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Tubule Formation (tubule + formation)
Selected AbstractsMicrotubule-dependent motility and orientation of the cortical endoplasmic reticulum in elongating characean internodal cellsCYTOSKELETON, Issue 3 2009Ilse Foissner Abstract Motility of the endoplasmic reticulum (ER) is predominantly microtubule- dependent in animal cells but thought to be entirely actomyosin-dependent in plant cells. Using live cell imaging and transmission electron microscopy to examine ER motility and structural organization in giant internodal cells of characean algae, we discovered that at the onset of cell elongation, the cortical ER situated near the plasma membrane formed a tight meshwork of predominantly transverse ER tubules that frequently coaligned with microtubules. Microtubule depolymerization increased mesh size and decreased the dynamics of the cortical ER. In contrast, perturbing the cortical actin array with cytochalasins did not affect the transverse orientation but decreased mesh size and increased ER dynamics. Our data suggest that myosin-dependent ER motility is confined to the ER strands in the streaming endoplasm, while the more sedate cortical ER uses microtubule-based mechanisms for organization and motility during early stages of cell elongation. We show further that the ER has an inherent, NEM-sensitive dynamics which can be altered via interaction with the cytoskeleton and that tubule formation and fusion events are cytoskeleton-independent. Cell Motil. Cytoskeleton 2009. © 2009 Wiley-Liss, Inc. [source] Pigment epithelium-derived factor binds to cell-surface F1 -ATP synthaseFEBS JOURNAL, Issue 9 2010Luigi Notari Pigment epithelium-derived factor (PEDF), a potent blocker of angiogenesis in vivo, and of endothelial cell migration and tubule formation, binds with high affinity to an as yet unknown protein on the surfaces of endothelial cells. Given that protein fingerprinting suggested a match of a , 60 kDa PEDF-binding protein in bovine retina with Bos taurus F1 -ATP synthase ,-subunit, and that F1Fo -ATP synthase components have been identified recently as cell-surface receptors, we examined the direct binding of PEDF to F1. Size-exclusion ultrafiltration assays showed that recombinant human PEDF formed a complex with recombinant yeast F1. Real-time binding as determined by surface plasmon resonance demonstrated that yeast F1 interacted specifically and reversibly with human PEDF. Kinetic evaluations revealed high binding affinity for PEDF, in agreement with PEDF affinities for endothelial cell surfaces. PEDF blocked interactions between F1 and angiostatin, another antiangiogenic factor, suggesting overlapping PEDF-binding and angiostatin-binding sites on F1. Surfaces of endothelial cells exhibited affinity for PEDF-binding proteins of , 60 kDa. Antibodies to F1,-subunit specifically captured PEDF-binding components in endothelial plasma membranes. The extracellular ATP synthesis activity of endothelial cells was examined in the presence of PEDF. PEDF significantly reduced the amount of extracellular ATP produced by endothelial cells, in agreement with direct interactions between cell-surface ATP synthase and PEDF. In addition to demonstrating that PEDF binds to cell-surface F1, these results show that PEDF is a ligand for endothelial cell-surface F1Fo -ATP synthase. They suggest that PEDF-mediated inhibition of ATP synthase may form part of the biochemical mechanisms by which PEDF exerts its antiangiogenic activity. Structured digital abstract ,,MINT-7711286: angiostatin (uniprotkb:P00747) physically interacts (MI:0915) with F-ATPase alpha subunit (uniprotkb:P07251), F-ATPase beta subunit (uniprotkb:P00830), F-ATPase gamma subunit (uniprotkb:P38077), F-ATPase delta subunit (uniprotkb:Q12165) and F-ATPase epsilon subunit (uniprotkb:P21306) by competition binding (MI:0405) ,,MINT-7711113: angiostatin (uniprotkb:P00747) physically interacts (MI:0915) with F-ATPase epsilon subunit (uniprotkb:P21306), F-ATPase delta subunit (uniprotkb:Q12165), F-ATPase gamma subunit (uniprotkb:P38077), F-ATPase beta subunit(uniprotkb:P00830) and F-ATPase alpha subunit (uniprotkb:P07251) by surface plasmon resonance (MI:0107) ,,MINT-7711060: F-ATPase gamma subunit (uniprotkb:P38077), F-ATPase beta subunit (uniprotkb:P00830), F-ATPase alpha subunit (uniprotkb:P07251) and PEDF (uniprotkb:P36955) physically interact (MI:0915) by molecular sieving (MI:0071) ,,MINT-7711313: F-ATPase epsilon subunit (uniprotkb:P21306), F-ATPase delta subunit (uniprotkb:Q12165), PEDF (uniprotkb:P36955), F-ATPase alpha subunit (uniprotkb:P07251), F-ATPase beta subunit (uniprotkb:P00830) and F-ATPase gamma subunit(uniprotkb:P38077) physically interact (MI:0915) by molecular sieving (MI:0071) ,,MINT-7711083: PEDF (uniprotkb:P36955) physically interacts (MI:0915) with F-ATPase epsilon subunit (uniprotkb:P21306), F-ATPase delta subunit (uniprotkb:Q12165), F-ATPase gamma subunit (uniprotkb:P38077), F-ATPase beta subunit (uniprotkb:P00830) and F-ATPase alpha subunit (uniprotkb:P07251) by surface plasmon resonance (MI:0107) [source] Inhibition of telomerase in the endothelial cells disrupts tumor angiogenesis in glioblastoma xenograftsINTERNATIONAL JOURNAL OF CANCER, Issue 6 2008Maria Laura Falchetti Abstract Tumor angiogenesis is a complex process that involves a series of interactions between tumor cells and endothelial cells (ECs). In vitro, glioblastoma multiforme (GBM) cells are known to induce an increase in proliferation, migration and tube formation by the ECs. We have previously shown that in human GBM specimens the proliferating ECs of the tumor vasculature express the catalytic component of telomerase, hTERT, and that telomerase can be upregulated in human ECs by exposing these cells to GBM in vitro. Here, we developed a controlled in vivo assay of tumor angiogenesis in which primary human umbilical vascular endothelial cells (HUVECs) were subcutaneously grafted with or without human GBM cells in immunocompromised mice as Matrigel implants. We found that primary HUVECs did not survive in Matrigel implants, and that telomerase upregulation had little effect on HUVEC survival. In the presence of GBM cells, however, the grafted HUVECs not only survived in Matrigel implants but developed tubule structures that integrated with murine microvessels. Telomerase upregulation in HUVECs enhanced such effect. More importantly, inhibition of telomerase in HUVECs completely abolished tubule formation and greatly reduced survival of these cells in the tumor xenografts. Our data demonstrate that telomerase upregulation by the ECs is a key requisite for GBM tumor angiogenesis. © 2007 Wiley-Liss, Inc. [source] Osteoprotegerin (OPG),a potential new role in the regulation of endothelialcell phenotype and tumour angiogenesis?INTERNATIONAL JOURNAL OF CANCER, Issue 8 2006Simon S. Cross Abstract The progression of cancer depends on the establishment of a tumour blood supply, and therefore tumour angiogenesis has been identified as a major target for new anticancer agents. Recent reports have suggested that osteoprotegerin (OPG) is involved in the control of endothelial cell survival through the inhibition of the activity of tumour necrosis factor- (TNF) related apoptosis-inducing ligand (TRAIL). The role of OPG in human tumour development and angiogenesis is currently unknown. In the present study we demonstrate the ability of OPG to support endothelial cell survival, as well as the formation of cord-like structures in vitro using a matrigel tubule formation assay. Investigation of various human cancers demonstrated endothelial OPG expression in 59% of malignant tumours (n = 512), but in contrast, OPG was absent in endothelial cells associated with benign tumours and normal tissues (n = 178). In a series of 400 breast tumours, endothelial OPG expression was associated with high tumour grade and certain histological types. Our data show a clear separation in endothelial OPG expression between malignant tumours and nonmalignant tissues, supporting a potential biological role for this molecule in the development and/or maintenance of the tumour vasculature. This is the first study to report the proangiogenic effects of OPG in vitro, as well as correlating expression of OPG by tumour endothelial cells with clinicopathological data in human tumours. © 2005 Wiley-Liss, Inc. [source] Recombinant vascular basement membrane derived multifunctional peptide blocks endothelial cell angiogenesis and neovascularization,JOURNAL OF CELLULAR