends strongly on particle composition. PMID:19558244

  8. The multiple personality disorder phenotype(s) of circulating endothelial cells in cancer.

    PubMed

    Bertolini, Francesco; Mancuso, Patrizia; Braidotti, Paola; Shaked, Yuval; Kerbel, Robert S

    2009-08-01

    Circulating endothelial cells (CECs) and circulating endothelial progenitors (CEPs) are currently being investigated in a variety of diseases as markers of vascular turnover or damage and, also in the case of CEPs, vasculogenesis. CEPs appear to have a "catalytic" role in different steps of cancer progression and recurrence after therapy, and there are preclinical and clinical data suggesting that CEC enumeration might be useful to select and stratify patients who are candidates for anti-angiogenic treatments. In some types of cancer, CECs and CEPs might be one of the possible hidden identities of cancer stem cells. The definition of CEC and CEP phenotype and the standardization of CEC and CEP enumeration strategies are highly desirable goals in order to exploit these cells as reliable biomarkers in oncology clinical trials.

  9. Quantitative assessment of angiogenesis, perfused blood vessels and endothelial tip cells in the postnatal mouse brain.

    PubMed

    Wälchli, Thomas; Mateos, José María; Weinman, Oliver; Babic, Daniela; Regli, Luca; Hoerstrup, Simon P; Gerhardt, Holger; Schwab, Martin E; Vogel, Johannes

    2015-01-01

    During development and in various diseases of the CNS, new blood vessel formation starts with endothelial tip cell selection and vascular sprout migration, followed by the establishment of functional, perfused blood vessels. Here we describe a method that allows the assessment of these distinct angiogenic steps together with antibody-based protein detection in the postnatal mouse brain. Intravascular and perivascular markers such as Evans blue (EB), isolectin B4 (IB4) or laminin (LN) are used alongside simultaneous immunofluorescence on the same sections. By using confocal laser-scanning microscopy and stereological methods for analysis, detailed quantification of the 3D postnatal brain vasculature for perfused and nonperfused vessels (e.g., vascular volume fraction, vessel length and number, number of branch points and perfusion status of the newly formed vessels) and characterization of sprouting activity (e.g., endothelial tip cell density, filopodia number) can be obtained. The entire protocol, from mouse perfusion to vessel analysis, takes ∼10 d.

  10. Effect of cerium oxide nanoparticles on inflammation in vascular endothelial cells.

    PubMed

    Gojova, Andrea; Lee, Jun-Tae; Jung, Heejung S; Guo, Bing; Barakat, Abdul I; Kennedy, Ian M

    2009-07-01

    Because vascular endothelial cell inflammation is critical in the development of cardiovascular pathology, we hypothesized that direct exposure of human aortic endothelial cells (HAECs) to ultrafine particles induces an inflammatory response. To test the hypothesis, we incubated HAECs for 4 h with different concentrations (0.001-50 microg/ml) of CeO(2) nanoparticles and subsequently measured mRNA levels of the three inflammatory markers intercellular adhesion molecule 1 (ICAM-1), interleukin (IL)-8, and monocyte chemotactic protein (MCP-1) using real-time polymerase chain reaction (PCR). Ceria nanoparticles caused very little inflammatory response in HAECs, even at the highest dose. This material is apparently rather benign in comparison with Y(2)O(3) and ZnO nanoparticles that we have studied previously. These results suggest that inflammation in HAECs following acute exposure to metal oxide nanoparticles depends strongly on particle composition.

  11. Characterization of a distinct population of circulating human non-adherent endothelial forming cells and their recruitment via intercellular adhesion molecule-3.

    PubMed

    Appleby, Sarah L; Cockshell, Michaelia P; Pippal, Jyotsna B; Thompson, Emma J; Barrett, Jeffrey M; Tooley, Katie; Sen, Shaundeep; Sun, Wai Yan; Grose, Randall; Nicholson, Ian; Levina, Vitalina; Cooke, Ira; Talbo, Gert; Lopez, Angel F; Bonder, Claudine S

    2012-01-01

    Circulating vascular progenitor cells contribute to the pathological vasculogenesis of cancer whilst on the other hand offer much promise in therapeutic revascularization in post-occlusion intervention in cardiovascular disease. However, their characterization has been hampered by the many variables to produce them as well as their described phenotypic and functional heterogeneity. Herein we have isolated, enriched for and then characterized a human umbilical cord blood derived CD133(+) population of non-adherent endothelial forming cells (naEFCs) which expressed the hematopoietic progenitor cell markers (CD133, CD34, CD117, CD90 and CD38) together with mature endothelial cell markers (VEGFR2, CD144 and CD31). These cells also expressed low levels of CD45 but did not express the lymphoid markers (CD3, CD4, CD8) or myeloid markers (CD11b and CD14) which distinguishes them from 'early' endothelial progenitor cells (EPCs). Functional studies demonstrated that these naEFCs (i) bound Ulex europaeus lectin, (ii) demonstrated acetylated-low density lipoprotein uptake, (iii) increased vascular cell adhesion molecule (VCAM-1) surface expression in response to tumor necrosis factor and (iv) in co-culture with mature endothelial cells increased the number of tubes, tubule branching and loops in a 3-dimensional in vitro matrix. More importantly, naEFCs placed in vivo generated new lumen containing vasculature lined by CD144 expressing human endothelial cells (ECs). Extensive genomic and proteomic analyses of the naEFCs showed that intercellular adhesion molecule (ICAM)-3 is expressed on their cell surface but not on mature endothelial cells. Furthermore, functional analysis demonstrated that ICAM-3 mediated the rolling and adhesive events of the naEFCs under shear stress. We suggest that the distinct population of naEFCs identified and characterized here represents a new valuable therapeutic target to control aberrant vasculogenesis.

  12. Characterization of a Distinct Population of Circulating Human Non-Adherent Endothelial Forming Cells and Their Recruitment via Intercellular Adhesion Molecule-3

    PubMed Central

    Thompson, Emma J.; Barrett, Jeffrey M.; Tooley, Katie; Sen, Shaundeep; Sun, Wai Yan; Grose, Randall; Nicholson, Ian; Levina, Vitalina; Cooke, Ira; Talbo, Gert; Lopez, Angel F.; Bonder, Claudine S.

    2012-01-01

    Circulating vascular progenitor cells contribute to the pathological vasculogenesis of cancer whilst on the other hand offer much promise in therapeutic revascularization in post-occlusion intervention in cardiovascular disease. However, their characterization has been hampered by the many variables to produce them as well as their described phenotypic and functional heterogeneity. Herein we have isolated, enriched for and then characterized a human umbilical cord blood derived CD133+ population of non-adherent endothelial forming cells (naEFCs) which expressed the hematopoietic progenitor cell markers (CD133, CD34, CD117, CD90 and CD38) together with mature endothelial cell markers (VEGFR2, CD144 and CD31). These cells also expressed low levels of CD45 but did not express the lymphoid markers (CD3, CD4, CD8) or myeloid markers (CD11b and CD14) which distinguishes them from ‘early’ endothelial progenitor cells (EPCs). Functional studies demonstrated that these naEFCs (i) bound Ulex europaeus lectin, (ii) demonstrated acetylated-low density lipoprotein uptake, (iii) increased vascular cell adhesion molecule (VCAM-1) surface expression in response to tumor necrosis factor and (iv) in co-culture with mature endothelial cells increased the number of tubes, tubule branching and loops in a 3-dimensional in vitro matrix. More importantly, naEFCs placed in vivo generated new lumen containing vasculature lined by CD144 expressing human endothelial cells (ECs). Extensive genomic and proteomic analyses of the naEFCs showed that intercellular adhesion molecule (ICAM)-3 is expressed on their cell surface but not on mature endothelial cells. Furthermore, functional analysis demonstrated that ICAM-3 mediated the rolling and adhesive events of the naEFCs under shear stress. We suggest that the distinct population of naEFCs identified and characterized here represents a new valuable therapeutic target to control aberrant vasculogenesis. PMID:23144795

  13. Connexin 43 expressed in endothelial cells modulates monocyte‑endothelial adhesion by regulating cell adhesion proteins.

    PubMed

    Yuan, Dongdong; Sun, Guoliang; Zhang, Rui; Luo, Chenfang; Ge, Mian; Luo, Gangjian; Hei, Ziqing

    2015-11-01

    Adhesion between circulating monocytes and vascular endothelial cells is a key initiator of atherosclerosis. In our previous studies, it was demonstrated that the expression of connexin (Cx)43 in monocytes modulates cell adhesion, however, the effects of the expression of Cx43 in endothelial cells remains to be elucidated. Therefore, the present study investigated the role of the expression of Cx43 in endothelial cells in the process of cell adhesion. A total of four different methods with distinct mechanisms were used to change the function and expression of Cx43 channels in human umbilical vein endothelial cells: Cx43 channel inhibitor (oleamide), enhancer (retinoic acid), overexpression of Cx43 by transfection with pcDNA‑Cx43 and knock‑down of the expression of Cx43 by small interfering RNA against Cx43. The results indicated that the upregulation of the expression of Cx43 enhanced monocyte‑endothelial adhesion and this was markedly decreased by downregulation of Cx43. This mechanism was associated with Cx43‑induced expression of vascular cell adhesion molecule‑1 and intercellular cell adhesion molecule‑1. The effects of Cx43 in endothelial cells was independent of Cx37 or Cx40. These experiments suggested that local regulation of endothelial Cx43 expression within the vasculature regulates monocyte‑endothelial adhesion, a critical event in the development of atherosclerosis and other inflammatory pathologies, with baseline adhesion set by the expression of Cx43. This balance may be crucial in controlling leukocyte involvement in inflammatory cascades.

  14. Developing a biohybrid lung - sufficient endothelialization of poly-4-methly-1-pentene gas exchange hollow-fiber membranes.

    PubMed

    Wiegmann, Bettina; von Seggern, Heide; Höffler, Klaus; Korossis, Sotirios; Dipresa, Daniele; Pflaum, Michael; Schmeckebier, Sabrina; Seume, Jörg; Haverich, Axel

    2016-07-01

    Working towards establishing a biohybrid lung with optimized hemocompatibility, this study analyzed the feasibility of establishing flow-resistant endothelium on heparin/albumin coated poly-4-methly-1-pentene hollow fiber gas exchange membranes (PMP-HFs). The seeding efficiency and proliferation of human cord blood derived endothelial cells (HCBEC) on PMP-HFs were analyzed under static conditions by WST-8 cell proliferation assay and fluorescence microscopy. The HCBEC monolayer integrity under different flow conditions was also assessed. Endothelial-specific phenotype verification, expression activation levels and thrombogenic state markers were quantified by real-time RT-PCR for cell-to-PMP-HF contact under static and dynamic conditions. The results demonstrated the feasibility of establishing a viable, confluent, and flow-resistant endothelial monolayer on the blood-contact surface of PMP-HFs, which maintained a physiological response to TNFα-stimulation and flow conditions. The endothelial phenotype, expression levels of adhesion molecules and thrombogenic state markers were unaffected by cell-to-PMP-HFs contact. These results represent a significant step towards establishing a biohybrid lung.

