Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.10.1 (ERK)
 95,504 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The characteristics for the ex vivo expansion of the endothelial progenitor cells (EPCs) were explored. CD34+ cells were selected from umbilical cord blood mononuclear cells (MNC) by MiniMACS system, expanded under the same conditions as those for total MNC, coincubation of CD34+ and CD34- from the same donor for EPCs. In addition, the effects of vessel endothelial growth factor (VEGF) and passage on cell differentiation, expansion kinetics and apoptosis were examined. EPCs were determined and quantified by immunocytochemistry and flow cytometry. The results showed that both coculture of CD34+ and CD34- and total MNC led to a significant increase in the expansion of CD34+ cells as compared with CD34 enrichment (P < 0.05). There was a tendency toward decreased apoptosis in cultures when early passage was performed immediately after cord like structures appeared. VEGF had no significant effect on apoptosis (P > 0.05). These differentiated EPCs were positive for CD34+, von Willebrand factor (vWF), KDR, CD31 staining and phagocytized acetylated low-density lipoprotein (LDL). CD34+ cells accounted for (68.2 +/- 6.3)% of attaching (AT) cells at day 7 of culture. It was suggested the most efficient method to ex vivo expansion of EPCs was coculture of CD34+ and CD34- or total MNC. Early passage makes cell apoptosis rate decrease. VEGF had no significant effect on ex vivo expansion of EPCs.
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PMID:Characteristics of ex vivo expansion of endothelial progenitor cells. 1712 Jul 35

The clinical use of endothelial progenitor cells is hampered by difficulties in obtaining an adequate number of functional progenitors. This study aimed to establish whether human thoracic aortas harvested from healthy multiorgan donors can be a valuable source of angiogenic progenitors. Immunohistochemical tissue studies showed that two distinct cell populations with putative stem cell capabilities, one composed of CD34+ cells and the other of c-kit+ cells, are present in between the media and adventitia of human thoracic aortas. Ki-67+ cells with high growth potential were located in an area corresponding to the site of CD34+ and c-kit+ cell residence. We thus isolated cells (0.5 approximately 2.0 x 10(4) aortic progenitors per 25 cm2) which, upon culturing, coexpressed molecules of mesenchymal stromal cells (i.e., CD44+, CD90+, CD105+) and showed a transcript expression of stem cell markers (e.g., OCT4, c-kit, BCRP-1, Interleukin-6) and BMI-1. Cell expansion was adequate for use in a clinical setting. A subset of cultured cells acquired the phenotype of endothelial cells in the presence of vascular endothelial growth factor (e.g., increased expression of KDR and von Willebrand factor positivity), as documented by flow cytometry, immunofluorescence, electron microscopy, and reverse transcription-polymerase chain reaction assays. An in vitro angiogenesis test kit revealed that cells were able to form capillary-like structures within 6 hours of seeding. This study demonstrates that thoracic aortas from multiorgan donors yield mesenchymal stromal cells with the ability to differentiate in vitro into endothelial cells. These cells can be used for the creation of an allogenic bank of angiogenic progenitors, thus providing new options for restoring vascularization at ischemic sites. Disclosure of potential conflicts of interest is found at the end of this article.
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PMID:Thoracic aortas from multiorgan donors are suitable for obtaining resident angiogenic mesenchymal stromal cells. 1744 60

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.
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PMID:Vascular progenitor cells isolated from human embryonic stem cells give rise to endothelial and smooth muscle like cells and form vascular networks in vivo. 1756 86

Isolation of endothelial progenitors from human umbilical cord blood generated great hope in vascular tissue engineering. However, before clinical use, progenitor derived endothelial cells (PDECs) have to be compared with mature endothelial cells (ECs). The aim of this study was to explore the behavior of PDECs exposed to a proinflammatory cytokine (interleukin-1alpha; IL-1alpha) according to the mitogen-activated protein (MAP) kinase and nuclear factor (NF)-kappaB signal transduction pathways as well as procoagulant activity (PCA). CD34(+) mononuclear cells were isolated using magnetic beads, cultured, and compared with human saphenous vein ECs (HSVECs). PDECs express endothelial markers: CD31, VE-cadherin, von Willebrand factor, KDR, and incorporate acetylated low-density lipoprotein (Dil-Ac-LDL). IL-1alpha similarly activates c-Jun N-terminal protein kinase (JNK) and p38 pathways in HSVECs and PDECs, whereas extracellular signal-related kinase (ERK)1/2 phosphorylation is lower in PDECs than in HSVECs. Low ERK1/2 phosphorylation in PDECs was specific to IL-1alpha as vascular endothelial growth factor (VEGF) similarly stimulated ERK1/2 pathway. With respect to inhibitor of NF-kappa B (Ikappa B) degradation, NF-kappa B translocation and phosphorylation, the NF-kappa B pathway is comparable in HSVECs and PDECs after stimulation. PCA and tissue factor level induced by IL-1alpha are lower in PDECs than in HSVECs. Thus, our data show that PDECs display the characteristics of functional mature ECs under IL-1alpha stimulation. However, we observed significant differences between PDECs and HSVECs related to both ERK1/2 pathway activation and tissue factor production.
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PMID:Signal transduction and procoagulant state of human cord blood--progenitor-derived endothelial cells after interleukin-1alpha stimulation. 1757 11

