Science.gov

Sample records for blood-derived endothelial progenitor

  1. A Novel Molecular and Functional Stemness Signature Assessing Human Cord Blood-Derived Endothelial Progenitor Cell Immaturity

    PubMed Central

    Pascaud, Juliette; Driancourt, Catherine; Boyer-Di-Ponio, Julie; Uzan, Georges

    2016-01-01

    Endothelial Colony Forming Cells (ECFCs), a distinct population of Endothelial Progenitor Cells (EPCs) progeny, display phenotypic and functional characteristics of endothelial cells while retaining features of stem/progenitor cells. Cord blood-derived ECFCs (CB-ECFCs) have a high clonogenic and proliferative potentials and they can acquire different endothelial phenotypes, this requiring some plasticity. These properties provide angiogenic and vascular repair capabilities to CB-ECFCs for ischemic cell therapies. However, the degree of immaturity retained by EPCs is still confused and poorly defined. Consequently, to better characterize CB-ECFC stemness, we quantified their clonogenic potential and demonstrated that they were reprogrammed into induced pluripotent stem cells (iPSCs) more efficiently and rapidly than adult endothelial cells. Moreover, we analyzed the transcriptional profile of a broad gene panel known to be related to stem cells. We showed that, unlike mature endothelial cells, CB-ECFCs expressed genes involved in the maintenance of embryonic stem cell properties such as DNMT3B, GDF3 or SOX2. Thus, these results provide further evidence and tools to appreciate EPC-derived cell stemness. Moreover this novel stem cell transcriptional signature of ECFCs could help better characterizing and ranging EPCs according to their immaturity profile. PMID:27043207

  2. Efficient Reprogramming of Human Fibroblasts and Blood-Derived Endothelial Progenitor Cells Using Nonmodified RNA for Reprogramming and Immune Evasion.

    PubMed

    Poleganov, Marco Alexander; Eminli, Sarah; Beissert, Tim; Herz, Stephanie; Moon, Jung-Il; Goldmann, Johanna; Beyer, Arianne; Heck, Rosario; Burkhart, Isabell; Barea Roldan, Diana; Türeci, Özlem; Yi, Kevin; Hamilton, Brad; Sahin, Ugur

    2015-11-01

    mRNA reprogramming results in the generation of genetically stable induced pluripotent stem (iPS) cells while avoiding the risks of genomic integration. Previously published mRNA reprogramming protocols have proven to be inconsistent and time-consuming and mainly restricted to fibroblasts, thereby demonstrating the need for a simple but reproducible protocol applicable to various cell types. So far there have been no published reports using mRNA to reprogram any cell type derived from human blood. Nonmodified synthetic mRNAs are immunogenic and activate cellular defense mechanisms, which can lead to cell death and inhibit mRNA translation upon repetitive transfection. Hence, to overcome RNA-related toxicity we combined nonmodified reprogramming mRNAs (OCT4, SOX2, KLF4, cMYC, NANOG, and LIN28 [OSKMNL]) with immune evasion mRNAs (E3, K3, and B18R [EKB]) from vaccinia virus. Additionally, we included mature, double-stranded microRNAs (miRNAs) from the 302/367 cluster, which are known to enhance the reprogramming process, to develop a robust reprogramming protocol for the generation of stable iPS cell lines from both human fibroblasts and human blood-outgrowth endothelial progenitor cells (EPCs). Our novel combination of RNAs enables the cell to tolerate repetitive transfections for the generation of stable iPS cell colonies from human fibroblasts within 11 days while requiring only four transfections. Moreover, our method resulted in the first known mRNA-vectored reprogramming of human blood-derived EPCs within 10 days while requiring only eight daily transfections. PMID:26381596

  3. Use of autologous blood-derived endothelial progenitor cells at point-of-care to protect against implant thrombosis in a large animal model

    PubMed Central

    Jantzen, Alexandra E.; Lane, Whitney O.; Gage, Shawn M.; Jamiolkowski, Ryan M.; Haseltine, Justin M.; Galinat, Lauren J.; Lin, Fu-Hsiung; Lawson, Jeffrey H.; Truskey, George A.; Achneck, Hardean E.

    2011-01-01

    Titanium (Ti) is commonly utilized in many cardiovascular devices, e.g. as a component of Nitinol stents, intra- and extracorporeal mechanical circulatory assist devices, but is associated with the risk of thromboemboli formation. We propose to solve this problem by lining the Ti blood-contacting surfaces with autologous peripheral blood-derived late outgrowth endothelial progenitor cells (EPCs) after having previously demonstrated that these EPCs adhere to and grow on Ti under physiological shear stresses and functionally adapt to their environment under flow conditions ex vivo. Autologous fluorescently-labeled porcine EPCs were seeded at the point-of-care in the operating room onto Ti tubes for 30 minutes and implanted into the pro-thrombotic environment of the inferior vena cava of swine (n = 8). After 3 days, Ti tubes were explanted, disassembled, and the blood-contacting surface was imaged. A blinded analysis found all 4 cell-seeded implants to be free of clot, whereas 4 controls without EPCs were either entirely occluded or partially thrombosed. Pre-labeled EPCs had spread and were present on all 4 cell-seeded implants while no endothelial cells were observed on control implants. These results suggest that late outgrowth autologous EPCs represent a promising source of lining Ti implants to reduce thrombosis in vivo. PMID:21840592

  4. Successful In Vitro Expansion and Differentiation of Cord Blood Derived CD34+ Cells into Early Endothelial Progenitor Cells Reveals Highly Differential Gene Expression

    PubMed Central

    Topcic, Denijal; Haviv, Izhak; Merivirta, Ruusu-Maaria; Agrotis, Alexander; Leitner, Ephraem; Jowett, Jeremy B.; Bode, Christoph; Lappas, Martha; Peter, Karlheinz

    2011-01-01

    Endothelial progenitor cells (EPCs) can be purified from peripheral blood, bone marrow or cord blood and are typically defined by a limited number of cell surface markers and a few functional tests. A detailed in vitro characterization is often restricted by the low cell numbers of circulating EPCs. Therefore in vitro culturing and expansion methods are applied, which allow at least distinguishing two different types of EPCs, early and late EPCs. Herein, we describe an in vitro culture technique with the aim to generate high numbers of phenotypically, functionally and genetically defined early EPCs from human cord blood. Characterization of EPCs was done by flow cytometry, immunofluorescence microscopy, colony forming unit (CFU) assay and endothelial tube formation assay. There was an average 48-fold increase in EPC numbers. EPCs expressed VEGFR-2, CD144, CD18, and CD61, and were positive for acetylated LDL uptake and ulex lectin binding. The cells stimulated endothelial tube formation only in co-cultures with mature endothelial cells and formed CFUs. Microarray analysis revealed highly up-regulated genes, including LL-37 (CAMP), PDK4, and alpha-2-macroglobulin. In addition, genes known to be associated with cardioprotective (GDF15) or pro-angiogenic (galectin-3) properties were also significantly up-regulated after a 72 h differentiation period on fibronectin. We present a novel method that allows to generate high numbers of phenotypically, functionally and genetically characterized early EPCs. Furthermore, we identified several genes newly linked to EPC differentiation, among them LL-37 (CAMP) was the most up-regulated gene. PMID:21858032

  5. Umbilical Cord Blood-Derived Mononuclear Cells Exhibit Pericyte-Like Phenotype and Support Network Formation of Endothelial Progenitor Cells In Vitro.

    PubMed

    Peters, Erica B; Liu, Betty; Christoforou, Nicolas; West, Jennifer L; Truskey, George A

    2015-10-01

    Umbilical cord blood represents a promising cell source for pro-angiogenic therapies. The present study examined the potential of mononuclear cells (MNCs) from umbilical cord blood to support endothelial progenitor cell (EPC) microvessel formation. MNCs were isolated from the cord blood of 20 separate donors and selected for further characterization based upon their proliferation potential and morphological resemblance to human vascular pericytes (HVPs). MNCs were screened for their ability to support EPC network formation using an in vitro assay (Matrigel™) as well as a reductionist, coculture system consisting of no additional angiogenic cytokines beyond those present in serum. In less than 15% of the isolations, we identified a population of highly proliferative MNCs that phenotypically resembled HVPs as assessed by expression of PDGFR-β, NG2, α-SMA, and ephrin-B2. Within a Matrigel™ system, MNCs demonstrated pericyte-like function through colocalization to EPC networks and similar effects as HVPs upon total EPC tubule length (p = 0.95) and number of branch points (p = 0.93). In a reductionist coculture system, MNCs served as pro-angiogenic mural cells by supporting EPC network formation to a significantly greater extent than HVP cocultures, by day 14 of coculture, as evidenced through EPC total tubule length (p < 0.0001) and number of branch points (p < 0.0001). Our findings are significant as we demonstrate mural cell progenitors can be isolated from umbilical cord blood and develop culture conditions to support their use in microvascular tissue engineering applications. PMID:25777295

  6. Endothelial progenitor cells: identity defined?

    PubMed Central

    Timmermans, Frank; Plum, Jean; Yöder, Mervin C; Ingram, David A; Vandekerckhove, Bart; Case, Jamie

    2009-01-01

    Abstract In the past decade, researchers have gained important insights on the role of bone marrow (BM)-derived cells in adult neovascularization. A subset of BM-derived cells, called endothelial progenitor cells (EPCs), has been of particular interest, as these cells were suggested to home to sites of neovascularization and neoendothelialization and differentiate into endothelial cells (ECs) in situ, a process referred to as postnatal vasculogenesis. Therefore, EPCs were proposed as a potential regenerative tool for treating human vascular disease and a possible target to restrict vessel growth in tumour pathology. However, conflicting results have been reported in the field, and the identification, characterization, and exact role of EPCs in vascular biology is still a subject of much discussion. The focus of this review is on the controversial issues in the field of EPCs which are related to the lack of a unique EPC marker, identification challenges related to the paucity of EPCs in the circulation, and the important phenotypical and functional overlap between EPCs, haematopoietic cells and mature ECs. We also discuss our recent findings on the origin of endothelial outgrowth cells (EOCs), showing that this in vitro defined EC population does not originate from circulating CD133+ cells or CD45+ haematopoietic cells. PMID:19067770

  7. Cord blood-derived endothelial colony-forming cell function is disrupted in congenital diaphragmatic hernia.

    PubMed

    Fujinaga, Hideshi; Fujinaga, Hiroko; Watanabe, Nobuyuki; Kato, Tomoko; Tamano, Moe; Terao, Miho; Takada, Shuji; Ito, Yushi; Umezawa, Akihiro; Kuroda, Masahiko

    2016-06-01

    Vascular growth is necessary for normal lung development. Although endothelial progenitor cells (EPCs) play an important role in vascularization, little is known about EPC function in congenital diaphragmatic hernia (CDH), a severe neonatal condition that is associated with pulmonary hypoplasia. We hypothesized that the function of endothelial colony-forming cells (ECFCs), a type of EPC, is impaired in CDH. Cord blood (CB) was collected from full-term CDH patients and healthy controls. We assessed CB progenitor cell populations as well as plasma vascular endothelial growth factor (VEGF) and stromal cell-derived factor 1α (SDF1α) levels. CB ECFC clonogenicity; growth kinetics; migration; production of VEGF, SDF1α, and nitric oxide (NO); vasculogenic capacity; and mRNA expression of VEGF-A, fms-related tyrosine kinase 1 (FLT1), kinase insert domain receptor (KDR), nitric oxide synthase (NOS) 1-3, SDF1, and chemokine (C-X-C motif) receptor 4 (CXCR4) were also assessed. Compared with controls, CB ECFCs were decreased in CDH. CDH ECFCs had reduced potential for self-renewal, clonogenicity, proliferation, and migration. Their capacity for NO production was enhanced but their response to VEGF was blunted in CDH ECFCs. In vivo potential for de novo vasculogenesis was reduced in CDH ECFCs. There was no difference in CB plasma VEGF and SDF1α concentrations, VEGF and SDF1α production by ECFCs, and ECFC mRNA expression of VEGF-A, FLT1, KDR, NOS1-3, SDF1, and CXCR4 between CDH and control subjects. In conclusion, CB ECFC function is disrupted in CDH, but these changes may be caused by mechanisms other than alteration of VEGF-NO and SDF1-CXCR4 signaling. PMID:27130531

  8. 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.