  15. Monocyte-derived microparticles and exosomes induce procoagulant and apoptotic effects on endothelial cells.

    PubMed

    Aharon, Anat; Tamari, Tal; Brenner, Benjamin

    2008-11-01

    Microvesicles (MVs) which include microparticles (MPs) and exosomes are found in blood circulation in normal physiologic conditions and are increased in a variety of diseases. This study evaluated the effects of MVs on human umbilical vein endothelial cells (HUVEC) by morphologic changes, apoptosis, and thrombogenicty, in vitro. Stimulation of monocyte cell line (THP-1) by starvation or by endotoxin and calcium ionophore A23187 resulted in the release of MVs which express exosome marker Tsg 101, negative phospholipids in their leaflets, monocyte markers (CD18, CD14) and active tissue factor (TF). MVs were found to disrupt EC integrity and rapidly induce membrane blebbing. Brief exposure (2-4 hours) to MVs resulted in EC membrane phospholipids "flip-flop" while longer stimulation (20 hours) led to two contradicting outcomes - tube formation as well as apoptosis, as assessed by nuclear fragmentation. Additionally, MVs exposure resulted in increased cell surface thrombogenicity and perturbation of the endothelial haemostatic balance, which were enhanced during longer exposure time. Activity, antigen level and mRNA expression of the coagulation initiator TF were elevated due to (i) adherence of MVs derived TF to the EC membrane, and (ii) an increase in endothelial TF expression. Furthermore, levels of the anticoagulant tissue factor pathway inhibitor (TFPI) and thrombomodulin (TM) were decreased. These findings demonstrate that monocyte MVs increase endothelial thrombogenicity and apoptosis. In addition, they induce tube formation which may indicate their angiogenic effect. These findings may clarify, in part, the role of MVs in EC dysfunction associated with inflammatory diseases and hypercoagulable states.

  16. Exclusion of histiocytes/endothelial cells and using endothelial cells as internal reference are crucial for interpretation of MGMT immunohistochemistry in glioblastoma.

    PubMed

    Hsu, Chih-Yi; Lin, Shih-Chieh; Ho, Hsiang-Ling; Chang-Chien, Yi-Chun; Hsu, Sanford P-C; Yen, Yu-Shu; Chen, Ming-Hsiung; Guo, Wan-You; Ho, Donald M-T

    2013-02-01

    We evaluated the predictive value of O6-methylguanine-DNA methyltransferase (MGMT) protein expression and MGMT promoter methylation status in glioblastomas (GBM) treated with temozolomide (TMZ) in a Taiwan medical center. Protein expression by immunohistochemical analysis (IHC) and MGMT promoter methylation detected by methylation-specific polymerase chain reaction (MSP) were performed in a series of 107 newly diagnosed GBMs. We used endothelial cells as an internal reference for IHC staining because the staining intensities of the MGMT-expressing cells in different specimens varied considerably; a positive result was defined as the staining intensity of the majority of tumor cells similar to that of the adjacent endothelial cells. Immunostainings for microglial/endothelial markers were included as part of the MGMT IHC evaluation, and in cases that were difficult to interpret, double-labeling helped to clarify the nature of reactive cells. The MGMT protein expression was reversely associated with MGMT promoter methylation status in 83.7% of cases (MSP/IHC and MSP/IHC; Pearson r=-0.644, P<0.001). Twenty-two of 24 (91.7%) IHC tumors did not respond to TMZ treatment. Combining MSP and IHC results, all the 15 MSP/IHC GBMs were TMZ resistant. The MGMT status detected by either IHC or MSP was significantly correlated with the TMZ treatment response (both P<0.001) and survival of GBM patients (both P<0.05).

  17. Referential Markers and Agreement Markers in Functional Discourse Grammar

    ERIC Educational Resources Information Center

    Hengeveld, Kees

    2012-01-01

    It follows from the ordering principles that are applied in Functional Discourse Grammar that the positional possibilities of markers of agreement and those of cross-reference are different. Markers of cross reference are predicted to occur closer to the verb stem, while markers of agreement would occupy peripheral positions. This paper tests…

  18. Vitamin D improves endothelial dysfunction and restores myeloid angiogenic cell function via reduced CXCL-10 expression in systemic lupus erythematosus

    PubMed Central

    Reynolds, John A.; Haque, Sahena; Williamson, Kate; Ray, David W.; Alexander, M. Yvonne; Bruce, Ian N.

    2016-01-01

    Patients with systemic lupus erythematosus (SLE) have accelerated cardiovascular disease and dysfunctional endothelial repair mechanisms. Myeloid angiogenic cells (MACs), derived from circulating monocytes, augment vascular repair by paracrine secretion of pro-angiogenic factors. We observed that SLE MACs are dysfunctional and secrete pro-inflammatory cytokines. We also found that the vitamin D receptor was transiently expressed during MAC differentiation and that in vitro, calcitriol increased differentiation of monocytes into MACs in both SLE and in a model using the prototypic SLE cytokine, interferon-alpha. The active form of vitamin D (calcitriol) restored the SLE MAC phenotype towards that of healthy subjects with reduced IL-6 secretion, and normalised surface marker expression. Calcitriol also augmented the angiogenic capacity of MACs via the down-regulation of CXCL-10. In SLE patients treated with cholecalciferol for 12 weeks, the improvement in endothelial function correlated with increase in serum 25(OH)D concentrations independently of disease activity. We also show that MACs were able to positively modulate eNOS expression in human endothelial cells in vitro, an effect further enhanced by calcitriol treatment of SLE MACs. The results demonstrate that vitamin D can positively modify endothelial repair mechanisms and thus endothelial function in a population with significant cardiovascular risk. PMID:26930567

  19. Lipoprotein marker for hypertriglyceridemia

    DOEpatents

    Cubicciotti, Roger S.; Karu, Alexander E.; Krauss, Ronald M.

    1986-01-01

    Methods and compositions are provided for the detection of a particular low density lipoprotein which has been found to be a marker for patients suffering from type IV hypertriglyceridemia. A monoclonal antibody capable of specifically binding to a characteristic epitopic site on this LDL subspecies can be utilized in a wide variety of immunoassays. Hybridoma cell line SPL.IVA5A1 was deposited at the American Type Culture Collection on Mar. 29, 1984, and granted accession no. HB 8535.

  20. Heterogeneity in immunohistochemical, genomic, and biological properties of human lymphatic endothelial cells between initial and collecting lymph vessels.

    PubMed

    Kawai, Yoshiko; Hosaka, Kayoko; Kaidoh, Maki; Minami, Takashi; Kodama, Tatsuhiko; Ohhashi, Toshio

    2008-01-01

    The immunohistochemical properties of selective lymph vessel markers, and NO synthase (NOS) and cyclo-oxygenase (COX) activities, were examined in two kinds of human lymphatic endothelial cells isolated from collecting (macro-) and initial (micro-) lymph vessels. The constitutively expressed genes in the two kinds of lymphatic endothelial cells were also evaluated by using oligonucleotide microarray analysis and RT-PCR. We also investigated the effects of oxygen concentration in culture conditions or growth factors such as basic fibroblast growth factor (bFGF), VEGF-A, and VEGF-C on proliferation activities of the two kinds of human lymphatic endothelial cells. Immunoreactivity to LYVE-1 and the RT-PCR expression level of LYVE-1 mRNA in endothelial cells of micro-lymph vessels were stronger than those of macro-lymph vessels. Immunoreactivity to VEGF R1 was also observed as significantly stronger in the micro-lymph vessels. In contrast, the immunoreactivity to Prox-1 and the RT-PCR expression level of Prox-1 mRNA in endothelial cells of macro-lymph vessels were stronger than those of micro-lymph vessels. Similarly, immunoreactivity to ecNOS, iNOS, COX1, and COX2 was also found as significantly higher than in macro-lymph vessels. In contrast, the increase of O(2) concentration ranging from 5% to 21% caused a significant reduction of the proliferation activity of endothelial cells in macro-lymph vessels. In conclusion, these findings suggest marked heterogeneity in the immunohistochemical, genomic, and proliferation activity of human lymphatic endothelial cells between micro-(initial) and macro-(collecting) lymph vessels.

  1. Serum tumor markers.

    PubMed

    Perkins, Greg L; Slater, Evan D; Sanders, Georganne K; Prichard, John G

    2003-09-15

    Monoclonal antibodies are used to detect serum antigens associated with specific malignancies. These tumor markers are most useful for monitoring response to therapy and detecting early relapse. With the exception of prostate-specific antigen (PSA), tumor markers do not have sufficient sensitivity or specificity for use in screening. Cancer antigen (CA) 27.29 most frequently is used to follow response to therapy in patients with metastatic breast cancer. Carcinoembryonic antigen is used to detect relapse of colorectal cancer, and CA 19-9 may be helpful in establishing the nature of pancreatic masses. CA 125 is useful for evaluating pelvic masses in postmenopausal women, monitoring response to therapy in women with ovarian cancer, and detecting recurrence of this malignancy. Alpha-fetoprotein (AFP), a marker for hepatocellular carcinoma, sometimes is used to screen highly selected populations and to assess hepatic masses in patients at particular risk for developing hepatic malignancy. Testing for the beta subunit of human chorionic gonadotropin (beta-hCG) is an integral part of the diagnosis and management of gestational trophoblastic disease. Combined AFP and beta-hCG testing is an essential adjunct in the evaluation and treatment of nonseminomatous germ cell tumors, and in monitoring the response to therapy. AFP and beta-hCG also may be useful in evaluating potential origins of poorly differentiated metastatic cancer. PSA is used to screen for prostate cancer, detect recurrence of the malignancy, and evaluate specific syndromes of adenocarcinoma of unknown primary.

  2. Parasite biomass-related inflammation, endothelial activation, microvascular dysfunction and disease severity in vivax malaria.

    PubMed

    Barber, Bridget E; William, Timothy; Grigg, Matthew J; Parameswaran, Uma; Piera, Kim A; Price, Ric N; Yeo, Tsin W; Anstey, Nicholas M

    2015-01-01

    Plasmodium vivax can cause severe malaria, however its pathogenesis is poorly understood. In contrast to P. falciparum, circulating vivax parasitemia is low, with minimal apparent sequestration in endothelium-lined microvasculature, and pathogenesis thought unrelated to parasite biomass. However, the relationships between vivax disease-severity and total parasite biomass, endothelial autocrine activation and microvascular dysfunction are unknown. We measured circulating parasitemia and markers of total parasite biomass (plasma parasite lactate dehydrogenase [pLDH] and PvLDH) in adults with severe (n = 9) and non-severe (n = 53) vivax malaria, and examined relationships with disease-severity, endothelial activation, and microvascular function. Healthy controls and adults with non-severe and severe falciparum malaria were enrolled for comparison. Median peripheral parasitemia, PvLDH and pLDH were 2.4-fold, 3.7-fold and 6.9-fold higher in severe compared to non-severe vivax malaria (p = 0.02, p = 0.02 and p = 0.015, respectively), suggesting that, as in falciparum malaria, peripheral P. vivax parasitemia underestimates total parasite biomass, particularly in severe disease. P. vivax schizonts were under-represented in peripheral blood. Severe vivax malaria was associated with increased angiopoietin-2 and impaired microvascular reactivity. Peripheral vivax parasitemia correlated with endothelial activation (angiopoietin-2, von-Willebrand-Factor [VWF], E-selectin), whereas markers of total vivax biomass correlated only with systemic inflammation (IL-6, IL-10). Activity of the VWF-cleaving-protease, ADAMTS13, was deficient in proportion to endothelial activation, IL-6, thrombocytopenia and vivax disease-severity, and associated with impaired microvascular reactivity in severe disease. Impaired microvascular reactivity correlated with lactate in severe vivax malaria. Findings suggest that tissue accumulation of P. vivax may occur, with the hidden

  3. Parasite Biomass-Related Inflammation, Endothelial Activation, Microvascular Dysfunction and Disease Severity in Vivax Malaria

    PubMed Central

    Barber, Bridget E.; William, Timothy; Grigg, Matthew J.; Parameswaran, Uma; Piera, Kim A.; Price, Ric N.; Yeo, Tsin W.; Anstey, Nicholas M.