BIOCHEMISTRY, Issue 2 2010Chengkun Wang Abstract Angiogenesis is an innovative target in the therapy of cancer and other diseases, but the effects of anti-angiogenic drugs have been rather modest in clinical trials. We have developed a small peptide, recombinant vascular basement membrane derived multifunctional peptide (rVBMDMP), which significantly inhibits endothelial cells in vitro. Here we test the mechanisms of rVBMDMP in angiogenesis balance in assays of tubule formation, colony formation, and apoptosis in HUVE-12 endothelial cells. We also analyzed the differential expression of phosphorylation proteins and related genes in a protein phosphorylation chip and extracellular matrix adhesion molecule cDNA microarray, and validated changes with Western blot or real-time quantitative PCR, respectively. rVBMDMP dose-dependently inhibited colony formation, induced apoptosis, and inhibited in vitro tubule formation. rVBMDMP increased the phosphorylation of 88 signal proteins, including caspase-3, death receptor 3, 4, and 5, and integrin ,V, ,1, and ,3, and down-regulated 41 signal proteins, including EGFR, pEGFR, VEGFR-1, and survivin versus control. rVBMDMP upregulated 14 genes, including collagen 4, 7, and 27, and down-regulated 21 genes, including integrin ,V,3, MMP10, and MMP12. Our study suggests that rVBMDMP inhibits angiogenesis and may be a viable drug candidate in anti-angiogenesis and anticancer therapies. J. Cell. Biochem. 111: 453,460, 2010. © 2010 Wiley-Liss, Inc. [source] Vascular endothelial growth factor (VEGF)-induced angiogenesis in herniated disc resorptionJOURNAL OF ORTHOPAEDIC RESEARCH, Issue 3 2002Hirotaka Haro Abstract Intervertebral disc herniation is a major cause of low back pain and sciatica. Spontaneous resorption of herniated disc (HD) is frequently detected by magnetic resonance imaging (MRI). Marked infiltration by macrophages and neo-vascularization are observed upon histogical examination of HD. In addition, enhanced MRI studies suggest that HD resorption occurs more frequently in those completely exposed to the epidural space and that this correlates with their degree of vascularization. We have postulated that the angiogenic factor, vascular endothelial growth factor (VEGF), may be implicated in the neo-vascularization of HD tissues. Here we demonstrate that VEGF and its receptors VEGFR-1 and VEGFR-2 are expressed in human surgical samples of HD. Using a co-culture system comprised of murine peritoneal macrophages and intervertebral disc tissue as a model of the acute phase of HD developed previously, an increase in macrophage VEGF protein and mRNA expression was observed upon exposure to disc tissue. Tumor necrosis factor alpha (TNF-,) was required for this induction of VEGF. Use of a novel angiogenesis assay revealed that addition of the conditioned media from the co-culture system resulted in an increase of vascular tubule formation. This effect was strongly inhibited by anti-VEGF antibody, but augmented by recombinant VEGF. We conclude that VEGF induction, under the co-culture conditions tested can result in neo-vascularization of intervertebral disc tissue and may thus play a role in the resorption of HD. © 2002 Orthopaedic Research Society. Published by Elsevier Science Ltd. All rights reserved. [source] Identification of key residues involved in mediating the in vivo anti-tumor/anti-endothelial activity of AlphastatinJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 4 2007C. A. STATON Summary., Background :,We have recently shown that Alphastatin, a 24-amino-acid peptide (ADSGEGDFLAEGGGVRGPRVVERH) derived from human fibrinogen has anti-endothelial properties in vitro and in vivo. Objectives:, The aim of this study was to determine the activity of a terminally modified (stabilized) form of Alphastatin in vitro and in vivo and to identify the key residues required for this activity. Methods:, The in vitro activity of modified Alphastatin, truncates and mutants was determined by endothelial cell (HuDMEC) tubule formation and migration. Active peptides were then assessed in vivo using syngeneic murine subcutaneous 4T1 mammary carcinomas. Results:, Modified Alphastatin-inhibited HuDMEC migration and tubule formation in response to multiple growth factors and