  9. Ca(2+) Signalling in Endothelial Progenitor Cells: Friend or Foe?

    PubMed

    Moccia, Francesco; Guerra, Germano

    2016-02-01

    Endothelial progenitor cells (EPCs) are mobilized either from the bone marrow and/or the arterial to replace dysfunctional endothelial cells and rescue blood perfusion in ischemic tissues. In addition, they may contribute to the angiogenic switch, thereby sustaining tumour growth and metastatization. Understanding the molecular mechanisms utilized by vascular endothelial growth factor (VEGF) to stimulate EPCs might unveil novel targets to enhance their clinical outcome in regenerative medicine and to adverse tumour vascularisation. VEGF stimulates peripheral blood-derived EPCs to undergo repetitive Ca(2+) oscillations shaped by the interaction between inositol-1,4,5-trisphosphate (InsP3 )-dependent Ca(2+) release and store-operated Ca(2+) entry (SOCE). However, the Ca(2+) machinery underlying VEGF-induced Ca(2+) spikes changes in umbilical cord blood-derived EPCs, which require TRPC3-mediated Ca(2+) entry to trigger the interplay between InsP3 and SOCE. Surprisingly, VEGF fails to elicit pro-angiogenic Ca(2+) signals when EPCs derive from renal cellular carcinoma patients, thus questioning the suitability of VEGFR-2 as a target for anti-angiogenic treatments in these individuals. The lack of response to VEGF is likely due to the dramatic rearrangement of the Ca(2+) toolkit occurring in RCC-derived EPCs. Finally, primary myelofibrosis-derived EPCs display a further pattern of reorganization of the Ca(2+) machinery and proliferate independently of SOCE. Thus, the Ca(2+) machinery in human ECFCs is extremely plastic and may change depending on the physio-pathological background of the donor. As a consequence, the Ca(2+) toolkit could properly be used to enhance the regenerative outcome of cell-based therapy or adverse tumor vascularisation. PMID:26247172

  10. Endothelial progenitor cells in hematologic malignancies

    PubMed Central

    Saulle, Ernestina; Castelli, Germana; Pelosi, Elvira

    2016-01-01

    Studies carried out in the last years have improved the understanding of the cellular and molecular mechanisms controlling angiogenesis during adult life in normal and pathological conditions. Some of these studies have led to the identification of some progenitor cells that sustain angiogenesis through indirect, paracrine mechanisms (hematopoietic angiogenic cells) and through direct mechanisms, i.e., through their capacity to generate a progeny of phenotypically and functionally competent endothelial cells [endothelial colony forming cells (ECFCs)]. The contribution of these progenitors to angiogenetic processes under physiological and pathological conditions is intensively investigated. Angiogenetic mechanisms are stimulated in various hematological malignancies, including chronic myeloid leukemia (CML), acute myeloid leukemia (AML), myelodysplastic syndromes and multiple myeloma, resulting in an increased angiogenesis that contributes to disease progression. In some of these conditions there is preliminary evidence that some endothelial cells could derive from the malignant clone, thus leading to the speculation that the leukemic cell derives from the malignant transformation of a hemangioblastic progenitor, i.e., of a cell capable of differentiation to the hematopoietic and to the endothelial cell lineages. Our understanding of the mechanisms underlying increased angiogenesis in these malignancies not only contributed to a better knowledge of the mechanisms responsible for tumor progression, but also offered the way for the discovery of new therapeutic targets. PMID:27583252

  11. Endothelial progenitor cells in hematologic malignancies.

    PubMed

    Testa, Ugo; Saulle, Ernestina; Castelli, Germana; Pelosi, Elvira

    2016-01-01

    Studies carried out in the last years have improved the understanding of the cellular and molecular mechanisms controlling angiogenesis during adult life in normal and pathological conditions. Some of these studies have led to the identification of some progenitor cells that sustain angiogenesis through indirect, paracrine mechanisms (hematopoietic angiogenic cells) and through direct mechanisms, i.e., through their capacity to generate a progeny of phenotypically and functionally competent endothelial cells [endothelial colony forming cells (ECFCs)]. The contribution of these progenitors to angiogenetic processes under physiological and pathological conditions is intensively investigated. Angiogenetic mechanisms are stimulated in various hematological malignancies, including chronic myeloid leukemia (CML), acute myeloid leukemia (AML), myelodysplastic syndromes and multiple myeloma, resulting in an increased angiogenesis that contributes to disease progression. In some of these conditions there is preliminary evidence that some endothelial cells could derive from the malignant clone, thus leading to the speculation that the leukemic cell derives from the malignant transformation of a hemangioblastic progenitor, i.e., of a cell capable of differentiation to the hematopoietic and to the endothelial cell lineages. Our understanding of the mechanisms underlying increased angiogenesis in these malignancies not only contributed to a better knowledge of the mechanisms responsible for tumor progression, but also offered the way for the discovery of new therapeutic targets. PMID:27583252

  12. Transplantation of human umbilical cord blood-derived adherent progenitors into the developing rodent brain.

    PubMed

    Coenen, Martin; Kögler, Gesine; Wernet, Peter; Brüstle, Oliver

    2005-08-01

    The results of several recent studies suggest that human umbilical cord blood (HUCB)-derived cells have the potential to undergo neural differentiation both in vitro and in vivo. Transplantation into the embryonic ventricular zone provides a unique opportunity to study the migration and differentiation of nonneural somatic progenitor cells in response to instructive cues within the developing neuroepithelium. We isolated an adherently growing population of HUCB-derived cells expressing CD13, CD29, CD49e, CD71, CD73, CD166, Flk-1, and vimentin but lacking CD34 and CD45. On transplantation into the ventricles of embryonic day 16.5 rat embryos, these cells formed subventricular clusters that extended into a variety of host brain regions, including striatum, cortex, hippocampus, thalamus, hypothalamus, tectum, pons, and cerebellum. Donor cells identified with an antibody to human nuclei or human-specific DNA in situ hybridization maintained expression of their original marker antigens and showed no expression of the neural markers MAP2 and NeuN (neurons), GFAP (astrocytes), and CNP (oligodendrocytes). In contrast to grafted primary neural cells, they remained largely confined to subventricular clusters with little evidence for intraparenchymal integration. Thus, the neurogenic environment of the embryonic ventricular zone does not promote the elaboration of a neural phenotype in HUCB-derived cells. PMID:16106216

  13. Retinal Endothelial Cell Apoptosis Stimulates Recruitment of Endothelial Progenitor Cells

    PubMed Central

    Bhatwadekar, Ashay D.; Glenn, Josephine V.; Curtis, Tim M.; Grant, Maria B.; Stitt, Alan W.; Gardiner, Tom A.

    2013-01-01

    Purpose Bone marrow–derived endothelial progenitor cells (EPCs) contribute to vascular repair although it is uncertain how local endothelial cell apoptosis influences their reparative function. This study was conducted to determine how the presence of apoptotic bodies at sites of endothelial damage may influence participation of EPCs in retinal microvascular repair. Methods Microlesions of apoptotic cell death were created in monolayers of retinal microvascular endothelial cells (RMECs) by using the photodynamic drug verteporfin. The adhesion of early-EPCs to these lesions was studied before detachment of the apoptotic cells or after their removal from the wound site. Apoptotic bodies were fed to normal RMECs and mRNA levels for adhesion molecules were analyzed. Results Endothelial lesions where apoptotic bodies were left attached at the wound site showed a fivefold enhancement in EPC recruitment (P < 0.05) compared with lesions where the apoptotic cells had been removed. In intact RMEC monolayers exposed to apoptotic bodies, expression of ICAM, VCAM, and E-selectin was upregulated by 5- to 15-fold (P < 0.05– 0.001). EPCs showed a characteristic chemotactic response (P < 0.05) to conditioned medium obtained from apoptotic bodies, whereas analysis of the medium showed significantly increased levels of VEGF, IL-8, IL-6, and TNF-α when compared to control medium; SDF-1 remained unchanged. Conclusions The data indicate that apoptotic bodies derived from retinal capillary endothelium mediate release of proangiogenic cytokines and chemokines and induce adhesion molecule expression in a manner that facilitates EPC recruitment. PMID:19474402

  14. Endothelial Progenitor Cells in Diabetic Retinopathy

    PubMed Central

    Lois, Noemi; McCarter, Rachel V.; O’Neill, Christina; Medina, Reinhold J.; Stitt, Alan W.

    2014-01-01

    Diabetic retinopathy (DR) is a leading cause of visual impairment worldwide. Patients with DR may irreversibly lose sight as a result of the development of diabetic macular edema (DME) and/or proliferative diabetic retinopathy (PDR); retinal blood vessel dysfunction and degeneration plays an essential role in their pathogenesis. Although new treatments have been recently introduced for DME, including intravitreal vascular endothelial growth factor inhibitors (anti-VEGFs) and steroids, a high proportion of patients (~40–50%) do not respond to these therapies. Furthermore, for people with PDR, laser photocoagulation remains a mainstay therapy despite this being an inherently destructive procedure. Endothelial progenitor cells (EPCs) are a low-frequency population of circulating cells known to be recruited to sites of vessel damage and tissue ischemia where they promote vascular healing and re-perfusion. A growing body of evidence suggests that the number and function of EPCs are altered in patients with varying degrees of diabetes duration, metabolic control, and in the presence or absence of DR. Although there are no clear-cut outcomes from these clinical studies, there is mounting evidence that some EPC sub-types may be involved in the pathogenesis of DR and may also serve as biomarkers for disease progression and stratification. Moreover, some EPC sub-types have considerable potential as therapeutic modalities for DME and PDR in the context of cell therapy. This study presents basic clinical concepts of DR and combines this with a general insight on EPCs and their relation to future directions in understanding and treating this important diabetic complication. PMID:24782825

  15. Point-of-Care Rapid-Seeding Ventricular Assist Device with Blood-Derived Endothelial Cells to Create a Living Antithrombotic Coating.

    PubMed

    Noviani, Maria; Jamiolkowski, Ryan M; Grenet, Justin E; Lin, Qiuyu; Carlon, Tim A; Qi, Le; Jantzen, Alexandra E; Milano, Carmelo A; Truskey, George A; Achneck, Hardean E

    2016-01-01

    The most promising alternatives to heart transplantation are left ventricular assist devices and artificial hearts; however, their use has been limited by thrombotic complications. To reduce these, sintered titanium (Ti) surfaces were developed, but thrombosis still occurs in approximately 7.5% of patients. We have invented a rapid-seeding technology to minimize the risk of thrombosis by rapid endothelialization of sintered Ti with human cord blood-derived endothelial cells (hCB-ECs). Human cord blood-derived endothelial cells were seeded within minutes onto sintered Ti and exposed to thrombosis-prone low fluid flow shear stresses. The hCB-ECs adhered and formed a confluent endothelial monolayer on sintered Ti. The exposure of sintered Ti to 4.4 dynes/cm for 20 hr immediately after rapid seeding resulted in approximately 70% cell adherence. The cell adherence was not significantly increased by additional ex vivo static culture of rapid-seeded sintered Ti before flow exposure. In addition, adherent hCB-ECs remained functional on sintered Ti, as indicated by flow-induced increase in nitric oxide secretion and reduction in platelet adhesion. After 15 day ex vivo static culture, the adherent hCB-ECs remained metabolically active, expressed endothelial cell functional marker thrombomodulin, and reduced platelet adhesion. In conclusion, our results demonstrate the feasibility of rapid-seeding sintered Ti with blood-derived hCB-ECs to generate a living antithrombotic surface. PMID:26809085

  16. Endothelial progenitor cells in acute ischemic stroke

    PubMed Central

    Martí-Fàbregas, Joan; Crespo, Javier; Delgado-Mederos, Raquel; Martínez-Ramírez, Sergi; Peña, Esther; Marín, Rebeca; Dinia, Lavinia; Jiménez-Xarrié, Elena; Fernández-Arcos, Ana; Pérez-Pérez, Jesús; Querol, Luis; Suárez-Calvet, Marc; Badimon, Lina

    2013-01-01

    Objectives The levels of circulating endothelial progenitor cells (EPCs) in ischemic stroke have not been studied extensively and reported results are inconsistent. We aimed to investigate the time course, the prognostic relevance, and the variables associated with EPC counts in patients with ischemic stroke at different time points. Material and methods We studied prospectively 146 consecutive patients with ischemic stroke within the first 48 h from the onset of symptoms (baseline). We evaluated demographic data, classical vascular risk factors, treatment with thrombolysis and statins, stroke etiology, National Institute of Health and Stroke Scale score and outcome (favorable when Rankin scale score 0–2). Blood samples were collected at baseline, at day 7 after stroke (n = 121) and at 3 months (n = 92). The EPC were measured by flow cytometry. Results We included 146 patients with a mean age of 70.8 ± 12.2 years. The circulating EPC levels were higher on day 7 than at baseline or at 3 months (P = 0.045). Pretreatment with statins (odds ratio [OR] 3.11, P = 0.008) and stroke etiology (P = 0.032) were predictive of EPC counts in the baseline sample. EPC counts were not associated with stroke severity or functional outcome in all the patients. However, using multivariate analyses, a better functional outcome was found in patients with higher EPC counts in large-artery atherosclerosis and small-vessel disease etiologic subtypes. Conclusions After acute ischemic stroke, circulating EPC counts peaked at day 7. Pretreatment with statins increased the levels of EPC. In patients with large-artery atherosclerosis and small-vessel disease subtypes, higher counts were related to better outcome at 3 months. PMID:24363968

  17. Inhibition of cyclooxygenase (COX)-2 affects endothelial progenitor cell proliferation

    SciTech Connect

    Colleselli, Daniela; Bijuklic, Klaudija; Mosheimer, Birgit A.; Kaehler, Christian M. . E-mail: C.M.Kaehler@uibk.ac.at

    2006-09-10

    Growing evidence indicates that inducible cyclooxygenase-2 (COX-2) is involved in the pathogenesis of inflammatory disorders and various types of cancer. Endothelial progenitor cells recruited from the bone marrow have been shown to be involved in the formation of new vessels in malignancies and discussed for being a key point in tumour progression and metastasis. However, until now, nothing is known about an interaction between COX and endothelial progenitor cells (EPC). Expression of COX-1 and COX-2 was detected by semiquantitative RT-PCR and Western blot. Proliferation kinetics, cell cycle distribution and rate of apoptosis were analysed by MTT test and FACS analysis. Further analyses revealed an implication of Akt phosphorylation and caspase-3 activation. Both COX-1 and COX-2 expression can be found in bone-marrow-derived endothelial progenitor cells in vitro. COX-2 inhibition leads to a significant reduction in proliferation of endothelial progenitor cells by an increase in apoptosis and cell cycle arrest. COX-2 inhibition leads further to an increased cleavage of caspase-3 protein and inversely to inhibition of Akt activation. Highly proliferating endothelial progenitor cells can be targeted by selective COX-2 inhibition in vitro. These results indicate that upcoming therapy strategies in cancer patients targeting COX-2 may be effective in inhibiting tumour vasculogenesis as well as angiogenic processes.