    2015-01-01

    Plasmodium vivax can cause severe malaria, however its pathogenesis is poorly understood. In contrast to P. falciparum, circulating vivax parasitemia is low, with minimal apparent sequestration in endothelium-lined microvasculature, and pathogenesis thought unrelated to parasite biomass. However, the relationships between vivax disease-severity and total parasite biomass, endothelial autocrine activation and microvascular dysfunction are unknown. We measured circulating parasitemia and markers of total parasite biomass (plasma parasite lactate dehydrogenase [pLDH] and PvLDH) in adults with severe (n = 9) and non-severe (n = 53) vivax malaria, and examined relationships with disease-severity, endothelial activation, and microvascular function. Healthy controls and adults with non-severe and severe falciparum malaria were enrolled for comparison. Median peripheral parasitemia, PvLDH and pLDH were 2.4-fold, 3.7-fold and 6.9-fold higher in severe compared to non-severe vivax malaria (p = 0.02, p = 0.02 and p = 0.015, respectively), suggesting that, as in falciparum malaria, peripheral P. vivax parasitemia underestimates total parasite biomass, particularly in severe disease. P. vivax schizonts were under-represented in peripheral blood. Severe vivax malaria was associated with increased angiopoietin-2 and impaired microvascular reactivity. Peripheral vivax parasitemia correlated with endothelial activation (angiopoietin-2, von-Willebrand-Factor [VWF], E-selectin), whereas markers of total vivax biomass correlated only with systemic inflammation (IL-6, IL-10). Activity of the VWF-cleaving-protease, ADAMTS13, was deficient in proportion to endothelial activation, IL-6, thrombocytopenia and vivax disease-severity, and associated with impaired microvascular reactivity in severe disease. Impaired microvascular reactivity correlated with lactate in severe vivax malaria. Findings suggest that tissue accumulation of P. vivax may occur, with the hidden

  4. Systemic inflammation, endothelial dysfunction, and activation in clinically healthy children exposed to air pollutants.

    PubMed

    Calderón-Garcidueñas, L; Villarreal-Calderon, R; Valencia-Salazar, G; Henríquez-Roldán, C; Gutiérrez-Castrellón, P; Torres-Jardón, R; Osnaya-Brizuela, N; Romero, L; Torres-Jardón, R; Solt, A; Reed, W

    2008-03-01

    Mexico City children are chronically exposed to significant concentrations of air pollutants and exhibit chronic respiratory-tract inflammation. Epidemiological, controlled human exposures, laboratory-based animal models, and in vitro/in vivo studies have shown that inflammatory, endothelial dysfunction, and endothelial damage mediators are upregulated upon exposure to particulate matter (PM). Endothelial dysfunction is a critical event in cardiovascular disease. The focus of this work was to investigate whether exposure to ambient air pollution including PM(2.5) produces systemic inflammation and endothelial injury in healthy children. We measured markers of endothelial activation, and inflammatory mediators in 52 children age 8.6+/-0.1 yr, residents of Mexico City (n: 28) or of Polotitlán (n: 24), a city with low levels of pollutants. Mexico City children had significant increases in inflammatory mediators and vasoconstrictors, including tumor necrosis factor (TNF)alpha, prostaglandin (PG) E2, C-reactive protein, interleukin-1beta, and endothelin-1. There was a significant anti-inflammatory response, and a downregulation of vascular adhesion molecule-1, intercellular adhesion molecule-1 and -2, and selectins sE and sL. Results from linear regression found TNF a positively associated with 24- and 48-h cumulative levels of PM(2.5), while the 7-d PM(2.5) value was negatively associated with the numbers of white blood cells in peripheral blood in highly exposed children. Systemic subclinical inflammation, increased endothelin- 1, and significant downregulation of soluble adhesion molecules are seen in Mexico City children. Children chronically exposed to fine PM above the standard could be at risk of developing cardiovascular diseases, atherosclerosis, stroke, and other systemic effects later in life.

  5. Endothelial cell adhesion molecule CD146: implications for its role in the pathogenesis of COPD.

    PubMed

    Kratzer, Adelheid; Chu, Hong Wei; Salys, Jonas; Moumen, Zakaria; Leberl, Maike; Bowler, Russ; Cool, Carlyne; Zamora, Martin; Taraseviciene-Stewart, Laima

    2013-08-01

    CD146 is an adhesion molecule localized at endothelial cell junctions and facilitates cell-cell interactions. The circulating soluble form (sCD146) lacks both the intracellular and the transmembrane domains. In this study we show that CD146 expression was significantly decreased in the lung tissue of smokers with chronic obstructive pulmonary disease (COPD) and also in rats exposed to second-hand smoke (SHS). Concurrently, levels of sCD146 were increased in both the plasma and bronchoalveolar lavage fluid (BALF) of COPD patients as well as in BALF from rats exposed to SHS. Decreased or abolished CD146 protein expression in rat pulmonary micro- and macrovascular endothelial cells was found after treatment with cigarette smoke extract (CSE), proinflammatory cytokine interleukin 18 (IL-18) or after silencing CD146 expression with siRNA. The decrease in CD146 protein was accompanied by increased endothelial monolayer permeability and enhanced macrophage infiltration in vitro. In CD146 knockout (KO) mice, distinct perivascular oedema was seen and increased numbers of inflammatory cells, along with increased protein levels in BALF. Increased sCD146 was found in BALF and plasma from patients with COPD. The circulating plasma levels of sCD146 correlated positively with the presence of anti-endothelial cell antibodies (AECAs). sCD146 in combination with AECAs may be useful markers for early detection of COPD. Our study indicates that loss of CD146 function damages pulmonary endothelial integrity. This damage may represent part of the pathophysiological processes that are involved in the basic aetiology of COPD/emphysema.

  6. IL-4 Causes Hyperpermeability of Vascular Endothelial Cells through Wnt5A Signaling

    PubMed Central

    Skaria, Tom; Burgener, Julia; Bachli, Esther; Schoedon, Gabriele

    2016-01-01

    Microvascular leakage due to endothelial barrier dysfunction is a prominent feature of T helper 2 (Th2) cytokine mediated allergic inflammation. Interleukin-4 (IL-4) is a potent Th2 cytokine, known to impair the barrier function of endothelial cells. However, the effectors mediating IL-4 induced cytoskeleton remodeling and consequent endothelial barrier dysfunction remain poorly defined. Here we have used whole genome transcriptome profiling and gene ontology analyses to identify the genes and processes regulated by IL-4 signaling in human coronary artery endothelial cells (HCAEC). The study revealed Wnt5A as an effector that can mediate actin cytoskeleton remodeling in IL-4 activated HCAEC through the regulation of LIM kinase (LIMK) and Cofilin (CFL). Following IL-4 treatment, LIMK and CFL were phosphorylated, thereby indicating the possibility of actin stress fiber formation. Imaging of actin showed the formation of stress fibers in IL-4 treated live HCAEC. Stress fiber formation was notably decreased in the presence of Wnt inhibitory factor 1 (WIF1). Non-invasive impedance measurements demonstrated that IL-4 increased the permeability and impaired the barrier function of HCAEC monolayers. Silencing Wnt5A significantly reduced permeability and improved the barrier function of HCAEC monolayers upon IL-4 treatment. Our study identifies Wnt5A as a novel marker of IL-4 activated vascular endothelium and demonstrates a critical role for Wnt5A in mediating IL-4 induced endothelial barrier dysfunction. Wnt5A could be a potential therapeutic target for reducing microvascular leakage and edema formation in Th2 driven inflammatory diseases. PMID:27214384

  7. Weight loss improves endothelial function independently of ADMA reduction in severe obesity.

    PubMed

    Rudofsky, G; Roeder, E; Merle, T; Hildebrand, M; Nawroth, P P; Wolfrum, C

    2011-05-01

    This prospective study was performed in order to establish whether improvement of endothelial function after weight reduction can be explained by a decrease of elevated asymmetric dimethyl arginine (ADMA), an inhibitor of endogenous NO-synthase (eNOS). Therefore, 21 obese subjects (BMI: 41.1±6.4 kg/m(2)) were studied at baseline and after 12 weeks of weight reduction with a very low calorie diet. Biochemical and clinical parameters of endothelial function were assessed before and after weight loss. Biochemical parameters were determined by measurement of ADMA and soluble intercellular adhesion molecule (sICAM). Clinical parameters were assessed by pulse wave analysis (PWA). Weight intervention resulted in a 21.4±6.8 kg reduction of body weight from 119.7±12.8 kg at study start to 98.3±11.6 kg at study end (p<0.001). Accordingly, biochemical markers improved under weight reduction (ADMA from 0.47±0.07 mmol/l to 0.42±0.08 mmol/l; p=0.002; ICAM from 276±42 ng/ml to 236±29 ng/ml; p<0.001). Further, clinical parameters of functional endothelial function improved with an increase of deltaRI after salbutamol inhalation from -1% before to -9% after weight reduction (p=0.02). Interestingly, improvement of endothelial function correlated with improved HOMA index only (r=-0.60, p=0.04) but not with reduced ADMA levels, improved hypertension or reduced body weight. In conclusion, weight reduction with a very low calorie diet improves endothelial function measured by pulse wave velocity. The missing correlation with ADMA suggests possible further mechanisms underlying this observed effect, for example, improvement of insulin resistance.

  8. Dietary phosphorus acutely impairs endothelial function.

    PubMed

    Shuto, Emi; Taketani, Yutaka; Tanaka, Rieko; Harada, Nagakatsu; Isshiki, Masashi; Sato, Minako; Nashiki, Kunitaka; Amo, Kikuko; Yamamoto, Hironori; Higashi, Yukihito; Nakaya, Yutaka; Takeda, Eiji

    2009-07-01

    Excessive dietary phosphorus may increase cardiovascular risk in healthy individuals as well as in patients with chronic kidney disease, but the mechanisms underlying this risk are not completely understood. To determine whether postprandial hyperphosphatemia may promote endothelial dysfunction, we investigated the acute effect of phosphorus loading on endothelial function in vitro and in vivo. Exposing bovine aortic endothelial cells to a phosphorus load increased production of reactive oxygen species, which depended on phosphorus influx via sodium-dependent phosphate transporters, and decreased nitric oxide production via inhibitory phosphorylation of endothelial nitric oxide synthase. Phosphorus loading inhibited endothelium-dependent vasodilation of rat aortic rings. In 11 healthy men, we alternately served meals containing 400 mg or 1200 mg of phosphorus in a double-blind crossover study and measured flow-mediated dilation of the brachial artery before and 2 h after the meals. The high dietary phosphorus load increased serum phosphorus at 2 h and significantly decreased flow-mediated dilation. Flow-mediated dilation correlated inversely with serum phosphorus. Taken together, these findings suggest that endothelial dysfunction mediated by acute postprandial hyperphosphatemia may contribute to the relationship between serum phosphorus level and the risk for cardiovascular morbidity and mortality.

  9. Endothelial cells and the IGF system.

    PubMed

    Bach, Leon A

    2015-02-01

    Endothelial cells line blood vessels and modulate vascular tone, thrombosis, inflammatory responses and new vessel formation. They are implicated in many disease processes including atherosclerosis and cancer. IGFs play a significant role in the physiology of endothelial cells by promoting migration, tube formation and production of the vasodilator nitric oxide. These actions are mediated by the IGF1 and IGF2/mannose 6-phosphate receptors and are modulated by a family of high-affinity IGF binding proteins. IGFs also increase the number and function of endothelial progenitor cells, which may contribute to protection from atherosclerosis. IGFs promote angiogenesis, and dysregulation of the IGF system may contribute to this process in cancer and eye diseases including retinopathy of prematurity and diabetic retinopathy. In some situations, IGF deficiency appears to contribute to endothelial dysfunction, whereas IGF may be deleterious in others. These differences may be due to tissue-specific endothelial cell phenotypes or IGFs having distinct roles in different phases of vascular disease. Further studies are therefore required to delineate the therapeutic potential of IGF system modulation in pathogenic processes.