Cells were isolated from four human amniotic membranes, and their biological characteristics analyzed during ex vivo expansion. Morphologically homogenous populations of fibroblast-like cells were obtained from the second or third passage. Under the appropriate culture conditions, these human amniotic membrane-derived mesenchymal cells (HAM) were shown to differentiate into adipocytes, osteocytes, chondrocytes and neuronal cells, as visualized by Oil Red O, von Kossa, alcian blue, anti-Neu N, and anti-Gal C antibody staining, respectively. Immunophenotype analysis of HAM cells revealed the presence of antigens for SSEA-3, SSEA-4, collagen type-I, -II, -III, -IV, -XII, fibronectin, alpha-SMA, vimentin, desmin, cytokeratin18 (CK18), HCAM-1, fibroblast surface protein, and human leukocyte antigen (HLA) ABC. ICAM-1 protein was weakly detectable, and proteins of TRA-1-60, VCAM-1, von Willebrand factor, PECAM-1, and HLA DR were not detected. HAM cells reached senescence after 14.5+/-0.9 passages, over a period of 146.8+/-8.9 days, and underwent an average of 36.9 4.7 population doublings. RT-PCR analysis showed that all four HAM cell lines consistently expressed genes of Oct-4, Rex-1, SCF, NCAM, nestin, BMP-4, GATA-4, HNF-4alpha, vimentin, and CK18, regardless of the passage number. The genes of Brachyury, FGF-5, Pax-6, and BMP2 were never expressed. Strikingly, alpha-fetoprotein (alphaFP), HLA ABC, and HLA DR genes were expressed in an earlier passage but not expressed in later passages. Telomerase activity of two HAM lines was discernable upon the third passage. These observations strongly suggest that HAM might be immune-privileged and, thus, advantageous as therapeutic cells.
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PMID:Ex vivo characteristics of human amniotic membrane-derived stem cells. 1815 18

The discoidin domain receptors, DDR1 and DDR2, are receptor tyrosine kinases that are activated by native triple-helical collagen. Here we have located three specific DDR2 binding sites by screening the entire triple-helical domain of collagen II, using the Collagen II Toolkit, a set of overlapping triple-helical peptides. The peptide sequence that bound DDR2 with highest affinity interestingly contained the sequence for the high affinity binding site for von Willebrand factor in collagen III. Focusing on this sequence, we used a set of truncated and alanine-substituted peptides to characterize the sequence GVMGFO (O is hydroxyproline) as the minimal collagen sequence required for DDR2 binding. Based on a recent NMR analysis of the DDR2 collagen binding domain, we generated a model of the DDR2-collagen interaction that explains why a triple-helical conformation is required for binding. Triple-helical peptides comprising the DDR2 binding motif not only inhibited DDR2 binding to collagen II but also activated DDR2 transmembrane signaling. Thus, DDR2 activation may be effected by single triple-helices rather than fibrillar collagen.
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PMID:Characterization of high affinity binding motifs for the discoidin domain receptor DDR2 in collagen. 1820 65

Biomaterials have been widely used to prepare synthetic vascular grafts over the past thirty years, but the inherent thrombogenicity of their surface can lead to graft failure. Endothelial progenitor cells (EPC) are circulating premature cells able to differentiate in either myocardial or endothelial cells (EC). The therapeutic potential of these cells and its easy obtaining technique are important reasons why these cells could be used to improve the performance of vascular grafts. In this study, two different stages of differentiation of EC derived from EPC (EC(1) and EC(2)) were characterized and cultured on poly(epsilon-caprolactone) (PCL) films treated with NaOH (PCL-NaOH). We investigated by immunolabeling the expression of CD31, von Willebrand factor (vWF), and endothelial nitric oxide synthase (eNOS) in these cells during the differentiation process. The proliferation, cell cycle, and mitochondrial function of EC(2) cultured on PCL-NaOH were evaluated at different times. The effect of this biomaterial on the nitric oxide (NO) content was also measured. The mature EC obtained from circulating progenitor cells (EC(2)) showed an appropriate growth and functionality on NaOH-treated films. They conserved their capacity to define vessel-like structures in culture and increased their basal NO production. These results underline the potential usefulness of these EC(2) to get a functional endothelialization of polymers with applications in vascular tissue engineering.
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PMID:Endothelial cells derived from circulating progenitors as an effective source to functional endothelialization of NaOH-treated poly(epsilon-caprolactone) films. 1825 77