  18. Endothelial progenitor cells and burn injury - exploring the relationship.

    PubMed

    Banyard, Derek A; Adnani, Blake O; Melkumyan, Satenik; Araniego, Cheryl Ann; Widgerow, Alan D

    2016-01-01

    Burn wounds result in varying degrees of soft tissue damage that are typically graded clinically. Recently a key participant in neovascularization, the endothelial progenitor cell, has been the subject of intense cardiovascular research to explore whether it can serve as a biomarker for vascular injury. In this review, we examine the identity of the endothelial progenitor cell as well as the evidence that support its role as a key responder after burn insult. While there is conflicting evidence with regards to the delta of endothelial progenitor cell mobilization and burn severity, it is clear that they play an important role in wound healing. Systematic and controlled studies are needed to clarify this relationship, and whether this population can serve as a biomarker for burn severity. PMID:27574674

  19. Regular Exercise Training Increases the Number of Endothelial Progenitor Cells and Decreases Homocysteine Levels in Healthy Peripheral Blood

    PubMed Central

    Choi, Jeong Kyu; Moon, Ki Myung; Jung, Seok Yun; Kim, Ji Yong; Choi, Sung Hyun; Kim, Da Yeon; Kang, Songhwa; Chu, Chong Woo

    2014-01-01

    Endothelial progenitor cells (EPCs) are known to play an important role in the repair of damaged blood vessels. We used an endothelial progenitor cell colony-forming assay (EPC-CFA) to determine whether EPC numbers could be increased in healthy individuals through regular exercise training. The number of functional EPCs obtained from human peripheral blood-derived AC133 stem cells was measured after a 28-day regular exercise training program. The number of total endothelial progenitor cell colony-forming units (EPC-CFU) was significantly increased compared to that in the control group (p=0.02, n=5). In addition, we observed a significant decrease in homocysteine levels followed by an increase in the number of EPC-CFUs (p=0.04, n=5), indicating that the 28-day regular exercise training could increase the number of EPC colonies and decrease homocysteine levels. Moreover, an inverse correlation was observed between small-endothelial progenitor cell colony-forming units (small-EPC-CFUs) and plasma homocysteine levels in healthy men (r=-0.8125, p=0.047). We found that regular exercise training could increase the number of EPC-CFUs and decrease homocysteine levels, thus decreasing the cardiovascular disease risk in men. PMID:24757379

  20. Endothelial progenitors in sepsis: vox clamantis in deserto?

    PubMed

    Goligorsky, Michael S

    2011-01-01

    In this issue of Critical Care, Patschan and colleagues present a study of endothelial progenitor cells (EPCs) in patients with sepsis. The importance of this study is in focusing attention on several frequently ignored aspects of sepsis. Among those are the phenomenon of microvascular dysfunction, which is potentially responsible for profound metabolic perturbations at the tissue level, and the role of endothelial progenitors in repair processes. Other important aspects of the study are the regenerative capacity of mobilized EPCs and the dissociation between the numerical value and clonogenic competence. Attempting to restore the competence to EPCs should be a priority in the future. PMID:21489327

  1. 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

  2. Enhanced adhesion of early endothelial progenitor cells to radiation-induced senescence-like vascular endothelial cells in vitro.

    PubMed

    Sermsathanasawadi, Nuttawut; Ishii, Hideto; Igarashi, Kaori; Miura, Masahiko; Yoshida, Masayuki; Inoue, Yoshinori; Iwai, Takehisa

    2009-09-01

    The effects of ionizing radiation (IR) on tumor neovascularization are still unclear. We previously reported that vascular endothelial cells (ECs) expressing the IR-induced senescence-like (IRSL) phenotype exhibit a significant decrease in angiogenic activity in vitro. In this study, we examined the effects of the IRSL phenotype on adhesion to early endothelial progenitor cells (early EPCs). Adhesion of human peripheral blood-derived early EPCs to human umbilical vein endothelial cells (HUVECs) expressing the IRSL phenotype was evaluated by an adhesion assay under static conditions. It was revealed that the IRSL HUVECs supported significantly more adhesion of early EPCs than normal HUVECs. Expressions of ICAM-1, VCAM-1 and E-selectin were up-regulated in IRSL HUVECs. Pre-treatment of IRSL HUVECs with adhesion-blocking monoclonal antibodies against E-selectin and VCAM-1 significantly reduced early EPC adhesion to IRSL HUVECs, suggesting a potential role for the E-selectin and VCAM-1 in the adhesion between IRSL ECs and early EPCs. Therefore, the IRSL phenotype expressed in ECs may enhance neovascularization via increased homing of early EPCs. Our findings are first to implicate the complex effects of this phenotype on tumor neovascularization following irradiation. PMID:19628926

  3. Molecular analysis of endothelial progenitor cell (EPC) subtypes reveals two distinct cell populations with different identities

    PubMed Central

    2010-01-01

    Background The term endothelial progenitor cells (EPCs) is currently used to refer to cell populations which are quite dissimilar in terms of biological properties. This study provides a detailed molecular fingerprint for two EPC subtypes: early EPCs (eEPCs) and outgrowth endothelial cells (OECs). Methods Human blood-derived eEPCs and OECs were characterised by using genome-wide transcriptional profiling, 2D protein electrophoresis, and electron microscopy. Comparative analysis at the transcript and protein level included monocytes and mature endothelial cells as reference cell types. Results Our data show that eEPCs and OECs have strikingly different gene expression signatures. Many highly expressed transcripts in eEPCs are haematopoietic specific (RUNX1, WAS, LYN) with links to immunity and inflammation (TLRs, CD14, HLAs), whereas many transcripts involved in vascular development and angiogenesis-related signalling pathways (Tie2, eNOS, Ephrins) are highly expressed in OECs. Comparative analysis with monocytes and mature endothelial cells clusters eEPCs with monocytes, while OECs segment with endothelial cells. Similarly, proteomic analysis revealed that 90% of spots identified by 2-D gel analysis are common between OECs and endothelial cells while eEPCs share 77% with monocytes. In line with the expression pattern of caveolins and cadherins identified by microarray analysis, ultrastructural evaluation highlighted the presence of caveolae and adherens junctions only in OECs. Conclusions This study provides evidence that eEPCs are haematopoietic cells with a molecular phenotype linked to monocytes; whereas OECs exhibit commitment to the endothelial lineage. These findings indicate that OECs might be an attractive cell candidate for inducing therapeutic angiogenesis, while eEPC should be used with caution because of their monocytic nature. PMID:20465783

  4. Intracerebroventricular Transplantation of Cord Blood-Derived Neural Progenitors in a Child With Severe Global Brain Ischemic Injury

    PubMed Central

    Jozwiak, Sergiusz; Habich, Aleksandra; Kotulska, Katarzyna; Sarnowska, Anna; Kropiwnicki, Tomasz; Janowski, Miroslaw; Jurkiewicz, Elzbieta; Lukomska, Barbara; Kmiec, Tomasz; Walecki, Jerzy; Roszkowski, Marcin; Litwin, Mieczyslaw; Oldak, Tomasz; Boruczkowski, Dariusz; Domanska-Janik, Krystyna

    2010-01-01

    Transplantation of neural stem/precursor cells has recently been proposed as a promising, albeit still controversial, approach to brain repair. Human umbilical cord blood could be a source of such therapeutic cells, proven beneficial in several preclinical models of stroke. Intracerebroventricular infusion of neutrally committed cord blood-derived cells allows their broad distribution in the CNS, whereas additional labeling with iron oxide nanoparticles (SPIO) enables to follow the fate of engrafted cells by MRI. A 16-month-old child at 7 months after the onset of cardiac arrest-induced global hypoxic/ischemic brain injury, resulting in a permanent vegetative state, was subjected to intracerebroventricular transplantation of the autologous neutrally committed cord blood cells. These cells obtained by 10-day culture in vitro in neurogenic conditions were tagged with SPIO nanoparticles and grafted monthly by three serial injections (12 × 106 cells/0.5 ml) into lateral ventricle of the brain. Neural conversion of cord blood cells and superparamagnetic labeling efficiency was confirmed by gene expression, immunocytochemistry, and phantom study. MRI examination revealed the discrete hypointense areas appearing immediately after transplantation in the vicinity of lateral ventricles wall with subsequent lowering of the signal during entire period of observation. The child was followed up for 6 months after the last transplantation and his neurological status slightly but significantly improved. No clinically significant adverse events were noted. This report indicates that intracerebroventricular transplantation of autologous, neutrally committed cord blood cells is a feasible, well tolerated, and safe procedure, at least during 6 months of our observation period. Moreover, a cell-related MRI signal persisted at a wall of lateral ventricle for more than 4 months and could be monitored in transplanted brain hemisphere. PMID:26966631

  5. Endothelial progenitor cells accelerate the resolution of deep vein thrombosis.

    PubMed

    Li, Wen-Dong; Li, Xiao-Qiang

    2016-08-01

    Deep vein thrombosis (DVT) causes high morbidity and mortality. Successful resolution of DVT-related thrombi is the key point in the treatment of DVT. Recently, endothelial progenitor cells (EPCs) which are multipotent progenitor cells mainly residing in human bone marrow have emerged as a promising therapeutic choice for DVT-related thrombus resolution. In this review, we discussed the mobilization and homing property of EPCs into the sites of thrombosis, mechanisms of EPCs in DVT-related thrombus resolution from the aspects of promoting endothelial regeneration, revascularization, vasoactive and angiogenic factor secretion, proteinase generation, thrombus propagation and recurrence prevention, and vein wall remodeling. In addition, we also provide suggestions on EPCs as a therapeutic choice for thrombus resolution. PMID:26187355

  6. Effects of physical activity on endothelial progenitor cells (EPCs)

    PubMed Central

    De Biase, Chiara; De Rosa, Roberta; Luciano, Rossella; De Luca, Stefania; Capuano, Ernesto; Trimarco, Bruno; Galasso, Gennaro

    2014-01-01

    Physical activity has a therapeutic role in cardiovascular disease (CVD), through its beneficial effects on endothelial function and cardiovascular system. Circulating endothelial progenitor cells (EPCs) are bone marrow (BM) derived cells that represent a novel therapeutic target in CVD patients, because of their ability to home to sites of ischemic injury and repair the damaged vessels. Several studies show that physical activity results in a significant increase in circulating EPCs, and, in particular, there are some evidence of the beneficial exercise-induced effects on EPCs activity in CVD settings, including coronary artery disease (CAD), heart failure (HF), and peripheral artery disease (PAD). The aim of this paper is to review the current evidence about the beneficial effects of physical exercise on endothelial function and EPCs levels and activity in both healthy subjects and patients with CVD. PMID:24550833

  7. 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

  8. 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

    2015-01-01

    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

  9. Redefining endothelial progenitor cells via clonal analysis and hematopoietic stem/progenitor cell principals

    PubMed Central

    Mead, Laura E.; Prater, Daniel; Krier, Theresa R.; Mroueh, Karim N.; Li, Fang; Krasich, Rachel; Temm, Constance J.; Prchal, Josef T.

    2007-01-01

    The limited vessel-forming capacity of infused endothelial progenitor cells (EPCs) into patients with cardiovascular dysfunction may be related to a misunderstanding of the biologic potential of the cells. EPCs are generally identified by cell surface antigen expression or counting in a commercially available kit that identifies “endothelial cell colony-forming units” (CFU-ECs). However, the origin, proliferative potential, and differentiation capacity of CFU-ECs is controversial. In contrast, other EPCs with blood vessel-forming ability, termed endothelial colony-forming cells (ECFCs), have been isolated from human peripheral blood. We compared the function of CFU-ECs and ECFCs and determined that CFU-ECs are derived from the hematopoietic system using progenitor assays, and analysis of donor cells from polycythemia vera patients harboring a Janus kinase 2 V617F mutation in hematopoietic stem cell clones. Further, CFU-ECs possess myeloid progenitor cell activity, differentiate into phagocytic macrophages, and fail to form perfused vessels in vivo. In contrast, ECFCs are clonally distinct from CFU-ECs, display robust proliferative potential, and form perfused vessels in vivo. Thus, these studies establish that CFU-ECs are not EPCs and the role of these cells in angiogenesis must be re-examined prior to further clinical trials, whereas ECFCs may serve as a potential therapy for vascular regeneration. PMID:17053059

  10. Redefining endothelial progenitor cells via clonal analysis and hematopoietic stem/progenitor cell principals.

    PubMed

    Yoder, Mervin C; Mead, Laura E; Prater, Daniel; Krier, Theresa R; Mroueh, Karim N; Li, Fang; Krasich, Rachel; Temm, Constance J; Prchal, Josef T; Ingram, David A

    2007-03-01

    The limited vessel-forming capacity of infused endothelial progenitor cells (EPCs) into patients with cardiovascular dysfunction may be related to a misunderstanding of the biologic potential of the cells. EPCs are generally identified by cell surface antigen expression or counting in a commercially available kit that identifies "endothelial cell colony-forming units" (CFU-ECs). However, the origin, proliferative potential, and differentiation capacity of CFU-ECs is controversial. In contrast, other EPCs with blood vessel-forming ability, termed endothelial colony-forming cells (ECFCs), have been isolated from human peripheral blood. We compared the function of CFU-ECs and ECFCs and determined that CFU-ECs are derived from the hematopoietic system using progenitor assays, and analysis of donor cells from polycythemia vera patients harboring a Janus kinase 2 V617F mutation in hematopoietic stem cell clones. Further, CFU-ECs possess myeloid progenitor cell activity, differentiate into phagocytic macrophages, and fail to form perfused vessels in vivo. In contrast, ECFCs are clonally distinct from CFU-ECs, display robust proliferative potential, and form perfused vessels in vivo. Thus, these studies establish that CFU-ECs are not EPCs and the role of these cells in angiogenesis must be re-examined prior to further clinical trials, whereas ECFCs may serve as a potential therapy for vascular regeneration. PMID:17053059

  11. Endothelial Progenitor Cells for Diagnosis and Prognosis in Cardiovascular Disease

    PubMed Central

    Cairo, Valentina; D'Ascola, Angela; Scuruchi, Michele; Basile, Giorgio; Mandraffino, Giuseppe