  10. Extensive Characterization and Comparison of Endothelial Cells Derived from Dermis and Adipose Tissue: Potential Use in Tissue Engineering

    PubMed Central

    Monsuur, Hanneke N.; Weijers, Ester M.; Niessen, Frank B.; Gefen, Amit; Koolwijk, Pieter; Gibbs, Susan; van den Broek, Lenie J.

    2016-01-01

    Tissue-engineered constructs need to become quickly vascularized in order to ensure graft take. One way of achieving this is to incorporate endothelial cells (EC) into the construct. The adipose tissue stromal vascular fraction (adipose-SVF) might provide an alternative source for endothelial cells as adipose tissue can easily be obtained by liposuction. Since adipose-EC are now gaining more interest in tissue engineering, we aimed to extensively characterize endothelial cells from adipose tissue (adipose-EC) and compare them with endothelial cells from dermis (dermal-EC). The amount of endothelial cells before purification varied between 4–16% of the total stromal population. After MACS selection for CD31 positive cells, a >99% pure population of endothelial cells was obtained within two weeks of culture. Adipose- and dermal-EC expressed the typical endothelial markers PECAM-1, ICAM-1, Endoglin, VE-cadherin and VEGFR2 to a similar extent, with 80–99% of the cell population staining positive. With the exception of CXCR4, which was expressed on 29% of endothelial cells, all other chemokine receptors (CXCR1, 2, 3, and CCR2) were expressed on less than 5% of the endothelial cell populations. Adipose-EC proliferated similar to dermal-EC, but responded less to the mitogens bFGF and VEGF. A similar migration rate was found for both adipose-EC and dermal-EC in response to bFGF. Sprouting of adipose-EC and dermal-EC was induced by bFGF and VEGF in a 3D fibrin matrix. After stimulation of adipose-EC and dermal-EC with TNF-α an increased secretion was seen for PDGF-BB, but not uPA, PAI-1 or Angiopoietin-2. Furthermore, secretion of cytokines and chemokines (IL-6, CCL2, CCL5, CCL20, CXCL1, CXCL8 and CXCL10) was also upregulated by both adipose- and dermal-EC. The similar characteristics of adipose-EC compared to their dermal-derived counterpart make them particularly interesting for skin tissue engineering. In conclusion, we show here that adipose tissue provides for an

  11. Effect of selected flavones on cancer and endothelial cells.

    PubMed

    Pilátová, Martina; Stupáková, Viktória; Varinská, Lenka; Sarisský, Marek; Mirossay, Ladislav; Mirossay, Andrej; Gál, Peter; Kraus, Vladimír; Dianisková, Katarína; Mojzis, Ján

    2010-06-01

    In our study we used quercetin (3,3 ,4 ,5,7-pentahydroxyflavone) as the reference standard to compare antiproliferative and antiangiogenic effects of chrysin (5,7-dihydroxyflavone) and 3-hydroxyflavone. Our data indicates that chrysin and 3-hydroxyflavone showed significantly higher cytotoxic effect than reference standard quercetin. These tested agents significantly decreased cell survival with the efficacy of 65-85% at the concentration 100 micromol/l for HUVEC, lung carcinoma and leukemic cells being the most sensitive. Cell cycle analysis indicates that quercetin and 3-hydroxyflavone might affect the cell cycle of Jurkat cells by a similar or the same mechanism of action which lead to G2/M arrest as well as to an increase in sub-G0/G1 fraction. Treatment of Jurkat cells with chrysin resulted only increase in the fraction of cells with sub-G0/G1 DNA content, which is considered to be a marker of apoptotic cell death. Apoptosis was confirmed by DNA fragmentation and by staining with annexin V. All three tested flavones inhibited endothelial cell migration after 24 h of incubation at a concentration 100 micromol/l. At a lower concentration (10 micromol/l) only quercetin significantly inhibited migration of endothelial cells. Furthermore, in our experiments decreased secretion of matrix metalloproteinases (MMP-2 and MMP-9) was observed after a 72 h treatment with quercetin. No decrease in secretion of MMP-2 and MMP-9 was seen after chrysin and 3-hydroxyflavone treatment. On the other hand, our results showed that none of three flavonoids blocked microcapillary tube formation. Further studies are necessary to investigate the mechanism of action and to find out the relationship between the structure, character and position of substituents of natural substances and their biological activities.

  12. Tesmilifene modifies brain endothelial functions and opens the blood-brain/blood-glioma barrier.

    PubMed

    Walter, Fruzsina R; Veszelka, Szilvia; Pásztói, Mária; Péterfi, Zoltán A; Tóth, András; Rákhely, Gábor; Cervenak, László; Ábrahám, Csongor S; Deli, Mária A

    2015-09-01

    Tesmilifene, a tamoxifen analog with antihistamine action, has chemopotentiating properties in experimental and clinical cancer studies. In our previous works, tesmilifene increased the permeability of the blood-brain barrier (BBB) in animal and culture models. Our aim was to investigate the effects of tesmilifene on brain microvessel permeability in the rat RG2 glioma model and to reveal its mode of action in brain endothelial cells. Tesmilifene significantly increased fluorescein extravasation in the glioma. Short-term treatment with tesmilifene reduced the resistance and increased the permeability for marker molecules in a rat triple co-culture BBB model. Tesmilifene also affected the barrier integrity in brain endothelial cells co-cultured with RG2 glioblastoma cells. Tesmilifene inhibited the activity of P-glycoprotein and multidrug resistance-associated protein-1 efflux pumps and down-regulated the mRNA expression of tight junction proteins, efflux pumps, solute carriers, and metabolic enzymes important for BBB functions. Among the possible signaling pathways that regulate BBB permeability, tesmilifene activated the early nuclear translocation of NFκB. The MAPK/ERK and PI3K/Akt kinase pathways were also involved. We demonstrate for the first time that tesmilifene increases permeability marker molecule extravasation in glioma and inhibits efflux pump activity in brain endothelial cells, which may have therapeutic relevance. Tesmilifene, a chemopotentiator in experimental and clinical cancer studies increases vascular permeability in RG2 glioma in rats and permeability for marker molecules in a culture model of the blood-brain barrier. Tesmilifene inhibits the activity of efflux pumps and down-regulates the mRNA expression of tight junction proteins, transporters, and metabolic enzymes important for the blood-brain barrier functions, which may have therapeutic relevance.

  13. Endothelial damage in major depression patients is modulated by SSRI treatment, as demonstrated by circulating biomarkers and an in vitro cell model

    PubMed Central

    Lopez-Vilchez, I; Diaz-Ricart, M; Navarro, V; Torramade, S; Zamorano-Leon, J; Lopez-Farre, A; Galan, A M; Gasto, C; Escolar, G

    2016-01-01

    There is a link between depression, cardiovascular events and inflammation. We have explored this connection through endothelial dysfunction, using in vivo and in vitro approaches. We evaluated circulating biomarkers of endothelial dysfunction in patients with major depression at their diagnosis (MD-0) and during antidepressant treatment with the selective serotonin reuptake inhibitor escitalopram, for 8 and 24 weeks (MD-8 and MD-24). Results were always compared with matched healthy controls (CON). We measured in vivo circulating endothelial cells (CECs) and endothelial progenitor cells (EPCs) in blood samples, and assessed plasma levels of soluble von Willebrand factor (VWF) and vascular cell adhesion molecule-1 (VCAM-1). CEC counts, soluble VWF and VCAM-1 were statistically elevated in MD-0 (P<0.01 versus CON) and gradually decreased during treatment. Conversely, EPC levels were lower in MD-0, tending to increase throughout treatment. In vitro studies were performed in human endothelial cells cultured in the presence of sera from each study group. Elevated expression of the inflammation marker intercellular adhesion molecule-1 and oxidative stress, with lower presence of endothelial nitric oxide synthase and higher reactive oxygen species production, were found in cells exposed to MD-0 sera (P<0.05 versus CON). These results were normalized in cells exposed to MD-24 sera. Thrombogenicity of extracellular matrices generated by these cells, measured as expression of VWF, tissue factor and platelet reactivity, showed non-significant differences. We provide a model of cultured endothelial cells reproducing endothelial dysfunction in naive patients with major depression, demonstrating endothelial damage and inflammation at diagnosis, and recovering with selective serotonin reuptake inhibitor treatment for 24 weeks. PMID:27598970

  14. Glassy Dynamics, Cell Mechanics and Endothelial Permeability

    PubMed Central

    Hardin, Corey; Rajendran, Kavitha; Manomohan, Greeshma; Tambe, Dhananjay T.; Butler, James P.; Fredberg, Jeffrey J.; Martinelli, Roberta; Carman, Christopher V.; Krishnan, Ramaswamy

    2013-01-01

    A key feature of all inflammatory processes is disruption of the vascular endothelial barrier. Such disruption is initiated in part through active contraction of the cytoskeleton of the endothelial cell (EC). Because contractile forces are propagated from cell to cell across a great many cell-cell junctions, this contractile process is strongly cooperative and highly nonlocal. We show here that the characteristic length scale of propagation is modulated by agonists and antagonists that impact permeability of the endothelial barrier. In the presence of agonists including thrombin, histamine, and H202, force correlation length increases, whereas in the presence of antagonists including sphingosine-1-phosphate, hepatocyte growth factor, and the rho kinase inhibitor, Y27632, force correlation length decreases. Intercellular force chains and force clusters are also evident, both of which are reminiscent of soft glassy materials approaching a glass transition. PMID:23638866

  15. Endothelial α3β1-Integrin Represses Pathological Angiogenesis and Sustains Endothelial-VEGF

    PubMed Central

    da Silva, Rita Graça; Tavora, Bernardo; Robinson, Stephen D.; Reynolds, Louise E.; Szekeres, Charles; Lamar, John; Batista, Sílvia; Kostourou, Vassiliki; Germain, Mitchel A.; Reynolds, Andrew R.; Jones, Dylan T.; Watson, Alan R.; Jones, Janet L.; Harris, Adrian; Hart, Ian R.; Iruela-Arispe, M. Luisa; DiPersio, C. Michael; Kreidberg, Jordan A.; Hodivala-Dilke, Kairbaan M.