Multi-potent adult progenitor cells (MAPCs) differentiate into endothelial cells (ECs) in the presence of vascular endothelial growth factor (VEGF). The mechanism(s) of VEGF-induced differentiation of MAPCs to ECs are not yet known. We, therefore, examined the role of mitogen-activated protein kinase/extracellular signal-regulated kinase (p42/44-MAPK/ERK1/2) signalling in endothelial differentiation from bone marrow stem cells. We observed that VEGF stimulation of MAPCs for 14 days results in a significant expression of endothelial-specific gene and/or proteins including von Willebrand factor (vWF), vascular endothelial-cadherin (VE-cadherin), VEGF receptor-2 (VEGFR2), and CD31. Up-regulation of EC-specific markers was accompanied by a cobblestone morphology, expression of endothelial nitric oxide synthase (eNOS), and Dil-Ac-LDL uptake, typical for EC morphology and function. VEGF induced a sustained activation of p42 MAPK/ERK, but not that of p44 MAPK/ERK during the course of MAPCs differentiation in a time-dependent manner up to 14 days. VEGF-induced activation of p42 MAPK/ERK also led to the nuclear translocation of MAPK/ERK1/2. Incubation of MAPCs with MAPK/ERK1/2 phosphorylation inhibitor PD98059 blocked the sustained VEGF-induced MAPK/ERK1/2 phosphorylation as well as its nuclear translocation in the differentiating MAPCs. Inhibition of MAPK/ERK1/2 phosphorylation by PD98059 also blocked the expression of EC-specific genes in these cells and their differentiation to ECs. These data suggest that VEGF induces MAPC differentiation into EC via a. MAPK/ERK1/2 signalling pathway-mediated mechanism in vitro.
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PMID:MAPK/ERK signalling mediates VEGF-induced bone marrow stem cell differentiation into endothelial cell. 1826 67

Fibrillar collagens provide the most fundamental platform in the vertebrate organism for the attachment of cells and matrix molecules. We have identified specific sites in collagens to which cells can attach, either directly or through protein intermediaries. Using Toolkits of triple-helical peptides, each peptide comprising 27 residues of collagen primary sequence and overlapping with its neighbours by nine amino acids, we have mapped the binding of receptors and other proteins on to collagens II or III. Integrin alpha2beta1 binds to several GXX'GER motifs within the collagens, the affinities of which differ sufficiently to control cell adhesion and migration independently of the cellular regulation of the integrin. The platelet receptor, Gp (glycoprotein) VI binds well to GPO (where O is hydroxyproline)-containing model peptides, but to very few Toolkit peptides, suggesting that sequence in addition to GPO triplets is important in defining GpVI binding. The Toolkits have been applied to the plasma protein vWF (von Willebrand factor), which binds to only a single sequence, identified by truncation and amino acid substitution within Toolkit peptides, as GXRGQOGVMGFO in collagens II and III. Intriguingly, the receptor tyrosine kinase, DDR2 (discoidin domain receptor 2) recognizes three sites in collagen II, including its vWF-binding site, although the amino acids that support the interaction differ slightly within this motif. Furthermore, the secreted protein BM-40 (basement membrane protein 40) also binds well to this same region. Thus the availability of extracellular collagen-binding proteins may be important in regulating and facilitating direct collagen-receptor interaction.
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PMID:Cell-collagen interactions: the use of peptide Toolkits to investigate collagen-receptor interactions. 1836 67

Endothelial progenitor cells (EPCs) are important initiators of vasculogenesis in the process of tumor neovascularization. However, it is unclear how circulating EPCs contribute to the formation of tumor microvessels. In this study, we isolated CD34(+)/CD133(+) cells from human umbilical cord blood (HUCB) and obtained EPCs with the capacities of forming colonies, uptaking acetylated low-density lipoprotein (ac-LDL), binding lectins and expressing vascular endothelial growth factor (VEGF) receptor 2 (VEGFR-2, KDR), CD31 and von Willebrand factor (vWF). These EPCs were actively proliferative and migratory, and could formed capillary-like tubules in response to VEGF. When injected into mice bearing subcutaneously implanted human malignant glioma, EPCs specifically accumulated at the sites of tumors and differentiated into mature endothelial cells (ECs), which accounted for 18% ECs of the tumor microvessels. The incorporation of circulating EPCs into tumor vessel walls significantly affected the morphology and structure of the vasculature. Our results suggest that circulating EPCs constitute important components of tumor microvessel network and contribute to tumor microvascular architecture phenotype heterogeneity.
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PMID:Incorporation of endothelial progenitor cells into the neovasculature of malignant glioma xenograft. 1905 96


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