    2016-01-01

    Objective. To identify, evaluate, and synthesize evidence on the predictive power of circulating endothelial progenitor cells (EPCs) in cardiovascular disease, through a systematic review of quantitative studies. Data Sources. MEDLINE was searched using keywords related to “endothelial progenitor cells” and “endothelium” and, for the different categories, respectively, “smoking”; “blood pressure”; “diabetes mellitus” or “insulin resistance”; “dyslipidemia”; “aging” or “elderly”; “angina pectoris” or “myocardial infarction”; “stroke” or “cerebrovascular disease”; “homocysteine”; “C-reactive protein”; “vitamin D”. Study Selection. Database hits were evaluated against explicit inclusion criteria. From 927 database hits, 43 quantitative studies were included. Data Syntheses. EPC count has been suggested for cardiovascular risk estimation in the clinical practice, since it is currently accepted that EPCs can work as proangiogenic support cells, maintaining their importance as regenerative/reparative potential, and also as prognostic markers. Conclusions. EPCs showed an important role in identifying cardiovascular risk conditions, and to suggest their evaluation as predictor of outcomes appears to be reasonable in different defined clinical settings. Due to their capability of proliferation, circulation, and the development of functional progeny, great interest has been directed to therapeutic use of progenitor cells in atherosclerotic diseases. This trial is registered with registration number: Prospero CRD42015023717. PMID:26839569

  12. The relationship between oxidised LDL, endothelial progenitor cells and coronary endothelial function in patients with CHD

    PubMed Central

    Watt, Jonathan; Kennedy, Simon; Ahmed, Nadeem; Hayhurst, James; McClure, John D; Berry, Colin; Wadsworth, Roger M; Oldroyd, Keith G

    2016-01-01

    Objective The balance between coronary endothelial dysfunction and repair is influenced by many protective and deleterious factors circulating in the blood. We studied the relationship between oxidised low-density lipoprotein (oxLDL), circulating endothelial progenitor cells (EPCs) and coronary endothelial function in patients with stable coronary heart disease (CHD). Methods 33 patients with stable CHD were studied. Plasma oxLDL was measured using ELISA, coronary endothelial function was assessed using intracoronary acetylcholine infusion and EPCs were quantified using flow cytometry for CD34+/KDR+ cells. Results Plasma oxLDL correlated positively with the number of EPCs in the blood (r=0.46, p=0.02). There was a positive correlation between the number of circulating EPCs and coronary endothelial function (r=0.42, p=0.04). There was no significant correlation between oxLDL and coronary endothelial function. Conclusions Plasma levels of oxLDL are associated with increased circulating EPCs in the blood of patients with CHD, which may reflect a host-repair response to endothelial injury. Patients with stable CHD had a high prevalence of coronary endothelial dysfunction, which was associated with lower numbers of circulating EPCs, suggesting a mechanistic link between endothelial dysfunction and the pathogenesis of atherosclerosis. PMID:26848395

  13. Effects of shear stress on endothelial progenitor cells.

    PubMed

    Obi, Syotaro; Yamamoto, Kimiko; Ando, Joji

    2014-10-01

    Endothelial progenitor cells (EPCs) are adult stem cells that play a central role in neovascularization. EPCs are mobilized from bone marrow into peripheral blood, attach to existing endothelial cells, and then transmigrate across the endothelium into tissues, where they proliferate, differentiate, and form new blood vessels. In the process, EPCs are exposed to shear stress, a biomechanical force generated by flowing blood and tissue fluid flow. When cultured EPCs are exposed to controlled levels of shear stress in a flow-loading device, their bioactivities in terms of proliferation, anti-apoptosis, migration, production of bioactive substances, anti-thrombosis, and tube formation increase markedly. Expression of endothelial marker genes and proteins by EPCs also increases in response to shear stress, and they differentiate into mature endothelial cells. Great advances have been made in elucidating the mechanisms by which mature endothelial cells sense and respond to shear stress, but not in EPCs. Further study of EPC responses to shear stress will be necessary to better understand the physiological and pathophysiological roles of EPCs and to apply EPCs to new therapies in the field of regenerative medicine. PMID:25992410

  14. Nitrative Stress Participates in Endothelial Progenitor Cell Injury in Hyperhomocysteinemia

    PubMed Central

    Dong, Yu; Sun, Qi; Liu, Teng; Wang, Huanyuan; Jiao, Kun; Xu, Jiahui; Liu, Xin; Liu, Huirong; Wang, Wen

    2016-01-01

    In order to investigate the role of nitrative stress in vascular endothelial injury in hyperhomocysteinemia (HHcy), thirty healthy adult female Wistar rats were randomly divided into three groups: control, hyperhomocysteinemia model, and hyperhomocysteinemia with FeTMPyP (peroxynitrite scavenger) treatment. The endothelium-dependent dilatation of thoracic aorta in vitro was determined by response to acetylcholine (ACh). The histological changes in endothelium were assessed by HE staining and scanning electron microscopy (SEM). The expression of 3-nitrotyrosine (NT) in thoracic aorta was demonstrated by immunohistochemistry and immunofluorescence, and the number of circulating endothelial progenitor cells (EPCs) was quantified by flow cytometry. Hyperhomocysteinemia caused significant endothelial injury and dysfunction including vasodilative and histologic changes, associated with higher expression of NT in thoracic aorta. FeTMPyP treatment reversed these injuries significantly. Further, the effect of nitrative stress on cultured EPCs in vitro was investigated by administering peroxynitrite donor (3-morpholino-sydnonimine, SIN-1) and peroxynitrite scavenger (FeTMPyP). The roles of nitrative stress on cell viability, necrosis and apoptosis were evaluated with 3-(4,5-dimethylthiazol)-2,5-diphenyl tetrazolium (MTT) assay, lactate dehydrogenase (LDH) release assay and terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) assay, respectively. Also, the phospho-eNOS expression and tube formation in Matrigel of cultured EPCs was detected. Our data showed that the survival of EPCs was much lower in SIN-1 group than in vehicle group, both the apoptosis and necrosis of EPCs were much more severe, and the p-eNOS expression and tube formation in Matrigel were obviously declined. Subsequent pretreatment with FeTMPyP reversed these changes. Further, pretreatment with FeTMPyP reversed homocysteine-induced EPC injury. In conclusion, this study indicates that

  15. Nitrative Stress Participates in Endothelial Progenitor Cell Injury in Hyperhomocysteinemia.

    PubMed

    Dong, Yu; Sun, Qi; Liu, Teng; Wang, Huanyuan; Jiao, Kun; Xu, Jiahui; Liu, Xin; Liu, Huirong; Wang, Wen

    2016-01-01

    In order to investigate the role of nitrative stress in vascular endothelial injury in hyperhomocysteinemia (HHcy), thirty healthy adult female Wistar rats were randomly divided into three groups: control, hyperhomocysteinemia model, and hyperhomocysteinemia with FeTMPyP (peroxynitrite scavenger) treatment. The endothelium-dependent dilatation of thoracic aorta in vitro was determined by response to acetylcholine (ACh). The histological changes in endothelium were assessed by HE staining and scanning electron microscopy (SEM). The expression of 3-nitrotyrosine (NT) in thoracic aorta was demonstrated by immunohistochemistry and immunofluorescence, and the number of circulating endothelial progenitor cells (EPCs) was quantified by flow cytometry. Hyperhomocysteinemia caused significant endothelial injury and dysfunction including vasodilative and histologic changes, associated with higher expression of NT in thoracic aorta. FeTMPyP treatment reversed these injuries significantly. Further, the effect of nitrative stress on cultured EPCs in vitro was investigated by administering peroxynitrite donor (3-morpholino-sydnonimine, SIN-1) and peroxynitrite scavenger (FeTMPyP). The roles of nitrative stress on cell viability, necrosis and apoptosis were evaluated with 3-(4,5-dimethylthiazol)-2,5-diphenyl tetrazolium (MTT) assay, lactate dehydrogenase (LDH) release assay and terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) assay, respectively. Also, the phospho-eNOS expression and tube formation in Matrigel of cultured EPCs was detected. Our data showed that the survival of EPCs was much lower in SIN-1 group than in vehicle group, both the apoptosis and necrosis of EPCs were much more severe, and the p-eNOS expression and tube formation in Matrigel were obviously declined. Subsequent pretreatment with FeTMPyP reversed these changes. Further, pretreatment with FeTMPyP reversed homocysteine-induced EPC injury. In conclusion, this study indicates that

  16. Regulation of human endothelial progenitor cell maturation by polyurethane nanocomposites.

    PubMed

    Hung, Huey-Shan; Yang, Yi-Chun; Lin, Yu-Chun; Lin, Shinn-Zong; Kao, Wei-Chien; Hsieh, Hsien-Hsu; Chu, Mei-Yun; Fu, Ru-Huei; Hsu, Shan-hui

    2014-08-01

    The mobilization and homing of endothelial progenitor cells (EPCs) are critical to the development of an antithrombotic cardiovascular prosthesis. Polyurethane (PU) with superior elasticity may provide a mechanical environment resembling that of the natural vascular tissues. The topographical cues of PU were maximized by making nanocomposites with a small amount of gold nanoparticles (AuNPs). The nanocomposites of PU-AuNPs ("PU-Au") with a favorable response of endothelial cells were previously established. In the current study, the effect of PU and PU-Au nanocomposites on the behavior of human peripheral blood EPCs was investigated in vitro and in vivo. It was found that PU-Au promoted EPCs to become differentiated endothelial cells in vitro, confirmed by the increased expressions of CD31 and VEGF-R2 surface markers. The increased maturation of EPCs was significantly more remarkable on PU-Au, probably through the stromal derived factor 1α (SDF-1α)/CXCR4 signaling pathway. In vivo experiments showed that EPCs seeded on PU-Au coated catheters effectively reduced thrombosis by differentiation into endothelial cells. Surface endothelialization with CD31 and CD34 expression as well as intimal formation with α-SMA expression was significantly accelerated in the group receiving EPC-seeded PU-Au catheters. Moreover, the analysis of collagen deposition revealed a reduction of fibrosis in the group receiving EPC-seeded PU-Au catheters as compared to the other groups. These results suggest that EPCs engineered with a proper elastic substrate may provide unique endothelialization and antithrombogenic properties that benefit vascular tissue regeneration. PMID:24836305

  17. Peripheral Blood-Derived Mesenchymal Stromal Cells Promote Angiogenesis via Paracrine Stimulation of Vascular Endothelial Growth Factor Secretion in the Equine Model.

    PubMed

    Bussche, Leen; Van de Walle, Gerlinde R

    2014-12-01

    Mesenchymal stromal cells (MSCs) have received much attention as a potential treatment of ischemic diseases, including ischemic tissue injury and cardiac failure. The beneficial effects of MSCs are thought to be mediated by their ability to provide proangiogenic factors, creating a favorable microenvironment that results in neovascularization and tissue regeneration. To study this in more detail and to explore the potential of the horse as a valuable translational model, the objectives of the present study were to examine the presence of angiogenic stimulating factors in the conditioned medium (CM) of peripheral blood-derived equine mesenchymal stromal cells (PB-MSCs) and to study their in vitro effect on angiogenesis-related endothelial cell (EC) behavior, including proliferation and vessel formation. Our salient findings were that CM from PB-MSCs contained significant levels of several proangiogenic factors. Furthermore, we found that CM could induce angiogenesis in equine vascular ECs and confirmed that endothelin-1, insulin growth factor binding protein 2, interleukin-8, and platelet-derived growth factor-AA, but not urokinase-type plasminogen activator, were responsible for this enhanced EC network formation by increasing the expression level of vascular endothelial growth factor-A, an important angiogenesis stimulator. PMID:25313202

  18. Circulating Progenitor and Mature Endothelial Cells in Deep Vein Thrombosis

    PubMed Central

    Alessio, Aline M; Beltrame, Miriam P; Nascimento, Mariane C Flores; Vicente, Cristina P; de Godoy, Juliana AP; Silva, Junia CR Santos; Bittar, Luis Fernando; Lorand-Metze, Irene; de Paula, Erich V; Annichino-Bizzacchi, Joyce M

    2013-01-01

    Introduction: Mature circulating endothelial cells (CEC) and circulating endothelial progenitor cells (EPC) have been described in several conditions associated with endothelial injury. Their role in deep vein thrombosis (DVT) has not been previously evaluated. Patients and Methods: In this pilot study we evaluated the time course of CEC and EPC release after vena cava experimental DVT in mice, using the FeCl3 model. We also evaluated their presence in patients with DVT at different phases of the disease (acute and chronic phase). CEC and EPC were evaluated by Flow Cytometry. Results: In mice, both CEC and EPC were increased 24 hours after DVT induction, peaking 48 hours thereafter. After 72 hours, CEC counts decreased sharply, whereas EPC counts decreased less substantially. In DVT patients we observed a significant increase in CEC counts immediately after DVT compared to healthy individuals. Patients with chronic disease also presented a significant elevation of these cell count. In a subgroup of patients for whom serial samples were available, CEC counts decreased significantly after 9-15 months of the acute event. Conclusions: Our results suggest the participation of these cells in the reparative processes that follows DVT, both at immediate and late time-points. The different kinetics of CEC and EPC release in experimental DVT suggests a heterogeneous role for these cells in the reparative events after DVT. PMID:24155660

  19. Acidic Fibroblast Growth Factor Promotes Endothelial Progenitor Cells Function via Akt/FOXO3a Pathway

    PubMed Central

    Wang, Yuqiang; Cao, Qing; Sang, Tiantian; Liu, Fang; Chen, Shuyan

    2015-01-01

    Acidic fibroblast growth factor (FGF1) has been suggested to enhance the functional activities of endothelial progenitor cells (EPCs). The Forkhead homeobox type O transcription factors (FOXOs), a key substrate of the survival kinase Akt, play important roles in regulation of various cellular processes. We previously have shown that FOXO3a is the main subtype of FOXOs expressed in EPCs. Here, we aim to determine whether FGF1 promotes EPC function through Akt/FOXO3a pathway. Human peripheral blood derived EPCs were transduced with adenoviral vectors either expressing a non-phosphorylable, constitutively active triple mutant of FOXO3a (Ad-TM-FOXO3a) or a GFP control (Ad-GFP). FGF1 treatment improved functional activities of Ad-GFP transduced EPCs, including cell viability, proliferation, antiapoptosis, migration and tube formation, whereas these beneficial effects disappeared by Akt inhibitor pretreatment. Moreover, EPC function was declined by Ad-TM-FOXO3a transduction and failed to be attenuated even with FGF1 treatment. FGF1 upregulated phosphorylation levels of Akt and FOXO3a in Ad-GFP transduced EPCs, which were repressed by Akt inhibitor pretreatment. However, FGF1 failed to recover Ad-TM-FOXO3a transduced EPCs from dysfunction. These data indicate that FGF1 promoting EPC function is at least in part mediated through Akt/FOXO3a pathway. Our study may provide novel ideas for enhancing EPC angiogenic ability and optimizing EPC transplantation therapy in the future. PMID:26061278

  20. Direct intracardiac injection of umbilical cord-derived stromal cells and umbilical cord blood-derived endothelial cells for the treatment of ischemic cardiomyopathy.