    2010-01-01

    Integrin α3β1 is a major receptor for laminin. The expression levels of laminins-8 and -10 in the basement membrane surrounding blood vessels are known to change during tumor angiogenesis. Although some studies have suggested that certain ligands of α3β1 can affect angiogenesis either positively or negatively, either a direct in vivo role for α3β1 in this process or its mechanism of action in endothelial cells during angiogenesis is still unknown. Because the global genetic ablation of α3-integrin results in an early lethal phenotype, we have generated conditional-knockout mice where α3 is deleted specifically in endothelial cells (ec-α3−/−). Here we show that ec-α3−/− mice are viable, fertile, and display enhanced tumor growth, elevated tumor angiogenesis, augmented hypoxia-induced retinal angiogenesis, and increased vascular endothelial growth factor (VEGF)-mediated neovascularization ex vivo and in vivo. Furthermore, our data provide a novel method by which an integrin may regulate angiogenesis. We show that α3β1 is a positive regulator of endothelial-VEGF and that, surprisingly, the VEGF produced by endothelial cells can actually repress VEGF-receptor 2 (Flk-1) expression. These data, therefore, identify directly that endothelial α3β1 negatively regulates pathological angiogenesis and implicate an unexpected role for low levels of endothelial-VEGF as an activator of neovascularization. PMID:20639457

  16. Apoptosis of Endothelial Cells by 13-HPODE Contributes to Impairment of Endothelial Barrier Integrity

    PubMed Central

    Ryman, Valerie E.; Packiriswamy, Nandakumar

    2016-01-01

    Inflammation is an essential host response during bacterial infections such as bovine mastitis. Endothelial cells are critical for an appropriate inflammatory response and loss of vascular barrier integrity is implicated in the pathogenesis of Streptococcus uberis-induced mastitis. Previous studies suggested that accumulation of linoleic acid (LA) oxygenation products derived from 15-lipoxygenase-1 (15-LOX-1) metabolism could regulate vascular functions. The initial LA derivative from the 15-LOX-1 pathway, 13-hydroperoxyoctadecadienoic acid (HPODE), can induce endothelial death, whereas the reduced hydroxyl product, 13-hydroxyoctadecadienoic acid (HODE), is abundantly produced during vascular activation. However, the relative contribution of specific LA-derived metabolites on impairment of mammary endothelial integrity is unknown. Our hypothesis was that S. uberis-induced LA-derived 15-LOX-1 oxygenation products impair mammary endothelial barrier integrity by apoptosis. Exposure of bovine mammary endothelial cells (BMEC) to S. uberis did not increase 15-LOX-1 LA metabolism. However, S. uberis challenge of bovine monocytes demonstrated that monocytes may be a significant source of both 13-HPODE and 13-HODE during mastitis. Exposure of BMEC to 13-HPODE, but not 13-HODE, significantly reduced endothelial barrier integrity and increased apoptosis. Changing oxidant status by coexposure to an antioxidant during 13-HPODE treatment prevented adverse effects of 13-HPODE, including amelioration of apoptosis. A better understanding of how the oxidant status of the vascular microenvironment impacts endothelial barrier properties could lead to more efficacious treatments for S. uberis mastitis. PMID:27818578

  17. Development of Endothelial-Specific Single Inducible Lentiviral Vectors for Genetic Engineering of Endothelial Progenitor Cells

    PubMed Central

    Yang, Guanghua; Kramer, M. Gabriela; Fernandez-Ruiz, Veronica; Kawa, Milosz P.; Huang, Xin; Liu, Zhongmin; Prieto, Jesus; Qian, Cheng

    2015-01-01

    Endothelial progenitor cells (EPC) are able to migrate to tumor vasculature. These cells, if genetically modified, can be used as vehicles to deliver toxic material to, or express anticancer proteins in tumor. To test this hypothesis, we developed several single, endothelial-specific, and doxycycline-inducible self-inactivating (SIN) lentiviral vectors. Two distinct expression cassettes were inserted into a SIN-vector: one controlled by an endothelial lineage-specific, murine vascular endothelial cadherin (mVEcad) promoter for the expression of a transactivator, rtTA2S-M2; and the other driven by an inducible promoter, TREalb, for a firefly luciferase reporter gene. We compared the expression levels of luciferase in different vector constructs, containing either the same or opposite orientation with respect to the vector sequence. The results showed that the vector with these two expression cassettes placed in opposite directions was optimal, characterized by a robust induction of the transgene expression (17.7- to 73-fold) in the presence of doxycycline in several endothelial cell lines, but without leakiness when uninduced. In conclusion, an endothelial lineage-specific single inducible SIN lentiviral vector has been developed. Such a lentiviral vector can be used to endow endothelial progenitor cells with anti-tumor properties. PMID:26612671

  18. Development of Endothelial-Specific Single Inducible Lentiviral Vectors for Genetic Engineering of Endothelial Progenitor Cells.

    PubMed

    Yang, Guanghua; Kramer, M Gabriela; Fernandez-Ruiz, Veronica; Kawa, Milosz P; Huang, Xin; Liu, Zhongmin; Prieto, Jesus; Qian, Cheng

    2015-11-27

    Endothelial progenitor cells (EPC) are able to migrate to tumor vasculature. These cells, if genetically modified, can be used as vehicles to deliver toxic material to, or express anticancer proteins in tumor. To test this hypothesis, we developed several single, endothelial-specific, and doxycycline-inducible self-inactivating (SIN) lentiviral vectors. Two distinct expression cassettes were inserted into a SIN-vector: one controlled by an endothelial lineage-specific, murine vascular endothelial cadherin (mVEcad) promoter for the expression of a transactivator, rtTA2S-M2; and the other driven by an inducible promoter, TREalb, for a firefly luciferase reporter gene. We compared the expression levels of luciferase in different vector constructs, containing either the same or opposite orientation with respect to the vector sequence. The results showed that the vector with these two expression cassettes placed in opposite directions was optimal, characterized by a robust induction of the transgene expression (17.7- to 73-fold) in the presence of doxycycline in several endothelial cell lines, but without leakiness when uninduced. In conclusion, an endothelial lineage-specific single inducible SIN lentiviral vector has been developed. Such a lentiviral vector can be used to endow endothelial progenitor cells with anti-tumor properties.

  19. KRIT1 protein depletion modifies endothelial cell behavior via increased vascular endothelial growth factor (VEGF) signaling.

    PubMed

    DiStefano, Peter V; Kuebel, Julia M; Sarelius, Ingrid H; Glading, Angela J

    2014-11-21

    Disruption of endothelial cell-cell contact is a key event in many cardiovascular diseases and a characteristic of pathologically activated vascular endothelium. The CCM (cerebral cavernous malformation) family of proteins (KRIT1 (Krev-interaction trapped 1), PDCD10, and CCM2) are critical regulators of endothelial cell-cell contact and vascular homeostasis. Here we show novel regulation of vascular endothelial growth factor (VEGF) signaling in KRIT1-depleted endothelial cells. Loss of KRIT1 and PDCD10, but not CCM2, increases nuclear β-catenin signaling and up-regulates VEGF-A protein expression. In KRIT1-depleted cells, increased VEGF-A levels led to increased VEGF receptor 2 (VEGFR2) activation and subsequent alteration of cytoskeletal organization, migration, and barrier function and to in vivo endothelial permeability in KRIT1-deficient animals. VEGFR2 activation also increases β-catenin phosphorylation but is only partially responsible for KRIT1 depletion-dependent disruption of cell-cell contacts. Thus, VEGF signaling contributes to modifying endothelial function in KRIT1-deficient cells and microvessel permeability in Krit1(+/-) mice; however, VEGF signaling is likely not the only contributor to disrupted endothelial cell-cell contacts in the absence of KRIT1.

  20. Immunostaining of macrophages, endothelial cells and smooth muscle cells in the atherosclerotic mouse aorta

    PubMed Central

    Menon, Prashanthi; Fisher, Edward A.

    2016-01-01

    The atherosclerotic mouse aorta consists of a heterogeneous population of cells, including macrophages, endothelial cells (EC) and smooth muscle cells (SMC), that play critical roles in cardiovascular disease. Identification of these vascular cells in the vessel wall is important to understanding their function in pathological conditions. Immunohistochemistry is an invaluable technique used to detect the presence of cells in different tissues. Here, we describe immunohistochemical techniques commonly used for the detection of the vascular cells in the atherosclerotic mouse aorta using cell specific markers. PMID:26445786

  1. Endothelial cell regulation of leukocyte infiltration in inflammatory tissues

    PubMed Central

    Mantovani, A.; Introna, M.; Dejana, E.

    1995-01-01

    Endothelial cells play an important, active role in the onset and regulation of inflammatory and immune reactions. Through the production of chemokines they attract leukocytes and activate their adhesive receptors. This leads to the anchorage of leukocytes to the adhesive molecules expressed on the endothelial surface. Leukocyte adhesion to endothelial cells is frequently followed by their extravasation. The mechanisms which regulate the passage of leukocytes through endothelial clefts remain to be clarified. Many indirect data suggest that leukocytes might transfer signals to endothelial cells both through the release of active agents and adhesion to the endothelial cell surface. Adhesive molecules (such as PECAM) on the endothelial cell surface might also ‘direct’ leukocytes through the intercellular junction by haptotaxis. The information available on the molecular structure and functional properties of endothelial chemokines, adhesive molecules or junction organization is still fragmentary. Further work is needed to clarify how they interplay in regulating leukocyte infiltration into tissues. PMID:18475659

  2. [Justifying genetic and immune markers of efficiency and sensitivity under combined exposure to risk factors in mining industry workers].

    PubMed

    Dolgikh, O V; Zaitseva, N V; Krivtsov, A V; Gorshkova, K G; Lanin, D V; Bubnova, O A; Dianova, D G; Lykhina, T S; Vdovina, N A

    2014-01-01

    The authors evaluated and justified immunologic and genetic markers under combined exposure to risk factors in mining industry workers. Analysis covered polymorphism features of 29 genes with variant alleles possibly participating in occupationally conditioned diseases formation and serving as sensitivity markers of these diseases risk. The genes association selected demonstrates reliably changed polymorphism vs. the reference group (SOD2 superoxidedismutase gene, ANKK1 dophamine receptor gene, SULT1A1 sulphtransaminase gene, MTHFR methylene tetrahydrofolate reductase gene, VEGF endothelial growth factor gene, TNF-alpha tumor necrosis factor gene). Under combined exposure to occupational hazards (sylvinite dust, noise) in mining industry, this association can serve as adequate marking complex of sensitivity to development of occupationally conditioned diseases. Increased-production of immune cytokine regulation markers: tumor necrosis factor and vascular endothelial growth factor. Genes SOD2, ANKK1, SULT1A1, VEGF, TNFalpha are recommended as sensitivity markers, and the coded cytokines (tumor necrosis factor and endothelial growth factor) are proposed as effect markers in evaluation of health risk for workers in mining industry.

  3. [Endothelial cell apoptosis in erectile dysfunction].

    PubMed

    Jiang, Rui

    2012-10-01

    Erectile dysfunction (ED) is one of the most common male diseases, which seriously affects the patient's quality of life. The risk factors of ED include aging, diabetes, hypertension, hyperlipidemia, and unhealthy lifestyle, and its exact mechanism remains unclear. The apoptosis of endothelial cells in the corpus cavernosum penis may reduce NOS activity, block NO synthesis, and affect penile erection, and the mechanisms of their apoptosis vary with different causes of ED. This article updates the relationship between the apoptosis of endothelial cells and the development of ED.

  4. Crossing the endothelial barrier during metastasis.

    PubMed

    Reymond, Nicolas; d'Água, Bárbara Borda; Ridley, Anne J

    2013-12-01

    During metastasis, cancer cells disseminate to other parts of the body by entering the bloodstream in a process that is called intravasation. They then extravasate at metastatic sites by attaching to endothelial cells that line blood vessels and crossing the vessel walls of tissues or organs. This Review describes how cancer cells cross the endothelial barrier during extravasation and how different receptors, signalling pathways and circulating cells such as leukocytes and platelets contribute to this process. Identification of the mechanisms that underlie cancer cell extravasation could lead to the development of new therapies to reduce metastasis.

  5. Glial cell line-derived neurotrophic factor induced the differentiation of amniotic fluid-derived stem cells into vascular endothelial-like cells in vitro.