caused a 45% inhibition in tumor growth when administered intravenously at 0.25 mg kg,1 (three times per week). Intravenous (i.v.) administration proved non-toxic at all doses investigated, whereas oral and intraperitoneal (i.p.) administration demonstrated neither anti-tumor activity nor toxicity. Truncations of Alphastatin revealed an 11-amino-acid peptide (DFLAEGGGVRG), termed AHN419, which inhibited endothelial cell activity in vitro; however, intravenous AHN419 caused a non-significant growth inhibition in vivo. Single amino acid substitutions to alanine along the entire length of Alphastatin indicated that additional residues outside the AHN419 sequence were required for full activity. Conclusions:, Terminal modification of Alphastatin altered the in vivo efficacy and these studies suggest that a hydrophobic cluster (Phe8, Leu9, Ala10 and Val15) is essential for the biological activity, but additional residues, including Ser3-Gly14, Pro18-Val20 and Arg23 are required for full inhibitory activity of Alphastatin. [source] The Role of Angiogenic and Wound Repair Factors During CMV-Accelerated Transplant Vascular Sclerosis in Rat Cardiac TransplantsAMERICAN JOURNAL OF TRANSPLANTATION, Issue 2 2008D. N. Streblow Human cytomegalovirus (HCMV) accelerates transplant vascular sclerosis (TVS), a consequence of angiogenesis (AG) and wound repair (WR). While HCMV can be localized to TVS lesions, the low number of infected cells suggests a global effect on target tissues. We used microarray analysis followed by real-time-polymerase chain reaction (RT-PCR) in an RCMV-accelerated TVS rat cardiac transplant model to determine whether CMV activates host WR and AG factors. Dysregulated cellular genes in allografts from RCMV-infected recipients were compared to those from uninfected recipients and native hearts. We demonstrated that RCMV upregulates the genes involved in WR and AG, which was highest during the critical time of TVS acceleration (21,28 days). Using a standard in vitro AG assay, virus and serum-free supernatants collected at 48 h postinfection significantly induced endothelial cell (EC) migration, branching and tubule formation compared to supernatants from mock-infected cells. Supernatants from ultraviolet (UV)-inactivated RCMV-infected cells failed to induce AG, indicating that virus replication is required. Upregulation of WR and AG genes occurs during the critical period of CMV-accelerated TVS. Targeting these genes may prevent this process and improve allograft survival. [source] Probability of axillary node involvement in patients with tubular carcinoma of the breast,BRITISH JOURNAL OF SURGERY (NOW INCLUDES EUROPEAN JOURNAL OF SURGERY), Issue 6 2001G. Papadatos Background: The aim of this study was to investigate the frequency of axillary metastasis in women with tubular carcinoma (TC) of the breast. Methods: Women who underwent axillary dissection for TC in the Western Sydney area (1984,1995) were identified retrospectively through a search of computerized records. A centralized pathology review was performed and tumours were classified as pure tubular (22) or mixed tubular (nine), on the basis of the invasive component containing 90 per cent or more, or 75,90 per cent tubule formation respectively. A Medline search of the literature was undertaken to compile a collective series (20 studies with a total of 680 patients) to address the frequency of nodal involvement in TC. A quantitative meta-analysis was used to combine the results of these studies. Results: The overall frequency of nodal metastasis was five of 31 (16 per cent); one of 22 pure tubular and four of nine mixed tumours (P = 0·019). None of the tumours with a diameter of 10 mm or less (n = 16) had nodal metastasis compared with five of 15 larger tumours (P = 0·018). The meta-analysis of 680 women showed an overall frequency of nodal metastasis in TC of 13·8 (95 per cent confidence interval 9·3,18·3) per cent. The frequency of nodal involvement was 6·6 (1·7,11·4) per cent in pure TC (n = 244) and 25·0 (12·5,37·6) per cent in mixed TC (n = 149). Conclusion: A case may be made for observing the clinically negative axilla in women with a small TC (10 mm or less in diameter). © 2001 British Journal of Surgery Society Ltd [source] | |||||||||||||