    PubMed

    Suss, Paula H; Capriglione, Luiz Guilherme A; Barchiki, Fabiane; Miyague, Lye; Jackowski, Danielle; Fracaro, Letícia; Schittini, Andressa V; Senegaglia, Alexandra C; Rebelatto, Carmen L K; Olandoski, Márcia; Correa, Alejandro; Brofman, Paulo R S

    2015-07-01

    The development of new therapeutic strategies is necessary to reduce the worldwide social and economic impact of cardiovascular disease, which produces high rates of morbidity and mortality. A therapeutic option that has emerged in the last decade is cell therapy. The aim of this study was to compare the effect of transplanting human umbilical cord-derived stromal cells (UCSCs), human umbilical cord blood-derived endothelial cells (UCBECs) or a combination of these two cell types for the treatment of ischemic cardiomyopathy (IC) in a Wistar rat model. IC was induced by left coronary artery ligation, and baseline echocardiography was performed seven days later. Animals with a left ventricular ejection fraction (LVEF) of ≤40% were selected for the study. On the ninth day after IC was induced, the animals were randomized into the following experimental groups: UCSCs, UCBECs, UCSCs plus UCBECs, or vehicle (control). Thirty days after treatment, an echocardiographic analysis was performed, followed by euthanasia. The animals in all of the cell therapy groups, regardless of the cell type transplanted, had less collagen deposition in their heart tissue and demonstrated a significant improvement in myocardial function after IC. Furthermore, there was a trend of increasing numbers of blood vessels in the infarcted area. The median value of LVEF increased by 7.19% to 11.77%, whereas the control group decreased by 0.24%. These results suggest that UCSCs and UCBECs are promising cells for cellular cardiomyoplasty and can be an effective therapy for improving cardiac function following IC. PMID:25576340

  1. Endothelial Progenitor Cells in Sprouting Angiogenesis: Proteases Pave the Way.

    PubMed

    Laurenzana, A; Fibbi, G; Margheri, F; Biagioni, A; Luciani, C; Del Rosso, M; Chillà, A

    2015-01-01

    Sprouting angiogenesis consists of the expansion and remodelling of existing vessels, where the vascular sprouts connect each other to form new vascular loops. Endothelial Progenitor Cells (EPCs) are a subtype of stem cells, with high proliferative potential, able to differentiate into mature Endothelial Cells (ECs) during the neovascularization process. In addition to this direct structural role EPCs improve neovascularization, also secreting numerous pro-angiogenic factors able to enhance the proliferation, survival and function of mature ECs, and other surrounding progenitor cells. While sprouting angiogenesis by mature ECs involves resident ECs, the vasculogenic contribution of EPCs is a high hurdle race. Bone marrowmobilized EPCs have to detach from the stem cell niche, intravasate into bone marrow vessels, reach the hypoxic area or tumour site, extravasate and incorporate into the new vessel lumen, thus complementing the resident mature ECs in sprouting angiogenesis. The goal of this review is to highlight the role of the main protease systems able to control each of these steps. The pivotal protease systems here described, involved in vascular patterning in sprouting angiogenesis, are the matrix-metalloproteinases (MMPs), the serineproteinases urokinase-type plasminogen activator (uPA) associated with its receptor (uPAR) and receptorassociated plasminogen/plasmin, the neutrophil elastase and the cathepsins. Since angiogenesis plays a critical role not only in physiological but also in pathological processes, such as in tumours, controlling the contribution of EPCs to the angiogenic process, through the regulation of the protease systems involved, could yield new opportunities for the therapeutic prospect of efficient control of pathological angiogenesis. PMID:26321757

  2. ECM-Dependence of Endothelial Progenitor Cell Features.

    PubMed

    Siavashi, Vahid; Nassiri, Seyed Mahdi; Rahbarghazi, Reza; Vafaei, Rana; Sariri, Reyhaneh

    2016-08-01

    Preserving self-renewal, multipotent capacity, and large-scale expansion of highly functional progenitor cells, including endothelial progenitor cells (EPCs), is a controversial issue. These current limitations, therefore, raise the need of developing promising in vitro conditions for prolonged expansion of EPCs without loss of their stemness feature. In the current study, the possible role of three different natural extracellular substrates, including collagen, gelatin, and fibronectin, on multiple parameters of EPCs such as cell morphology, phenotype, clonogenic, and vasculogenic properties was scrutinized. Next, EPCs from GFP-positive mice were pre-expanded on each of these ECM substrates and then systemically transplanted into sublethaly irradiated mice to analyze the potency of these cells for marrow reconstitution. Our results revealed considerable promise for fibronectin for EPC expansion with maintenance of stemness characteristics, whereas gelatin and collagen matrices directed the cells toward a mature endothelial phenotype. Transplantation of EPCs pre-expanded on fibronectin resulted in widespread distribution and appropriate engraftment to various tissues with habitation in close association with the microvasculature. In addition, fibronectin pre-expanded cells were gradually enriched in the bone marrow after transplantation, resulting in marrow repopulation and hematologic recovery, leading to improved survival of recipient mice whereas gelatin- and collagen-expanded cells failed to reconstitute the bone marrow. This study demonstrated that, cell characteristics of in vitro expanded EPCs are determined by the subjacent matrix. Fibronectin-expanded EPCs are heralded as a source of great promise for bone marrow reconstitution and neo-angiogenesis in therapeutic bone marrow transplantation. J. Cell. Biochem. 117: 1934-1946, 2016. © 2016 Wiley Periodicals, Inc. PMID:26756870

  3. Circulating endothelial cells and their progenitors in acute myeloid leukemia

    PubMed Central

    Zahran, Asmaa Mohammed; Aly, Sanaa Shaker; Altayeb, Hanan Ahmed; Ali, Arwa Mohammed

    2016-01-01

    Acute myeloid leukemia (AML) is an aggressive hematological malignancy characterized by the accumulation of immature myeloid progenitor cells in the bone marrow. Studies are required to investigate the prognostic and predictive value of surrogate biomarkers. Given the importance of angiogenesis in oncology in terms of pathogenesis as well as being a target for treatment, circulating endothelial cells (CECs) and endothelial progenitor cells (EPCs) are promising candidates to serve as such markers. The aim of the present study was to quantify CECs and EPCs in patients with AML at initial diagnosis and following induction chemotherapy, and to correlate these findings with the response to treatment in AML patients. The present study included 40 patients with de novo AML and 20 age- and gender-matched healthy controls. CECs and EPCs were evaluated by flow cytometry at initial diagnosis and after induction chemotherapy (3+7 protocol for AML other than M3 and all-trans-retinoic acid plus anthracycline for M3 disease). CECs and EPCs were significantly higher in AML patients at diagnosis and after induction chemotherapy than in controls. After induction chemotherapy, CECs and EPCs were significantly decreased compared with the levels at initial diagnosis. Patients who achieved complete response (n=28) had lower initial CEC and EPC levels compared with patients who did not respond to treatment. These results suggest that CEC levels are higher in AML patients and may correlate with disease status and treatment response. Further investigations are required to better determine the predictive value and implication of these cells in AML management. PMID:27602121

  4. Endothelial Progenitor Cells in Diabetic Microvascular Complications: Friends or Foes?

    PubMed Central

    Yu, Cai-Guo; Zhang, Ning; Yuan, Sha-Sha; Ma, Yan; Yang, Long-Yan; Feng, Ying-Mei; Zhao, Dong

    2016-01-01

    Despite being featured as metabolic disorder, diabetic patients are largely affected by hyperglycemia-induced vascular abnormality. Accumulated evidence has confirmed the beneficial effect of endothelial progenitor cells (EPCs) in coronary heart disease. However, antivascular endothelial growth factor (anti-VEGF) treatment is the main therapy for diabetic retinopathy and nephropathy, indicating the uncertain role of EPCs in the pathogenesis of diabetic microvascular disease. In this review, we first illustrate how hyperglycemia induces metabolic and epigenetic changes in EPCs, which exerts deleterious impact on their number and function. We then discuss how abnormal angiogenesis develops in eyes and kidneys under diabetes condition, focusing on “VEGF uncoupling with nitric oxide” and “competitive angiopoietin 1/angiopoietin 2” mechanisms that are shared in both organs. Next, we dissect the nature of EPCs in diabetic microvascular complications. After we overview the current EPCs-related strategies, we point out new EPCs-associated options for future exploration. Ultimately, we hope that this review would uncover the mysterious nature of EPCs in diabetic microvascular disease for therapeutics. PMID:27313624

  5. Endothelial Progenitor Cells in Diabetic Microvascular Complications: Friends or Foes?

    PubMed

    Yu, Cai-Guo; Zhang, Ning; Yuan, Sha-Sha; Ma, Yan; Yang, Long-Yan; Feng, Ying-Mei; Zhao, Dong

    2016-01-01

    Despite being featured as metabolic disorder, diabetic patients are largely affected by hyperglycemia-induced vascular abnormality. Accumulated evidence has confirmed the beneficial effect of endothelial progenitor cells (EPCs) in coronary heart disease. However, antivascular endothelial growth factor (anti-VEGF) treatment is the main therapy for diabetic retinopathy and nephropathy, indicating the uncertain role of EPCs in the pathogenesis of diabetic microvascular disease. In this review, we first illustrate how hyperglycemia induces metabolic and epigenetic changes in EPCs, which exerts deleterious impact on their number and function. We then discuss how abnormal angiogenesis develops in eyes and kidneys under diabetes condition, focusing on "VEGF uncoupling with nitric oxide" and "competitive angiopoietin 1/angiopoietin 2" mechanisms that are shared in both organs. Next, we dissect the nature of EPCs in diabetic microvascular complications. After we overview the current EPCs-related strategies, we point out new EPCs-associated options for future exploration. Ultimately, we hope that this review would uncover the mysterious nature of EPCs in diabetic microvascular disease for therapeutics. PMID:27313624

  6. Endothelial progenitor cell recruitment in a microfluidic vascular model.

    PubMed

    Lewis, Daniel M; Abaci, Hasan E; Xu, Yu; Gerecht, Sharon

    2015-12-01

    During vessel injury, endothelial progenitors cells (EPCs) are recruited from bone marrow and directed to the hypoxic injury site. The hypoxic conditions in the damaged blood vessel promote TNF-α, which upregulates intercellular adhesion molecule-1 (ICAM-1). EPCs attach to endothelial cell lining using ICAM-1. Here we aimed to examine EPC attachment to ECs in an injured-blood vessel conditions. We first determined ICAM-1 expression in stimulated HUVECs. We stimulated HUVECs with 21% oxygen (atmospheric), atmospheric with TNF-α-supplemented media, 1% oxygen (hypoxia), and hypoxia with TNF-α-supplemented media and found the highest ECFC attachment on HUVECs stimulated with TNF-α and hypoxia, correlating with the highest ICAM-1 expression. We next designed, fabricated and tested a three-dimensional microbioreactor (3D MBR) system with precise control and monitoring of dissolve oxygen and media flow rate in the cellular environment. We utilized a step-wise seeding approach, producing monolayer of HUVECs on all four walls. When stimulated with both TNF-α and hypoxia, ECFC retention on HUVECs was significantly increased under low shear stress compared to static controls. Overall, the 3D MBR system mimics the pathological oxygen tension and shear stress in the damaged vasculature, providing a platform to model vascular-related disorders. PMID:26693599

  7. Directed Endothelial Progenitor Differentiation from Human Pluripotent Stem Cells Via Wnt Activation Under Defined Conditions.

    PubMed

    Bao, Xiaoping; Lian, Xiaojun; Palecek, Sean P

    2016-01-01

    Efficient derivation of endothelial cells and their progenitors from human pluripotent stem cells (hPSCs) can facilitate studies of human vascular development, disease modeling, drug discovery, and cell-based therapy. Here we provide a detailed protocol for directing hPSCs to functional endothelial cells and their progenitors in a completely defined, growth factor- and serum-free system by temporal modulation of Wnt/β-catenin signaling via small molecules. We demonstrate a 10-day, two-stage process that recapitulates endothelial cell development, in which hPSCs first differentiate to endothelial progenitors that then generate functional endothelial cells and smooth muscle cells. Methods to characterize endothelial cell identity and function are also described. PMID:27590162

  8. Characterization of Endothelial Progenitor Cell Interactions with Human Tropoelastin

    PubMed Central

    Yu, Young; Wise, Steven G.; Michael, Praveesuda L.; Bax, Daniel V.; Yuen, Gloria S. C.; Hiob, Matti A.; Yeo, Giselle C.; Filipe, Elysse C.; Dunn, Louise L.; Chan, Kim H.; Hajian, Hamid; Celermajer, David S.; Weiss, Anthony S.; Ng, Martin K. C.