    PubMed

    Zhang, Ruyu; Lu, Ying; Li, Ju; Wang, Jia; Liu, Caixia; Gao, Fang; Sun, Dong

    2016-02-01

    Amniotic fluid-derived stem cells (AFSCs) are a novel source of stem cells that are isolated and cultured from second trimester amniocentesis. Glial cell line-derived neurotrophic factor (GDNF) acts as a tissue morphogen and regulates stem cell proliferation and differentiation. This study investigated the effect of an adenovirus-mediated GDNF gene, which was engineered into AFSCs, on the cells' biological properties and whether GDNF in combination with AFSCs can be directionally differentiated into vascular endothelial-like cells in vitro. AFSCs were isolated and cultured using the plastic adherence method in vitro and identified by the transcription factor Oct-4, which is the primary marker of pluripotent stem cells. AFSCs were efficiently transfected by a GFP-labeled plasmid system of an adenovirus vector carrying the GDNF gene (Ad-GDNF-GFP). Transfected AFSCs stably expressed GDNF. Transfected AFSCs were cultured in endothelial growth medium-2 containing vascular endothelial growth factor. After 1 week, AFSCs were positive for von Willebrand factor (vWF) and CD31, which are markers of endothelial cells, and the recombinant GDNF group was significantly higher than undifferentiated controls and the GFP only group. These results demonstrated that AFSCs differentiated into vascular endothelial-like cells in vitro, and recombinant GDNF promoted differentiation. The differentiation-induced AFSCs may be used as seed cells to provide a new manner of cell and gene therapies for transplantation into the vascular injury site to promote angiogenesis.

  6. Nucleic Acid Delivery for Endothelial Dysfunction in Cardiovascular Diseases

    PubMed Central

    Deshpande, Dipti; Janero, David R.; Segura-Ibarra, Victor; Blanco, Elvin; Amiji, Mansoor M.

    2016-01-01

    Endothelial dysfunction has been implicated in the pathophysiology of multiple cardiovascular diseases and involves components of both innate and acquired immune mechanisms. Identifying signature patterns and targets associated with endothelial dysfunction can help in the development of novel nanotherapeutic platforms for treatment of vascular diseases. This review discusses nucleic acid-based regulation of endothelial function and the different nucleic acid-based nanotherapeutic approaches designed to target endothelial dysfunction in cardiovascular disorders. PMID:27826366

  7. Endothelial cell promotion of early liver and pancreas development.

    PubMed

    Freedman, Deborah A; Kashima, Yasushige; Zaret, Kenneth S

    2007-01-01

    Different steps of embryonic pancreas and liver development require inductive signals from endothelial cells. During liver development, interactions between newly specified hepatic endoderm cells and nascent endothelial cells are crucial for the endoderm's subsequent growth and morphogenesis into a liver bud. Reconstitution of endothelial cell stimulation of hepatic cell growth with embryonic tissue explants demonstrated that endothelial signalling occurs independent of the blood supply. During pancreas development, midgut endoderm interactions with aortic endothelial cells induce Ptf1a, a crucial pancreatic determinant. Endothelial cells also have a later effect on pancreas development, by promoting survival of the dorsal mesenchyme, which in turn produces factors supporting pancreatic endoderm. A major goal of our laboratory is to determine the endothelial-derived molecules involved in these inductive events. Our data show that cultured endothelial cells induce Ptf1a in dorsal endoderm explants lacking an endogenous vasculature. We are purifying endothelial cell line product(s) responsible for this effect. We are also identifying endothelial-responsive regulatory elements in genes such as Ptf1a by genetic mapping and chromatin-based assays. These latter approaches will allow us to track endothelial-responsive signal pathways from DNA targets within progenitor cells. The diversity of organogenic steps dependent upon endothelial cell signalling suggests that cross-regulation of tissue development with its vasculature is a general phenomenon.

  8. Iterative marker excision system.

    PubMed

    Myronovskyi, Maksym; Rosenkränzer, Birgit; Luzhetskyy, Andriy

    2014-05-01

    The deletions of large genomic DNA fragments and consecutive gene knockouts are prerequisites for the generation of organisms with improved properties. One of the key issues in this context is the removal of antibiotic resistance markers from engineered organisms without leaving an active recombinase recognition site. Here, we report the establishment of an iterative marker excision system (IMES) that solves this problem. Based on the phiC31 integrase and its mutant att sites, IMES can be used for highly effective deletion of DNA fragments between inversely oriented B-CC and P-GG sites. The B-CC and P-GG sites are derived from attB and attP by substitution of the central core TT dinucleotide with CC and GG, respectively. An unnatural RR site that resides in the chromosome following deletion is the joining product of the right shoulders of B-CC and P-GG. We show that the RR sites do not recombine with each other as well as the RR site recombines with B-CC. The recombination efficiencies between RR and P-GG or RR and LL are only 0.1 % and 1 %, respectively. Thus, IMES can be used for multistep genomic engineering without risking unwanted DNA recombination. The fabrication of multi-purpose antibiotic cassettes and examples of the utilisation of IMES are described.

  9. Evaluation of expression profiles of hematopoietic stem cell, endothelial cell, and myeloid cell antigens in spontaneous and chemically induced hemangiosarcomas and hemangiomas in mice.

    PubMed

    Kakiuchi-Kiyota, Satoko; Crabbs, Torrie A; Arnold, Lora L; Pennington, Karen L; Cook, Jon C; Malarkey, David E; Cohen, Samuel M

    2013-07-01

    It is unclear whether the process of spontaneous and chemically induced hemangiosarcoma and hemangioma formation in mice involves the transformation of differentiated endothelial cells (ECs) or recruitment of multipotential bone marrow-derived hematopoietic stem cells or endothelial progenitor cells (EPCs), which show some degree of endothelial differentiation. In the present study, immunohistochemical staining for hematopoietic stem cell markers (CD45 and CD34), EC markers (vascular endothelial growth factor receptor 2 [VEGFR2], CD31, and factor VIII-related antigen), and a myeloid lineage marker (CD14) was employed to better define the origin of hemangiosarcomas and hemangiomas in mice. Staining was negative for CD45, factor VIII-related antigen, and CD14 and positive for CD34, VEGFR2, and CD31, indicating that mouse hemangiosarcomas and hemangiomas are composed of cells derived from EPCs expressing CD34, VEGFR2, and CD31 but not factor VIII-related antigen. The lack of CD45 expression suggests that mouse vascular tumors may arise from EPCs that are at a stage later than hematopoietic stem cells. Since factor VIII-related antigen expression is known to occur later than CD31 expression in EPCs, our observations may indicate that these tumor cells are arrested at a stage prior to complete differentiation.  In addition, myeloid lineage cells do not appear to contribute to hemangiosarcoma and hemangioma formation in mice.

  10. Intravascular papillary endothelial hyperplasia of the foot.

    PubMed

    Cisco, R W; McCormac, R M

    1994-01-01

    Intravascular papillary endothelial hyperplasia is a rare benign reactive lesion usually found in thrombosed subcutaneous blood vessels. The lesion resembles malignant angiosarcoma clinically and histopathologically, and must be diagnosed correctly to avoid inappropriate treatment. The following is a case presentation involving the foot.

  11. The function of vascular endothelial growth factor.

    PubMed

    Nieves, Bonnie J; D'Amore, Patricia A; Bryan, Brad A

    2009-01-01

    Vascular endothelial growth factor (VEGF) is considered the master regulator of angiogenesis during growth and development, as well as in disease states such as cancer, diabetes, and macular degeneration. This review details our current understanding of VEGF signaling and discusses the benefits and unexpected side effects of promising anti-angiogenic therapeutics that are currently being used to inhibit neovacularization in tumors.

  12. Brain endothelial dysfunction in cerebral adrenoleukodystrophy.

    PubMed

    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.

  13. Endothelial Progenitors: A Consensus Statement on Nomenclature.

    PubMed

    Medina, Reinhold J; Barber, Chad L; Sabatier, Florence; Dignat-George, Francoise; Melero-Martin, Juan M; Khosrotehrani, Kiarash; Ohneda, Osamu; Randi, Anna M; Chan, Jerry K Y; Yamaguchi, Teruhide; Van Hinsbergh, Victor W M; Yoder, Mervin C; Stitt, Alan W

    2017-03-10

    Endothelial progenitor cell (EPC) nomenclature remains ambiguous and there is a general lack of concordance in the stem cell field with many distinct cell subtypes continually grouped under the term "EPC." It would be highly advantageous to agree standards to confirm an endothelial progenitor phenotype and this should include detailed immunophenotyping, potency assays, and clear separation from hematopoietic angiogenic cells which are not endothelial progenitors. In this review, we seek to discourage the indiscriminate use of "EPCs," and instead propose precise terminology based on defining cellular phenotype and function. Endothelial colony forming cells and myeloid angiogenic cells are examples of two distinct and well-defined cell types that have been considered EPCs because they both promote vascular repair, albeit by completely different mechanisms of action. It is acknowledged that scientific nomenclature should be a dynamic process driven by technological and conceptual advances; ergo the ongoing "EPC" nomenclature ought not to be permanent and should become more precise in the light of strong scientific evidence. This is especially important as these cells become recognized for their role in vascular repair in health and disease; and, in some cases, progress toward use in cell therapy. © Stem Cells Translational Medicine 2017.

  14. Endothelial dysfunction: the early predictor of atherosclerosis.

    PubMed

    Mudau, Mashudu; Genis, Amanda; Lochner, Amanda; Strijdom, Hans

    2012-05-01

    Since the discovery in the 1980s that nitric oxide (NO) is in fact the elusive endothelium-derived relaxing factor, it has become evident that NO is not only a major cardiovascular signalling molecule, but that changes in its bioavailability are crucial in determining whether atherosclerosis will develop or not. Sustained high levels of harmful circulating stimuli associated with cardiovascular risk factors such as diabetes mellitus elicit responses in endothelial cells that appear sequentially, namely endothelial cell activation and endothelial dysfunction (ED). ED, characterised by reduced NO bioavailability, is now recognised by many as an early, reversible precursor of atherosclerosis. The pathogenesis of ED is multifactorial; however, oxidative stress appears to be the common underlying cellular mechanism in the ensuing loss of vaso-active, inflammatory, haemostatic and redox homeostasis in the body's vascular system. The role of ED as a pathophysiological link between early endothelial cell changes associated with cardiovascular risk factors and the development of ischaemic heart disease is of importance to basic scientists and clinicians alike.

  15. Endothelialized ePTFE Graft by Nanobiotechnology

    ClinicalTrials.gov

    2013-11-29

    The Apparatus for Processing the Tubular Graft Modification Will be Designed and Evaluated.; The On-site Capturing of the Endothelial (Progenitor) Cells by Peptide-mediated Selective Adhesion in Vitro and in Vivo Will Also be Elucidated.; The Patency Rate of ITRI-made Artificial Blood Vessels Will be Evaluated by the Porcine Animal Model.

  16. Regulation of endothelial cell differentiation and specification

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The circulatory system is the first organ system to develop in the vertebrate embryo and is critical throughout gestation for the delivery of oxygen and nutrients to, as well as removal of metabolic waste products from, growing tissues. Endothelial cells, which constitute the luminal layer of all bl...

  17. Lymphatic endothelial lineage assemblage during corneal lymphangiogenesis

    PubMed Central

    Connor, Alicia L.; Kelley, Philip M.; Tempero, Richard M.

    2015-01-01

    Post natal inflammatory lymphangiogenesis presumably requires precise regulatory processes to properly assemble proliferating lymphatic endothelial cells (LECs). The specific mechanisms that regulate the assembly of LECs during new lymphatic vessel synthesis are unclear. Dynamic endothelial shuffling and rearrangement has been proposed as a mechanism of blood vessel growth. We developed genetic lineage tracing strategies using an inductive transgenic technology to track the fate of entire tandem dimer tomato positive (tdT) lymphatic vessels or small, in some cases clonal, populations of LECs. We coupled this platform with a suture induced mouse model of corneal lymphangiogenesis and used different analytic microscopy techniques including serial live imaging to study the spatial properties of proliferating tdT+ LEC progenies. LEC precursors and their progeny expanded from the corneal limbal lymphatic vessel and were assembled contiguously to comprise a subunit within a new lymphatic vessel. VE-cadherin blockade induced morphologic abnormalities in newly synthesized lymphatic vessels, but did not disrupt the tdT+ lymphatic endothelial lineage assembly. Analysis of this static and dynamic data based largely on direct in vivo observations supports a model of lymphatic endothelial lineage assemblage during corneal inflammatory lymphangiogenesis. PMID:26658452

  18. Enteric vascular endothelial response to bacterial endotoxin.

    PubMed Central

    Koshi, R.; Mathan, V. I.; David, S.; Mathan, M. M.

    1993-01-01

    The response of enteric vasculature to endotoxin was examined at the ultrastructural level using a murine model of endotoxin-induced acute diarrhoea. Morphological changes indicative of endothelial damage were evident as early as 15 minutes following endotoxin challenge. These changes, characterized by widening of intercellular spaces, increased microvillous projections and the appearance of stress fibres, preceded the leucocytic response. Endothelial damage increased with time, being associated with progressive degenerative changes in the plasma membrane, cytoplasm and organelles, ultimately leading to desquamation. These latter changes were temporally associated with margination of neutrophils and platelet adhesion to the denuded subendothelium. The venules were the primary site of these changes while the capillaries were the least affected. The arterioles were markedly constricted with minimal endothelial damage. These changes suggest that the enteric vascular endothelium may be an important target organ, and the resultant endothelial injury may have implications in host responses to endotoxin. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 PMID:8292557

  19. Podosomes as novel players in endothelial biology.

    PubMed

    Seano, Giorgio; Daubon, Thomas; Génot, Elisabeth; Primo, Luca

    2014-10-01

    Podosomes and invadopodia, collectively known as invadosomes, are specialized cell-matrix contacts with an inherent ability to degrade extracellular matrix. Their occurrence in either normal (podosomes) or cancer cells (invadopodia) is thus traditionally associated with cell invasiveness and tissue remodelling. These specialized micro-domains of the plasma membrane are characterized by enrichment of F-actin, cortactin and metalloproteases. Recent developments in the field show that, under some circumstances, vascular endothelial cells (ECs) can be induced to form this kind of peculiar structures. Cultured ECs contain either 0.5-1-μm-wide individual podosomes or 5 to 10 μm wide ring-like clusters of podosomes (podosome rosettes). The formation of individual podosomes or podosome rosettes in ECs can be induced by soluble factors, such as TGFβ, VEGF, TNFα or pharmacological agents, such as phorbol esters. Recently, the evidence of the existence of such structures in vascular endothelium has been provided by ex vivo observation. Endothelial podosome rosettes have recently been functionally linked to arterial remodelling and sprouting angiogenesis. Concerted efforts aim now at confirming the relevance of endothelial podosomes in these patho-physiological processes in vivo. In the current review, we will introduce some general considerations regarding ECs in the vascular system. From there on, we will review the various EC types where podosomes have been described and the state-of-art knowledge hitherto generated regarding endothelial podosome features.

  20. Dobesilate enhances endothelial nitric oxide synthase-activity in macro- and microvascular endothelial cells

    PubMed Central

    Suschek, Christoph; Kolb, Hubert; Kolb-Bachofen, Victoria

    1997-01-01

    Dobesilate is used for normalizing vascular dysfunction in a number of diseases. In search for an effect on endothelial NO production, macrovascular endothelial cells from rat aorta, microvascular endothelial cells from rat exocrine pancreatic tissue, and capillary endothelial cells from rat islets, were cultured in the presence or absence of Mg-Dobesilate. The activity of constitutive nitric oxide synthase (ecNOS) in resident cells as well as of inducible nitric oxide synthase (iNOS) in cytokine-activated cells was measured indirectly by recording the citrulline concentrations in culture supernatants.In each of the different endothelial cells Mg-Dobesilate incubation (0.25–1 mM) for 24 h led to a significant and concentration-dependent increase in ecNOS-activities. With cytokine-activated endothelial cell cultures only moderate effects were seen with little or no concentration-dependency. Addition of the NOS-inhibitor NG-monomethyl-L-arginine led to a significant suppression of citrulline formation in all cultures as an evidence for the enzyme specificity of these effects.iNOS- and ecNOS-specific reverse transcription and semi-quantitative polymerase chain reaction (RT–PCR) with RNA from resident or cytokine-activated endothelial cells gave no evidence for an increase in NOS-specific mRNA after Mg-Dobesilate-treatment. Furthermore, Dobesilate-mediated enhancement of NO synthesis in resting endothelial cells was not due to iNOS induction in these cells, as no iNOS-specific signal was found by RT–PCR. PMID:9421302

  1. Genetic Variation in the Platelet Endothelial Aggregation Receptor 1 Gene Results in Endothelial Dysfunction.

    PubMed

    Fisch, Adam S; Yerges-Armstrong, Laura M; Backman, Joshua D; Wang, Hong; Donnelly, Patrick; Ryan, Kathleen A; Parihar, Ankita; Pavlovich, Mary A; Mitchell, Braxton D; O'Connell, Jeffrey R; Herzog, William; Harman, Christopher R; Wren, Jonathan D; Lewis, Joshua P

    2015-01-01

    Platelet Endothelial Aggregation Receptor 1 (PEAR1) is a newly identified membrane protein reported to be involved in multiple vascular and thrombotic processes. While most studies to date have focused on the effects of this receptor in platelets, PEAR1 is located in multiple tissues including the endothelium, where it is most highly expressed. Our first objective was to evaluate the role of PEAR1 in endothelial function by examining flow-mediated dilation of the brachial artery in 641 participants from the Heredity and Phenotype Intervention Heart Study. Our second objective was to further define the impact of PEAR1 on cardiovascular disease computationally through meta-analysis of 75,000 microarrays, yielding insights regarding PEAR1 function, and predictions of phenotypes and diseases affected by PEAR1 dysregulation. Based on the results of this meta-analysis we examined whether genetic variation in PEAR1 influences endothelial function using an ex vivo assay of endothelial cell migration. We observed a significant association between rs12041331 and flow-mediated dilation in participants of the Heredity and Phenotype Intervention Heart Study (P = 0.02). Meta-analysis results revealed that PEAR1 expression is highly correlated with several genes (e.g. ANG2, ACVRL1, ENG) and phenotypes (e.g. endothelial cell migration, angiogenesis) that are integral to endothelial function. Functional validation of these results revealed that PEAR1 rs12041331 is significantly associated with endothelial migration (P = 0.04). Our results suggest for the first time that genetic variation of PEAR1 is a significant determinant of endothelial function through pathways implicated in cardiovascular disease.

  2. Genetic Variation in the Platelet Endothelial Aggregation Receptor 1 Gene Results in Endothelial Dysfunction

    PubMed Central

    Fisch, Adam S.; Yerges-Armstrong, Laura M.; Backman, Joshua D.; Wang, Hong; Donnelly, Patrick; Ryan, Kathleen A.; Parihar, Ankita; Pavlovich, Mary A.; Mitchell, Braxton D.; O’Connell, Jeffrey R.; Herzog, William; Harman, Christopher R.; Wren, Jonathan D.; Lewis, Joshua P.

    2015-01-01

    Platelet Endothelial Aggregation Receptor 1 (PEAR1) is a newly identified membrane protein reported to be involved in multiple vascular and thrombotic processes. While most studies to date have focused on the effects of this receptor in platelets, PEAR1 is located in multiple tissues including the endothelium, where it is most highly expressed. Our first objective was to evaluate the role of PEAR1 in endothelial function by examining flow-mediated dilation of the brachial artery in 641 participants from the Heredity and Phenotype Intervention Heart Study. Our second objective was to further define the impact of PEAR1 on cardiovascular disease computationally through meta-analysis of 75,000 microarrays, yielding insights regarding PEAR1 function, and predictions of phenotypes and diseases affected by PEAR1 dysregulation. Based on the results of this meta-analysis we examined whether genetic variation in PEAR1 influences endothelial function using an ex vivo assay of endothelial cell migration. We observed a significant association between rs12041331 and flow-mediated dilation in participants of the Heredity and Phenotype Intervention Heart Study (P = 0.02). Meta-analysis results revealed that PEAR1 expression is highly correlated with several genes (e.g. ANG2, ACVRL1, ENG) and phenotypes (e.g. endothelial cell migration, angiogenesis) that are integral to endothelial function. Functional validation of these results revealed that PEAR1 rs12041331 is significantly associated with endothelial migration (P = 0.04). Our results suggest for the first time that genetic variation of PEAR1 is a significant determinant of endothelial function through pathways implicated in cardiovascular disease. PMID:26406321

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

    SciTech Connect

    Lin, Ming-Chung; Chen, Chia-Ling; Yang, Tsan-Tzu; Choi, Pui-Ching; Hsing, Chung-Hsi; Lin, Chiou-Feng

    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

  4. Corneal Donor Tissue Preparation for Endothelial Keratoplasty

    PubMed Central

    Woodward, Maria A.; Titus, Michael; Mavin, Kyle; Shtein, Roni M.

    2012-01-01

    Over the past ten years, corneal transplantation surgical techniques have undergone revolutionary changes1,2. Since its inception, traditional full thickness corneal transplantation has been the treatment to restore sight in those limited by corneal disease. Some disadvantages to this approach include a high degree of post-operative astigmatism, lack of predictable refractive outcome, and disturbance to the ocular surface. The development of Descemet's stripping endothelial keratoplasty (DSEK), transplanting only the posterior corneal stroma, Descemet's membrane, and endothelium, has dramatically changed treatment of corneal endothelial disease. DSEK is performed through a smaller incision; this technique avoids 'open sky' surgery with its risk of hemorrhage or expulsion, decreases the incidence of postoperative wound dehiscence, reduces unpredictable refractive outcomes, and may decrease the rate of transplant rejection3-6. Initially, cornea donor posterior lamellar dissection for DSEK was performed manually1 resulting in variable graft thickness and damage to the delicate corneal endothelial tissue during tissue processing. Automated lamellar dissection (Descemet's stripping automated endothelial keratoplasty, DSAEK) was developed to address these issues. Automated dissection utilizes the same technology as LASIK corneal flap creation with a mechanical microkeratome blade that helps to create uniform and thin tissue grafts for DSAEK surgery with minimal corneal endothelial cell loss in tissue processing. Eye banks have been providing full thickness corneas for surgical transplantation for many years. In 2006, eye banks began to develop methodologies for supplying precut corneal tissue for endothelial keratoplasty. With the input of corneal surgeons, eye banks have developed thorough protocols to safely and effectively prepare posterior lamellar tissue for DSAEK surgery. This can be performed preoperatively at the eye bank. Research shows no significant difference

  5. Kisspeptin-10 induces endothelial cellular senescence and impaired endothelial cell growth.

    PubMed

    Usui, Sayaka; Iso, Yoshitaka; Sasai, Masahiro; Mizukami, Takuya; Mori, Hiroyoshi; Watanabe, Takuya; Shioda, Seiji; Suzuki, Hiroshi

    2014-07-01

    The KPs (kisspeptins) are a family of multifunctional peptides with established roles in cancer metastasis, puberty and vasoconstriction. The effects of KPs on endothelial cells have yet to be determined. The aim of the present study was to investigate the effects of KP-10 on endothelial cell growth and the mechanisms underlying those effects. The administration of recombinant KP-10 into the hindlimbs of rats with ischaemia significantly impaired blood flow recovery, as shown by laser Doppler, and capillary growth, as shown using histology, compared with the controls. HUVECs (human umbilical vein endothelial cells) express the KP receptor and were treated with KP-10 in culture studies. KP-10 inhibited endothelial cell tube formation and proliferation in a significant and dose-dependent manner. The HUVECs treated with KP exhibited the senescent phenotype, as determined using a senescence-associated β-galactosidase assay, cell morphology analysis, and decreased Sirt1 (sirtuin 1) expression and increased p53 expression shown by Western blot analysis. Intriguingly, a pharmacological Rho kinase inhibitor, Y-27632, was found to increase the proliferation of HUVECs and to reduce the number of senescent phenotype cells affected by KP-10. In conclusion, KP-10 suppressed endothelial cells growth both in vivo and in vitro in the present study. The adverse effect of KP on endothelial cells was attributable, at least in part, to the induction of cellular senescence.