    2015-01-01

    The deployment of endovascular implants such as stents in the treatment of cardiovascular disease damages the vascular endothelium, increasing the risk of thrombosis and promoting neointimal hyperplasia. The rapid restoration of a functional endothelium is known to reduce these complications. Circulating endothelial progenitor cells (EPCs) are increasingly recognized as important contributors to device re-endothelialization. Extracellular matrix proteins prominent in the vessel wall may enhance EPC-directed re-endothelialization. We examined attachment, spreading and proliferation on recombinant human tropoelastin (rhTE) and investigated the mechanism and site of interaction. EPCs attached and spread on rhTE in a dose dependent manner, reaching a maximal level of 56±3% and 54±3%, respectively. EPC proliferation on rhTE was comparable to vitronectin, fibronectin and collagen. EDTA, but not heparan sulfate or lactose, reduced EPC attachment by 81±3%, while full attachment was recovered after add-back of manganese, inferring a classical integrin-mediated interaction. Integrin αVβ3 blocking antibodies decreased EPC adhesion and spreading on rhTE by 39±3% and 56±10% respectively, demonstrating a large contribution from this specific integrin. Attachment of EPCs on N-terminal rhTE constructs N25 and N18 accounted for most of this interaction, accompanied by comparable spreading. In contrast, attachment and spreading on N10 was negligible. αVβ3 blocking antibodies reduced EPC spreading on both N25 and N18 by 45±4% and 42±14%, respectively. In conclusion, rhTE supports EPC binding via an integrin mechanism involving αVβ3. N25 and N18, but not N10 constructs of rhTE contribute to EPC binding. The regulation of EPC activity by rhTE may have implications for modulation of the vascular biocompatibility of endovascular implants. PMID:26115013

  9. Characterization of Endothelial Progenitor Cell Interactions with Human Tropoelastin.

    PubMed

    Yu, Young; Wise, Steven G; Michael, Praveesuda L; Bax, Daniel V; Yuen, Gloria S C; Hiob, Matti A; Yeo, Giselle C; Filipe, Elysse C; Dunn, Louise L; Chan, Kim H; Hajian, Hamid; Celermajer, David S; Weiss, Anthony S; Ng, Martin K C

    2015-01-01

    The deployment of endovascular implants such as stents in the treatment of cardiovascular disease damages the vascular endothelium, increasing the risk of thrombosis and promoting neointimal hyperplasia. The rapid restoration of a functional endothelium is known to reduce these complications. Circulating endothelial progenitor cells (EPCs) are increasingly recognized as important contributors to device re-endothelialization. Extracellular matrix proteins prominent in the vessel wall may enhance EPC-directed re-endothelialization. We examined attachment, spreading and proliferation on recombinant human tropoelastin (rhTE) and investigated the mechanism and site of interaction. EPCs attached and spread on rhTE in a dose dependent manner, reaching a maximal level of 56±3% and 54±3%, respectively. EPC proliferation on rhTE was comparable to vitronectin, fibronectin and collagen. EDTA, but not heparan sulfate or lactose, reduced EPC attachment by 81±3%, while full attachment was recovered after add-back of manganese, inferring a classical integrin-mediated interaction. Integrin αVβ3 blocking antibodies decreased EPC adhesion and spreading on rhTE by 39±3% and 56±10% respectively, demonstrating a large contribution from this specific integrin. Attachment of EPCs on N-terminal rhTE constructs N25 and N18 accounted for most of this interaction, accompanied by comparable spreading. In contrast, attachment and spreading on N10 was negligible. αVβ3 blocking antibodies reduced EPC spreading on both N25 and N18 by 45±4% and 42±14%, respectively. In conclusion, rhTE supports EPC binding via an integrin mechanism involving αVβ3. N25 and N18, but not N10 constructs of rhTE contribute to EPC binding. The regulation of EPC activity by rhTE may have implications for modulation of the vascular biocompatibility of endovascular implants. PMID:26115013

  10. Infection of hepatitis B virus in extrahepatic endothelial tissues mediated by endothelial progenitor cells

    PubMed Central

    Rong, Qifei; Huang, Jun; Su, Enben; Li, Jun; Li, Jianyong; Zhang, Lili; Cao, Kejiang

    2007-01-01

    Background Hepatitis B virus (HBV) replication has been reported to be involved in many extrahepatic viral disorders; however, the mechanism by which HBV is trans-infected into extrahepatic tissues such as HBV associated myocarditis remains largely unknown. Results In this study, we showed that human cord blood endothelial progenitor cells (EPCs), but not human umbilical vein endothelial cells (HUVECs) could be effectively infected by uptake of HBV in vitro. Exposure of EPCs with HBV resulted in HBV DNA and viral particles were detected in EPCs at day 3 after HBV challenge, which were peaked around day 7 and declined in 3 weeks. Consistently, HBV envelope surface and core antigens were first detected in EPCs at day 3 after virus challenge and were retained to be detectable for 3 weeks. In contrast, HBV covalently closed circular DNA was not detected in EPCs at any time after virus challenge. Intravenous transplantation of HBV-treated EPCs into myocardial infarction and acute renal ischemia mouse model resulted in incorporation of HBV into injured heart, lung, and renal capillary endothelial tissues. Conclusion These results strongly support that EPCs serve as virus carrier mediating HBV trans-infection into the injured endothelial tissues. The findings might provide a novel mechanism for HBV-associated myocarditis and other HBV-related extrahepatic diseases as well. PMID:17407553

  11. Endothelial cells are progenitors of cardiac pericytes and vascular smooth muscle cells.

    PubMed

    Chen, Qi; Zhang, Hui; Liu, Yang; Adams, Susanne; Eilken, Hanna; Stehling, Martin; Corada, Monica; Dejana, Elisabetta; Zhou, Bin; Adams, Ralf H

    2016-01-01

    Mural cells of the vessel wall, namely pericytes and vascular smooth muscle cells, are essential for vascular integrity. The developmental sources of these cells and molecular mechanisms controlling their progenitors in the heart are only partially understood. Here we show that endocardial endothelial cells are progenitors of pericytes and vascular smooth muscle cells in the murine embryonic heart. Endocardial cells undergo endothelial-mesenchymal transition and convert into primitive mesenchymal progenitors expressing the platelet-derived growth factor receptors, PDGFRα and PDGFRβ. These progenitors migrate into the myocardium, differentiate and assemble the wall of coronary vessels, which requires canonical Wnt signalling involving Frizzled4, β-catenin and endothelial cell-derived Wnt ligands. Our findings identify a novel and unexpected population of progenitors for coronary mural cells with potential relevance for heart function and disease conditions. PMID:27516371

  12. 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

  13. Reduced circulating endothelial progenitor cells in reversible cerebral vasoconstriction syndrome

    PubMed Central

    2014-01-01

    Background The pathophysiology of reversible cerebral vasoconstriction syndrome (RCVS) remains elusive. Endothelial dysfunction might play a role, but direct evidence is lacking. This study aimed to explore whether patients with RCVS have a reduced level of circulating circulating endothelial progenitor cells (EPCs) to repair the dysfunctional endothelial vasomotor control. Methods We prospectively recruited 24 patients with RCVS within one month of disease onset and 24 healthy age- and sex-matched controls. Flow cytometry was used to quantify the numbers of circulating EPCs, defined as KDR+CD133+, CD34+CD133+, and CD34+KDR+ double-positive mononuclear cells. The Lindegaard index, an index of vasoconstriction, was calculated by measuring the mean flow velocity of middle cerebral arteries and distal extracranial internal carotid arteries via color-coded sonography on the same day as blood drawing. A Lindegaard index of 2 was chosen as the cutoff value for significant vasoconstriction of middle cerebral arteries based on our previous study. Results Patients with RCVS had a reduced number of CD34+KDR+ cells (0.009 ± 0.006% vs. 0.014 ± 0.010%, p = 0.031) but not KDR+CD133+ cells or CD34+CD133+ EPCs, in comparison with controls. The number of CD34+KDR+ cells was inversely correlated with the Lindegaard index (rs = -0.418, p = 0.047). Of note, compared to controls, patients with a Lindegaard index > 2 (n = 13) had a reduced number of CD34+KDR+ cells (0.007 ± 0.005% vs. 0.014 ± 0.010%, p = 0.010), but those with a Lindegaard index ≤ 2 did not. Conclusions Patients with RCVS had reduced circulating CD34+KDR+ EPCs, which were correlated with the severity of vasoconstriction. Endothelial dysfunction might contribute to the pathogenesis of RCVS. PMID:25466718

  14. The Hemogenic Competence of Endothelial Progenitors Is Restricted by Runx1 Silencing during Embryonic Development.

    PubMed

    Eliades, Alexia; Wareing, Sarah; Marinopoulou, Elli; Fadlullah, Muhammad Z H; Patel, Rahima; Grabarek, Joanna B; Plusa, Berenika; Lacaud, Georges; Kouskoff, Valerie

    2016-06-01

    It is now well-established that hematopoietic stem cells (HSCs) and progenitor cells originate from a specialized subset of endothelium, termed hemogenic endothelium (HE), via an endothelial-to-hematopoietic transition. However, the molecular mechanisms determining which endothelial progenitors possess this hemogenic potential are currently unknown. Here, we investigated the changes in hemogenic potential in endothelial progenitors at the early stages of embryonic development. Using an ETV2::GFP reporter mouse to isolate emerging endothelial progenitors, we observed a dramatic decrease in hemogenic potential between embryonic day (E)7.5 and E8.5. At the molecular level, Runx1 is expressed at much lower levels in E8.5 intra-embryonic progenitors, while Bmi1 expression is increased. Remarkably, the ectopic expression of Runx1 in these progenitors fully restores their hemogenic potential, as does the suppression of BMI1 function. Altogether, our data demonstrate that hemogenic competency in recently specified endothelial progenitors is restrained through the active silencing of Runx1 expression. PMID:27239041

  15. CD133 positive progenitor endothelial cell lines from human cord blood.

    PubMed

    Paprocka, Maria; Krawczenko, Agnieszka; Dus, Danuta; Kantor, Aneta; Carreau, Aude; Grillon, Catherine; Kieda, Claudine

    2011-08-01

    Endothelial progenitor cells (EPCs) modulate postnatal vascularization and contribute to vessel regeneration in adults. Stem cells and progenitor cells were found in umbilical cord blood, bone marrow, and mobilized peripheral blood cells, from where they were isolated and cultured. However, the yield of progenitor cells is usually not sufficient for clinical application and the quality of progenitor cells varies. The aim of the study was the immortalization of early progenitor cells with high proliferative potential, capable to differentiate to EPCs and, further, toward endothelial cells. Two cell lines, namely HEPC-CB.1 and HEPC-CB.2 (human endothelial progenitor cells-cord blood) were isolated. As assessed by specific antibody labeling and flow cytometric analysis, they express a panel of stem cell markers: CD133, CD13, CD271, CD90 and also endothelial cell markers: CD202b, CD309 (VEGFR2), CD146, CD105, and CD143 but they do not present markers of finally differentiated endothelial cells: CD31, vWf, nor CD45 which is a specific hematopoietic cell marker. Using the multiplex Cytometric Bead Assay, the simultaneous production of proangiogenic cytokines IL8, angiogenin, and VEGF was demonstrated in normoxia and was shown to be increased by hypoxia. Both cell lines, similarly as mature endothelial cells, underwent in vitro pre-angiogenic process, formed pseudovessel structures and present an accelerated angiogenesis in hypoxic conditions. To date, these are the first CD133 positive established cell lines from human cord blood cells. PMID:21710642

  16. Effect of Human Endothelial Progenitor Cell (EPC)- or Mouse Vascular Endothelial Growth Factor-Derived Vessel Formation on the Survival of Vitrified/Warmed Mouse Ovarian Grafts

    PubMed Central

    Cha, Soo Kyung; Shin, Dong Hyuk; Kim, Bo Yeun; Yoon, Sook-Young; Yoon, Tae Ki; Lee, Woo Sik

    2014-01-01

    The aim of this study was to evaluate the effectiveness of improving angiogenesis at graft sites on the survival of follicles in transplanted ovarian tissue. Matrigel containing 5 × 105 of cord blood-derived endothelial progenitor cells (EPCs) or 200 ng of mouse vascular endothelial growth factor (VEGF) was injected subcutaneously into BALB/c-Nu mice. After 1 week, vitrified/warmed ovaries from female B6D2F1 mice were subcutaneously transplanted into the injection sites. After 1, 2, and 4 weeks posttransplantation, the ovaries were recovered and subjected to histological analysis. Oocytes were collected from the transplanted ovaries, and their fertilization, embryonic development, and delivery were also observed. Vitrified/warmed ovaries transplanted into EPC- or VEGF-treated sites developed more blood vessels and showed better follicle survival than those transplanted into sham-injected sites. Normal embryonic development and consequent live births were obtained using oocytes recovered from cryopreserved/transplanted ovaries. Treatment with EPCs or VEGF could prevent the ischemic damage during the early revascularization stage of ovarian transplantation. PMID:24401473

  17. Differential Effects of Isoxazole-9 on Neural Stem/Progenitor Cells, Oligodendrocyte Precursor Cells, and Endothelial Progenitor Cells

    PubMed Central

    Maki, Takakuni; Shindo, Akihiro; Osumi, Noriko; Zhao, Jing; Lin, Hong; Holder, Julie C.; Chuang, Tsu Tshen; McNeish, John D.; Arai, Ken; Lo, Eng H.

    2015-01-01

    Adult mammalian brain can be plastic after injury and disease. Therefore, boosting endogenous repair mechanisms would be a useful therapeutic approach for neurological disorders. Isoxazole-9 (Isx-9) has been reported to enhance neurogenesis from neural stem/progenitor cells (NSPCs). However, the effects of Isx-9 on other types of progenitor/precursor cells remain mostly unknown. In this study, we investigated the effects of Isx-9 on the three major populations of progenitor/precursor cells in brain: NSPCs, oligodendrocyte precursor cells (OPCs), and endothelial progenitor cells (EPCs). Cultured primary NSPCs, OPCs, or EPCs were treated with various concentrations of Isx-9 (6.25, 12.5, 25, 50 μM), and their cell numbers were counted in a blinded manner. Isx-9 slightly increased the number of NSPCs and effectively induced neuronal differentiation of NSPCs. However, Isx-9 significantly decreased OPC number in a concentration-dependent manner, suggesting cytotoxicity. Isx-9 did not affect EPC cell number. But in a matrigel assay of angiogenesis, Isx-9 significantly inhibited tube formation in outgrowth endothelial cells derived from EPCs. This potential anti-tube-formation effect of Isx-9 was confirmed in a brain endothelial cell line. Taken together, our data suggest that mechanisms and targets for promoting stem/progenitor cells in the central nervous system may significantly differ between cell types. PMID:26407349

  18. Transdifferentiation of human endothelial progenitors into smooth muscle cells.

    PubMed

    Ji, HaYeun; Atchison, Leigh; Chen, Zaozao; Chakraborty, Syandan; Jung, Youngmee; Truskey, George A; Christoforou, Nicolas; Leong, Kam W

    2016-04-01

    Access to smooth muscle cells (SMC) would create opportunities for tissue engineering, drug testing, and disease modeling. Herein we report the direct conversion of human endothelial progenitor cells (EPC) to induced smooth muscle cells (iSMC) by induced expression of MYOCD. The EPC undergo a cytoskeletal rearrangement resembling that of mesenchymal cells within 3 days post initiation of MYOCD expression. By day 7, the reprogrammed cells show upregulation of smooth muscle markers ACTA2, MYH11, and TAGLN by qRT-PCR and ACTA2 and MYH11 expression by immunofluorescence. By two weeks, they resemble umbilical artery SMC in microarray gene expression analysis. The iSMC, in contrast to EPC control, show calcium transients in response to phenylephrine stimulation and a contractility an order of magnitude higher than that of EPC as determined by traction force microscopy. Tissue-engineered blood vessels constructed using iSMC show functionality with respect to flow- and drug-mediated vasodilation and vasoconstriction. PMID:26874281

  19. Endothelial progenitor cells regenerate infracted myocardium with neovascularisation development.

    PubMed

    Abd El Aziz, M T; Abd El Nabi, E A; Abd El Hamid, M; Sabry, D; Atta, H M; Rahed, L A; Shamaa, A; Mahfouz, S; Taha, F M; Elrefaay, S; Gharib, D M; Elsetohy, Khaled A

    2015-03-01

    We achieved possibility of isolation, characterization human umbilical cord blood endothelial progenitor cells (EPCs), examination potency of EPCs to form new blood vessels and differentiation into cardiomyoctes in canines with acute myocardial infarction (AMI). EPCs were separated and cultured from umbilical cord blood. Their phenotypes were confirmed by uptake of double stains dioctadecyl tetramethylindocarbocyanine-labeled acetylated LDL and FITC-labeled Ulex europaeus agglutinin 1 (DILDL-UEA-1). EPCs of cord blood were counted. Human VEGFR-2 and eNOS from the cultured EPCs were assessed by qPCR. Human EPCs was transplanted intramyocardially in canines with AMI. ECG and cardiac enzymes (CK-MB and Troponin I) were measured to assess severity of cellular damage. Histopathology was done to assess neovascularisation. Immunostaining was done to detect EPCs transdifferentiation into cardiomyocytes in peri-infarct cardiac tissue. qPCR for human genes (hVEGFR-2, and eNOS) was done to assess homing and angiogenic function of transplanted EPCs. Cultured human cord blood exhibited an increased number of EPCs and significant high expression of hVEGFR-2 and eNOS genes in the culture cells. Histopathology showed increased neovascularization and immunostaining showed presence of EPCs newly differentiated into cardiomyocyte-like cells. Our findings suggested that hEPCs can mediate angiogenesis and differentiate into cardiomyoctes in canines with AMI. PMID:25750747

  20. Endothelial progenitor cells regenerate infracted myocardium with neovascularisation development☆

    PubMed Central

    Abd El Aziz, M.T.; Abd El Nabi, E.A.; Abd El Hamid, M.; Sabry, D.; Atta, H.M.; Rahed, L.A.; Shamaa, A.; Mahfouz, S.; Taha, F.M.; Elrefaay, S.; Gharib, D.M.; Elsetohy, Khaled A.

    2013-01-01

    We achieved possibility of isolation, characterization human umbilical cord blood endothelial progenitor cells (EPCs), examination potency of EPCs to form new blood vessels and differentiation into cardiomyoctes in canines with acute myocardial infarction (AMI). EPCs were separated and cultured from umbilical cord blood. Their phenotypes were confirmed by uptake of double stains dioctadecyl tetramethylindocarbocyanine-labeled acetylated LDL and FITC-labeled Ulex europaeus agglutinin 1 (DILDL-UEA-1). EPCs of cord blood were counted. Human VEGFR-2 and eNOS from the cultured EPCs were assessed by qPCR. Human EPCs was transplanted intramyocardially in canines with AMI. ECG and cardiac enzymes (CK-MB and Troponin I) were measured to assess severity of cellular damage. Histopathology was done to assess neovascularisation. Immunostaining was done to detect EPCs transdifferentiation into cardiomyocytes in peri-infarct cardiac tissue. qPCR for human genes (hVEGFR-2, and eNOS) was done to assess homing and angiogenic function of transplanted EPCs. Cultured human cord blood exhibited an increased number of EPCs and significant high expression of hVEGFR-2 and eNOS genes in the culture cells. Histopathology showed increased neovascularization and immunostaining showed presence of EPCs newly differentiated into cardiomyocyte-like cells. Our findings suggested that hEPCs can mediate angiogenesis and differentiate into cardiomyoctes in canines with AMI. PMID:25750747

  1. Hyperoxia, Endothelial Progenitor Cell Mobilization, and Diabetic Wound Healing

    PubMed Central

    Liu, Zhao-Jun

    2008-01-01

    Abstract Diabetic foot disease is a major health problem, which affects 15% of the 200 million patients with diabetes worldwide. Diminished peripheral blood flow and decreased local neovascularization are critical factors that contribute to the delayed or nonhealing wounds in these patients. The correction of impaired local angiogenesis may be a key component in developing therapeutic protocols for treating chronic wounds of the lower extremity and diabetic foot ulcers. Endothelial progenitor cells (EPCs) are the key cellular effectors of postnatal neovascularization and play a central role in wound healing, but their circulating and wound-level numbers are decreased in diabetes, implicating an abnormality in EPC mobilization and homing mechanisms. The deficiency in EPC mobilization is presumably due to impairment of eNOS-NO cascade in bone marrow (BM). Hyperoxia, induced by a clinically relevant hyperbaric oxygen therapy (HBO) protocol, can significantly enhance the mobilization of EPCs from the BM into peripheral blood. However, increased circulating EPCs failed to reach to wound tissues. This is partly a result of downregulated production of SDF-1α in local wound lesions with diabetes. Administration of exogenous SDF-1α into wounds reversed the EPC homing impairment and, with hyperoxia, synergistically enhanced EPC mobilization, homing, neovascularization, and wound healing. Antioxid. Redox Signal. 10, 1869–1882. PMID:18627349

  2. Estrogen Stimulates Homing of Endothelial Progenitor Cells to Endometriotic Lesions.

    PubMed

    Rudzitis-Auth, Jeannette; Nenicu, Anca; Nickels, Ruth M; Menger, Michael D; Laschke, Matthias W

    2016-08-01

    The incorporation of endothelial progenitor cells (EPCs) into microvessels contributes to the vascularization of endometriotic lesions. Herein, we analyzed whether this vasculogenic process is regulated by estrogen. Estrogen- and vehicle-treated human EPCs were analyzed for migration and tube formation. Endometriotic lesions were induced in irradiated FVB/N mice, which were reconstituted with bone marrow from FVB/N-TgN (Tie2/green fluorescent protein) 287 Sato mice. The animals were treated with 100 μg/kg β-estradiol 17-valerate or vehicle (control) over 7 and 28 days. Lesion growth, cyst formation, homing of green fluorescent protein(+)/Tie2(+) EPCs, vascularization, cell proliferation, and apoptosis were analyzed by high-resolution ultrasonography, caliper measurements, histology, and immunohistochemistry. Numbers of blood circulating EPCs were assessed by flow cytometry. In vitro, estrogen-treated EPCs exhibited a higher migratory and tube-forming capacity when compared with controls. In vivo, numbers of circulating EPCs were not affected by estrogen. However, estrogen significantly increased the number of EPCs incorporated into the lesions' microvasculature, resulting in an improved early vascularization. Estrogen further stimulated the growth of lesions, which exhibited massively dilated glands with a flattened layer of stroma. This was mainly because of an increased glandular secretory activity, whereas cell proliferation and apoptosis were not markedly affected. These findings indicate that vasculogenesis in endometriotic lesions is dependent on estrogen, which adds a novel hormonally regulated mechanism to the complex pathophysiology of endometriosis. PMID:27315780

  3. CD34 expression modulates tube-forming capacity and barrier properties of peripheral blood-derived endothelial colony-forming cells (ECFCs).

    PubMed

    Tasev, Dimitar; Konijnenberg, Lara S F; Amado-Azevedo, Joana; van Wijhe, Michiel H; Koolwijk, Pieter; van Hinsbergh, Victor W M

    2016-07-01

    Endothelial colony-forming cells (ECFC) are grown from circulating CD34(+) progenitors present in adult peripheral blood, but during in vitro expansion part of the cells lose CD34. To evaluate whether the regulation of CD34 characterizes the angiogenic phenotypical features of PB-ECFCs, we investigated the properties of CD34(+) and CD34(-) ECFCs with respect to their ability to form capillary-like tubes in 3D fibrin matrices, tip-cell gene expression, and barrier integrity. Selection of CD34(+) and CD34(-) ECFCs from subcultured ECFCs was accomplished by magnetic sorting (FACS: CD34(+): 95 % pos; CD34(-): 99 % neg). Both fractions proliferated at same rate, while CD34(+) ECFCs exhibited higher tube-forming capacity and tip-cell gene expression than CD3(4-) cells. However, during cell culture CD34(-) cells re-expressed CD34. Cell-seeding density, cell-cell contact formation, and serum supplements modulated CD34 expression. CD34 expression in ECFCs was strongly suppressed by newborn calf serum. Stimulation with FGF-2, VEGF, or HGF prepared in medium supplemented with 3 % albumin did not change CD34 mRNA or surface expression. Silencing of CD34 with siRNA resulted in strengthening of cell-cell contacts and increased barrier function of ECFC monolayers as measured by ECIS. Furthermore, CD34 siRNA reduced tube formation by ECFC, but did not affect tip-cell gene expression. These findings demonstrate that CD34(+) and CD34(-) cells are different phenotypes of similar cells and that CD34 (1) can be regulated in ECFC; (2) is positively involved in capillary-like sprout formation; (3) is associated but not causally related to tip-cell gene expression; and (4) can affect endothelial barrier function. PMID:27043316

  4. Rapamycin inhibits re-endothelialization after percutaneous coronary intervention by impeding the proliferation and migration of endothelial cells and inducing apoptosis of endothelial progenitor cells.

    PubMed

    Liu, Hai-Tao; Li, Fei; Wang, Wen-Yong; Li, Xiao-Jing; Liu, Yi-Meng; Wang, Rui-An; Guo, Wen-Yi; Wang, Hai-Chang

    2010-01-01

    Endothelial-cell function is important in the healing of damaged endothelium after percutaneous coronary artery damage. In 3 different animal models, we sought to determine whether rapamycin (sirolimus) affects the proliferation and migration of endothelial cells and endothelial progenitor cells. First, after we implanted stents in dogs, we found that re-endothelialization was impeded more by drug-eluting stents than by bare-metal stents, 30 days after percutaneous coronary intervention. Second, in vitro in rats, we found that 1-100 ng/mL of rapamycin time- and dose-dependently inhibited proliferation over 72 hr (with effects evident as early as 24 hr) and also dose-dependently induced endothelial progenitor-cell apoptosis. Finally, in vivo in rats, we observed that vascular endothelial growth factor expression was decreased after 5 days of rapamycin treatment. We conclude that rapamycin impedes re-endothelialization after drug-eluting stent implantation by inhibiting the proliferation and migration of coronary endothelial cells, inducing endothelial progenitor-cell apoptosis, and decreasing vascular endothelial growth factor expression in the circulation. PMID:20401293

  5. Neutrophil Elastase-Generated Fragment of Vascular Endothelial Growth Factor-A Stimulates Macrophage and Endothelial Progenitor Cell Migration

    PubMed Central

    Kurtagic, Elma; Rich, Celeste B.; Buczek-Thomas, Jo Ann; Nugent, Matthew A.