  6. Endothelial Cell Density to Predict Endothelial Graft Failure After Penetrating Keratoplasty

    PubMed Central

    Lass, Jonathan H.; Sugar, Alan; Benetz, Beth Ann; Beck, Roy W.; Dontchev, Mariya; Gal, Robin L.; Kollman, Craig; Gross, Robert; Heck, Ellen; Holland, Edward J.; Mannis, Mark J.; Raber, Irving; Stark, Walter; Stulting, R. Doyle

    2010-01-01

    Objective To determine whether preoperative and/or postoperative central endothelial cell density (ECD) and its rate of decline postoperatively are predictive of graft failure caused by endothelial decompensation following penetrating keratoplasty to treat a moderate-risk condition, principally, Fuchs dystrophy or pseudophakic corneal edema. Methods In a subset of Cornea Donor Study participants, a central reading center determined preoperative and postoperative ECD from available specular images for 17 grafts that failed because of endothelial decompensation and 483 grafts that did not fail. Results Preoperative ECD was not predictive of graft failure caused by endothelial decompensation (P = .91). However, the 6-month ECD was predictive of subsequent failure (P < .001). Among those that had not failed within the first 6 months, the 5-year cumulative incidence (±95% confidence interval) of failure was 13% (±12%) for the 33 participants with a 6-month ECD of less than 1700 cells/mm2 vs 2%(±3%) for the 137 participants with a 6-monthECDof 2500 cells/mm2 or higher. After 5 years’ follow-up, 40 of 277 participants (14%) with a clear graft had an ECD below 500 cells/mm2. Conclusions Preoperative ECD is unrelated to graft failure from endothelial decompensation, whereas there is a strong correlation of ECD at 6 months with graft failure from endothelial decompensation. A graft can remain clear after 5 years even when the ECD is below 500 cells/mm2. PMID:20065219

  7. In vitro endothelial cell susceptibility to xenobiotics: comparison of three cell types.

    PubMed

    L'Azou, B; Fernandez, P; Bareille, R; Beneteau, M; Bourget, C; Cambar, J; Bordenave, L

    2005-03-01

    In three different endothelial cell (EC) cultures (primary human umbilical cord vein, so-called HUVEC; and immortalized cell lines HBMEC and EA-hy-926), the effects of different xenobiotics were studied in order to standardize vascular EC models for in vitro pharmacotoxicological studies. Cell characteristics were first investigated by the production and the mRNA levels of known endothelial markers in the three EC culture models. EC secretory products, tissue plasminogen activator (tPA) and von Willebrand factor (vWF), were present in the supernatant of the immortalized cell lines. The mRNA levels of vWF, tPA, platelet endothelial cell adhesion molecule-1 (PECAM-1/CD31), and beta -integrin subunit, which are involved in the control of platelet function, coagulation, and fibrinolysis as well as in cell-matrix interactions, were investigated in all EC types. For at least three parameters, cultured cells provided marked characteristics of EC phenotype, in HUVEC and in immortalized cell lines, regardless of their origin from the macro- or microcirculation. Toxicity experiments were assessed after 24 h exposure to cadmium, cyclosporin A and cisplatin by MTT assay. These experiments show nonsignificant difference in susceptibility to cyclosporin A and cadmium on HUVEC, HBMEC, and EA-hy-926. However, HBMEC, seems to be highly susceptible to cisplatin compared to HUVEC, the latter being more sensitive than EA-hy-926. For experiments conducted with cyclosporin and cadmium, cell lines could constitute an alternative material for routine cytotoxicity studies.

  8. Phloroglucinol Inhibits the in vitro Differentiation Potential of CD34 Positive Cells into Endothelial Progenitor Cells

    PubMed Central

    Kwon, Yi-Hong; Lee, Jun-Hee; Jung, Seok-Yun; Kim, Jae-Won; Lee, Sang-Hun; Lee, Dong-Hyung; Lee, Kyu-Sup; Lee, Boo-Yong; Kwon, Sang-Mo

    2012-01-01

    Inhibiting the bioactivities of circulating endothelial progenitor cells (EPCs) results in significant inhibition of neovessel formation during tumor angiogenesis. To investigate the potential effect of phloroglucinol as an EPC inhibitor, we performed several in vitro functional assays using CD34+ cells isolated from human umbilical cord blood (HUCB). Although a high treatment dose of phloroglucinol did not show any cell toxicity, it specifically induced the cell death of EPCs under serum free conditions through apoptosis. In the EPC colony-forming assay (EPC-CFA), we observed a significant decreased in the small EPC-CFUs for the phloroglucinol group, implying that phloroglucinol inhibited the early stage of EPC commitment. In addition, in the in vitro expansion assay using CD34+ cells, treatment with phloroglucinol was shown to inhibit endothelial lineage commitment, as demonstrated by the decrease in endothelial surface markers of EPCs including CD34+, CD34+/CD133+, CD34+/CD31+ and CD34+/CXCR4+. This is the first report to demonstrate that phloroglucinol can inhibit the functional bioactivities of EPCs, indicating that phloroglucinol may be used as an EPC inhibitor in the development of biosafe anti-tumor drugs that target tumor angiogenesis. PMID:24116289

  9. G-protein-coupled receptor kinase 2 and endothelial dysfunction: molecular insights and pathophysiological mechanisms

    PubMed Central

    Taguchi, Kumiko; Matsumoto, Takayuki; Kobayashi, Tsuneo

    2015-01-01

    Smooth muscle cells (SMC) and endothelial cells are the major cell types in blood vessels. The principal function of vascular SMC in the body is to regulate blood flow and pressure through contraction and relaxation. The endothelium performs a crucial role in maintaining vascular integrity by achieving whole-organ metabolic homeostasis via the production of factors associated with vasoconstriction or vasorelaxation. In this review, we have focused on the production of nitric oxide (NO), a vasorelaxation factor. The extent of NO production represents a key marker in vascular health. A decrease in NO is capable of inducing pathological conditions associated with endothelial dysfunction, such as obesity, diabetes, cardiovascular disease, and atherosclerosis. Recent studies have strongly implicated the involvement of G-protein-coupled receptor kinase 2 (GRK2) in the progression of cardiovascular disease. Vasculature which is affected by insulin resistance and type 2 diabetes expresses high levels of GRK2, which may induce endothelial dysfunction by reducing intracellular NO. GRK2 activation also induces changes in the subcellular localization of GRK2 itself and also of β-arrestin 2, a downstream protein. In this review, we describe the pathophysiological mechanisms of insulin resistance and diabetes, focusing on the signal transduction for NO production via GRK2 and β-arrestin 2, providing novel insights into the potential field of translational investigation in the treatment of diabetic complications. PMID:26447102

  10. Functional Imaging of Working Memory and Peripheral Endothelial Function in Middle-Aged Adults

    PubMed Central

    Gonzales, Mitzi M.; Tarumi, Takashi; Tanaka, Hirofumi; Sugawara, Jun; Swann-Sternberg, Tali; Goudarzi, Katyoon; Haley, Andreana P.

    2010-01-01

    The current study examined the relationship between a prognostic indicator of vascular health, flow-mediated dilation (FMD), and working memory-related brain activation in healthy middle-aged adults. Forty-two participants underwent functional magnetic resonance imaging while completing a 2-Back working memory task. Brachial artery endothelial-dependent flow-mediated dilation (FMD) was assessed using B-mode ultrasound. The relationship between FMD and task-related brain activation in a priori regions of interest was modeled using hierarchical linear regression. Brachial FMD, was significantly related to reduced working memory-related activation in the right superior parietal lobule (β=0.338, p=0.027), independent of age, sex, systolic blood pressure, and full scale IQ (F(5,36)=2.66, p=0.038). These data provide preliminary support for the association between a preclinical marker of endothelial dysfunction and cerebral hemodynamic alterations in healthy middle-aged adults. Considering the modifiable nature of endothelial function, additional investigations on the prognostic significance of FMD on future cognitive impairment are warranted. PMID:20493622

  11. Dose-dependent surface endothelialization and biocompatibility of polyurethane noble metal nanocomposites.

    PubMed

    Hess, Christian; Schwenke, Andreas; Wagener, Philipp; Franzka, Steffen; Laszlo Sajti, Csaba; Pflaum, Michael; Wiegmann, Bettina; Haverich, Axel; Barcikowski, Stephan

    2014-06-01

    Surface pre-endothelialization is a promising approach to improve the hemocompatibility of implants, medical devices, and artificial organs. To promote the adhesive property of thermoplastic polyurethane (TPU) for endothelial cells (ECs), up to 1 wt % of gold (Au) or platinum (Pt) nanoparticles, fabricated by pulsed laser ablation in polymer solution, were embedded into the polymer matrix. The analysis of these nanocomposites showed a homogenous dispersion of the nanoparticles, with average diameters of 7 nm for Au or 9 nm for Pt. A dose-dependent effect was found when ECs were seeded onto nanocomposites comprising different nanoparticle concentrations, resulting in a fivefold improvement of proliferation at 0.1 wt % nanoparticle load. This effect was associated with a nanoparticle concentration-dependent hydrophilicity and negative charge of the nanocomposite. In dynamic flow tests, nanocomposites containing 0.1 wt % Au or Pt nanoparticles allowed for the generation of a confluent and resistant EC layer. Real-time polymerase chain reaction quantification of specific markers for EC activation indicated that ECs cultivated on nanocomposites remain in an inactivated, nonthrombogenic and noninflammatory state; however, maintain the ability to trigger an inflammatory response upon stimulation. These findings were confirmed by a platelet and leukocyte adhesion assay. The results of this study suggest the possible applicability of TPU nanocomposites, containing 0.1 wt % Au or Pt nanoparticles, for the generation of pre-endothelialized surfaces of medical devices.

  12. G-protein-coupled receptor kinase 2 and endothelial dysfunction: molecular insights and pathophysiological mechanisms.

    PubMed

    Taguchi, Kumiko; Matsumoto, Takayuki; Kobayashi, Tsuneo

    2015-01-01

    Smooth muscle cells (SMC) and endothelial cells are the major cell types in blood vessels. The principal function of vascular SMC in the body is to regulate blood flow and pressure through contraction and relaxation. The endothelium performs a crucial role in maintaining vascular integrity by achieving whole-organ metabolic homeostasis via the production of factors associated with vasoconstriction or vasorelaxation. In this review, we have focused on the production of nitric oxide (NO), a vasorelaxation factor. The extent of NO production represents a key marker in vascular health. A decrease in NO is capable of inducing pathological conditions associated with endothelial dysfunction, such as obesity, diabetes, cardiovascular disease, and atherosclerosis. Recent studies have strongly implicated the involvement of G-protein-coupled receptor kinase 2 (GRK2) in the progression of cardiovascular disease. Vasculature which is affected by insulin resistance and type 2 diabetes expresses high levels of GRK2, which may induce endothelial dysfunction by reducing intracellular NO. GRK2 activation also induces changes in the subcellular localization of GRK2 itself and also of β-arrestin 2, a downstream protein. In this review, we describe the pathophysiological mechanisms of insulin resistance and diabetes, focusing on the signal transduction for NO production via GRK2 and β-arrestin 2, providing novel insights into the potential field of translational investigation in the treatment of diabetic complications.

  13. Endothelial