    2015-01-01

    Elastase released from neutrophils as part of the innate immune system has been implicated in chronic diseases such as emphysema and cardiovascular disease. We have previously shown that neutrophil elastase targets vascular endothelial growth factor-A (VEGF) for partial degradation to generate a fragment of VEGF (VEGFf) that has distinct activities. Namely, VEGFf binds to VEGF receptor 1 but not to VEGF receptor 2 and shows altered signaling compared to intact VEGF. In the present study we investigated the chemotactic function of VEGF and VEGFf released from cells by neutrophil elastase. We found that endothelial cells migrated in response to intact VEGF but not VEGFf whereas RAW 264.7 macrophages/monocytes and embryonic endothelial progenitor cells were stimulated to migrate by either VEGF or VEGFf. To investigate the role of elastase-mediated release of VEGF from cells/extracellular matrices, a co-culture system was established. High or low VEGF producing cells were co-cultured with macrophages, endothelial or endothelial progenitor cells and treated with neutrophil elastase. Elastase treatment stimulated macrophage and endothelial progenitor cell migration with the response being greater with the high VEGF expressing cells. However, elastase treatment led to decreased endothelial cell migration due to VEGF cleavage to VEGF fragment. These findings suggest that the tissue response to NE-mediated injury might involve the generation of diffusible VEGF fragments that stimulate inflammatory cell recruitment. PMID:26672607

  6. Long-Term Expansion in Platelet Lysate Increases Growth of Peripheral Blood-Derived Endothelial-Colony Forming Cells and Their Growth Factor-Induced Sprouting Capacity

    PubMed Central

    Tasev, Dimitar; van Wijhe, Michiel H.; Weijers, Ester M.; van Hinsbergh, Victor W. M.; Koolwijk, Pieter

    2015-01-01

    Introduction Efficient implementation of peripheral blood-derived endothelial-colony cells (PB-ECFCs) as a therapeutical tool requires isolation and generation of a sufficient number of cells in ex vivo conditions devoid of animal-derived products. At present, little is known how the isolation and expansion procedure in xenogeneic-free conditions affects the therapeutical capacity of PB-ECFCs. Results The findings presented in this study indicate that human platelet lysate (PL) as a serum substitute yields twice more colonies per mL blood compared to the conventional isolation with fetal bovine serum (FBS). Isolated ECFCs displayed a higher proliferative ability in PL supplemented medium than cells in FBS medium during 30 days expansion. The cells at 18 cumulative population doubling levels (CPDL) retained their proliferative capacity, showed higher sprouting ability in fibrin matrices upon stimulation with FGF-2 and VEGF-A than the cells at 6 CPDL, and displayed low β-galactosidase activity. The increased sprouting of PB-ECFCs at 18 CPDL was accompanied by an intrinsic activation of the uPA/uPAR fibrinolytic system. Induced deficiency of uPA (urokinase-type plasminogen activator) or uPAR (uPA receptor) by siRNA technology completely abolished the angiogenic ability of PB-ECFCs in fibrin matrices. During the serial expansion, the gene induction of the markers associated with inflammatory activation such as VCAM-1 and ICAM-1 did not occur or only to limited extent. While further propagation up to 31 CPDL proceeded at a comparable rate, a marked upregulation of inflammatory markers occurred in all donors accompanied by a further increase of uPA/uPAR gene induction. The observed induction of inflammatory genes at later stages of long-term propagation of PB-ECFCs underpins the necessity to determine the right time-point for harvesting of sufficient number of cells with preserved therapeutical potential. Conclusion The presented isolation method and subsequent cell

  7. Isolation of Foreign Material-Free Endothelial Progenitor Cells Using CD31 Aptamer and Therapeutic Application for Ischemic Injury

    PubMed Central

    Heo, Soon Chul; Kwon, Yang Woo; Choi, Eun Jung; Bae, Kwang-Hee; Suh, Dong-Soo; Kim, Seung-Chul; Han, Seungmin; Haam, Seungjoo; Jung, Jongha; Kim, Kiseok; Ryu, Sung Ho; Kim, Jae Ho

    2015-01-01

    Endothelial progenitor cells (EPCs) can be isolated from human bone marrow or peripheral blood and reportedly contribute to neovascularization. Aptamers are 40-120-mer nucleotides that bind to a specific target molecule, as antibodies do. To utilize apatmers for isolation of EPCs, in the present study, we successfully generated aptamers that recognize human CD31, an endothelial cell marker. CD31 aptamers bound to human umbilical cord blood-derived EPCs and showed specific interaction with human CD31, but not with mouse CD31. However, CD31 aptamers showed non-specific interaction with CD31-negative 293FT cells and addition of polyanionic competitor dextran sulfate eliminated non-specific interaction without affecting cell viability. From the mixture of EPCs and 293FT cells, CD31 aptamers successfully isolated EPCs with 97.6% purity and 94.2% yield, comparable to those from antibody isolation. In addition, isolated EPCs were decoupled from CD31 aptamers with a brief treatment of high concentration dextran sulfate. EPCs isolated with CD31 aptamers and subsequently decoupled from CD31 aptamers were functional and enhanced the restoration of blood flow when transplanted into a murine hindlimb ischemia model. In this study, we demonstrated isolation of foreign material-free EPCs, which can be utilized as a universal protocol in preparation of cells for therapeutic transplantation. PMID:26148001

  8. The effect of Heparin-VEGF multilayer on the biocompatibility of decellularized aortic valve with platelet and endothelial progenitor cells.

    PubMed

    Ye, Xiaofeng; Wang, Haozhe; Zhou, Jingxin; Li, Haiqing; Liu, Jun; Wang, Zhe; Chen, Anqing; Zhao, Qiang

    2013-01-01

    The application of polyelectrolyte multilayer films is a new, versatile approach to surface modification of decellularized tissue, which has the potential to greatly enhance the functionality of engineered tissue constructs derived from decellularized organs. In the present study, we test the hypothesis that Heparin- vascular endothelial growth factor (VEGF) multilayer film can not only act as an antithrombotic coating reagent, but also induce proliferation of endothelial progenitor cells (EPCs) on the decellularized aortic heart valve. SEM demonstrated the adhesion and geometric deformation of platelets. The quantitative assay of platelet activation was determined by measuring the production of soluble P-selectin. Binding and subsequent release of heparin and VEGF from valve leaflets were assessed qualitatively by laser confocal scanning microscopy and quantitatively by ELISA methods. Human blood derived EPCs were cultured and the adhesion and growth of EPCs on the surface modified valvular scaffolds were assessed. The results showed that Heparin-VEGF multilayer film improved decellularized valve haemocompatibility with respect to a substantial reduction of platelet adhesion. Release of VEGF from the decellularized heart valve leaflets at physiological conditions was sustained over 5 days. In vitro biological tests demonstrated that EPCs achieved better adhesion, proliferation and migration on the coatings with Heparin-VEGF multilayer film. Combined, these results indicate that Heparin-VEGF multilayer film could be used to cover the decellularized porcine aortic valve to decrease platelet adhesion while exhibiting excellent EPCs biocompatibility. PMID:23359625

  9. Endothelial cells are progenitors of cardiac pericytes and vascular smooth muscle cells

    PubMed Central

    Chen, Qi; Zhang, Hui; Liu, Yang; Adams, Susanne; Eilken, Hanna; Stehling, Martin; Corada, Monica; Dejana, Elisabetta; Zhou, Bin; Adams, Ralf H.

    2016-01-01

    Mural cells of the vessel wall, namely pericytes and vascular smooth muscle cells, are essential for vascular integrity. The developmental sources of these cells and molecular mechanisms controlling their progenitors in the heart are only partially understood. Here we show that endocardial endothelial cells are progenitors of pericytes and vascular smooth muscle cells in the murine embryonic heart. Endocardial cells undergo endothelial–mesenchymal transition and convert into primitive mesenchymal progenitors expressing the platelet-derived growth factor receptors, PDGFRα and PDGFRβ. These progenitors migrate into the myocardium, differentiate and assemble the wall of coronary vessels, which requires canonical Wnt signalling involving Frizzled4, β-catenin and endothelial cell-derived Wnt ligands. Our findings identify a novel and unexpected population of progenitors for coronary mural cells with potential relevance for heart function and disease conditions. PMID:27516371

  10. Endothelial progenitor cells and asymmetric dimethylarginine after renal transplantation.

    PubMed

    Teplan, Vladimír; Mahrová, Andrea; Králová-Lesná, Ivana; Racek, Jaroslav; Valkovský, Ivo; Štollová, Milena

    2015-03-01

    Levels of the endogenous nitric oxide synthase inhibitor asymmetrical dimethylarginine (ADMA) are elevated and endothelial progenitor cells (EPCs) decreased in patients undergoing renal transplantation (Tx) and may contribute to cardiovascular complications. In this study, we tested the hypothesis that elevated ADMA and decreased EPC can be positively influenced with regular physical exercise early after Tx. Blood samples for analysis of ADMA and EPC were obtained from randomly selected 64 patients after Tx who agreed to participate in a supervised aerobic exercise program for 6 months (group I). Samples were collected before the training began, 1 month after surgery (with stabilized renal function), and at 6 months after initiation. Sixty-two age, sex, human leukocyte antigens (HLA) typing, duration of previous dialysis, history of cardiovascular disease, and immunosupression regimen-matched transplant patients who did not exercise regularly were examined as controls (group II). There were no differences in ADMA levels and EPC count between both groups before the training program began. After 6 months of exercise, ADMA concentration in the group I decreased (3.50 ± 0.45 vs. 2.11 ± 0.35 μmol/L; P < .01) and was also lower comparing with group II (2.11 ± 0.23 vs. 3.25 ± 0.35 μmol/L; P < .01). In the same period, EPC cells increased from 2.085 ± 650 cells/mL versus 3.991 ± 560 cells/mL, P < .01 in group I; but in group II, changes were nonsignificant (P = .11). Blood lipids, HbA1c, insulin, and systolic blood pressure were also affected by the training program. Elevated ADMA level and decreased EPC count were significantly influenced by early regular exercise in patients after Tx. PMID:25576240

  11. The Novel Methods for Analysis of Exosomes Released from Endothelial Cells and Endothelial Progenitor Cells

    PubMed Central

    Wang, Jinju; Guo, Runmin; Yang, Yi; Jacobs, Bradley; Chen, Suhong; Iwuchukwu, Ifeanyi; Gaines, Kenneth J.; Chen, Yanfang; Simman, Richard; Lv, Guiyuan; Wu, Keng; Bihl, Ji C.

    2016-01-01

    Exosomes (EXs) are cell-derived vesicles that mediate cell-cell communication and could serve as biomarkers. Here we described novel methods for purification and phenotyping of EXs released from endothelial cells (ECs) and endothelial progenitor cells (EPCs) by combining microbeads and fluorescence quantum dots (Q-dots®) techniques. EXs from the culture medium of ECs and EPCs were isolated and detected with cell-specific antibody conjugated microbeads and second antibody conjugated Q-dots by using nanoparticle tracking analysis (NTA) system. The sensitivities of the cell origin markers for ECs (CD105, CD144) and EPCs (CD34, KDR) were evaluated. The sensitivity and specificity were determined by using positive and negative markers for EXs (CD63), platelets (CD41), erythrocytes (CD235a), and microvesicles (Annexin V). Moreover, the methods were further validated in particle-free plasma and patient samples. Results showed that anti-CD105/anti-CD144 and anti-CD34/anti-KDR had the highest sensitivity and specificity for isolating and detecting EC-EXs and EPC-EXs, respectively. The methods had the overall recovery rate of over 70% and were able to detect the dynamical changes of circulating EC-EXs and EPC-EXs in acute ischemic stroke. In conclusion, we have developed sensitive and specific microbeads/Q-dots fluorescence NTA methods for EC-EX and EPC-EX isolation and detection, which will facilitate the functional study and biomarker discovery. PMID:27118976

  12. Concise Review: Ex Vivo Expansion of Cord Blood-Derived Hematopoietic Stem and Progenitor Cells: Basic Principles, Experimental Approaches, and Impact in Regenerative Medicine

    PubMed Central

    Flores-Guzmán, Patricia; Fernández-Sánchez, Verónica

    2013-01-01

    Hematopoietic stem cells (HSCs) and hematopoietic progenitor cells (HPCs) play key roles in the production of mature blood cells and in the biology and clinical outcomes of hematopoietic transplants. The numbers of these cells, however, are extremely low, particularly in umbilical cord blood (UCB); thus, ex vivo expansion of human UCB-derived HSCs and HPCs has become a priority in the biomedical field. Expansion of progenitor cells can be achieved by culturing such cells in the presence of different combinations of recombinant stimulatory cytokines; in contrast, expansion of actual HSCs has proved to be more difficult because, in addition to needing recombinant cytokines, HSCs seem to deeply depend on the presence of stromal cells and/or elements that promote the activation of particular self-renewal signaling pathways. Hence, there is still controversy regarding the optimal culture conditions that should be used to achieve this. To date, UCB transplants using ex vivo-expanded cells have already been performed for the treatment of different hematological disorders, and although results are still far from being optimal, the advances are encouraging. Recent studies suggest that HSCs may also give rise to nonhematopoietic cells, such as neural, cardiac, mesenchymal, and muscle cells. Such plasticity and the possibility of producing nonhematopoietic cells at the clinical scale could bring new alternatives for the treatment of neural, metabolic, orthopedic, cardiac, and neoplastic disorders. Once standardized, ex vivo expansion of human HSCs/HPCs will surely have a positive impact in regenerative medicine. PMID:24101670

  13. Impairment of endothelial progenitor cell function and vascularization capacity by aldosterone in mice and humans

    PubMed Central

    Thum, Thomas; Schmitter, Kerstin; Fleissner, Felix; Wiebking, Volker; Dietrich, Bernd; Widder, Julian D.; Jazbutyte, Virginija; Hahner, Stefanie; Ertl, Georg; Bauersachs, Johann

    2011-01-01

    Aims Hyperaldosteronism is associated with vascular injury and increased cardiovascular events. Bone marrow-derived endothelial progenitor cells (EPCs) play an important role in endothelial repair and vascular homeostasis. We hypothesized that hyperaldosteronism impairs EPC function and vascularization capacity in mice and humans. Methods and results We characterized the effects of aldosterone and mineralocorticoid receptor (MR) blockade on EPC number and function as well as vascularization capacity and endothelial function. Treatment of human EPC with aldosterone induced translocation of the MR and impaired multiple cellular functions of EPC, such as differentiation, migration, and proliferation in vitro. Impaired EPC function was rescued by pharmacological blockade or genetic ablation of the MR. Aldosterone protein kinase A (PKA) dependently increased reactive oxygen species formation in EPC. Aldosterone infusion in mice impaired EPC function, EPC homing to vascular structures and vascularization capacity in a MR-dependent but blood pressure-independent manner. Endothelial progenitor cells from patients with primary hyperaldosteronism compared with controls of similar age displayed reduced migratory potential. Impaired EPC function was associated with endothelial dysfunction. MR blockade in patients with hyperaldosteronism improved EPC function and arterial stiffness. Conclusion Endothelial progenitor cells express a MR that mediates functional impairment by PKA-dependent increase of reactive oxygen species. Normalization of EPC function may represent a novel mechanism contributing to the beneficial effects of MR blockade in cardiovascular disease prevention and treatment. PMID:20926363

  14. PRC2 inhibition counteracts the culture-associated loss of engraftment potential of human cord blood-derived hematopoietic stem and progenitor cells

    PubMed Central

    Varagnolo, Linda; Lin, Qiong; Obier, Nadine; Plass, Christoph; Dietl, Johannes; Zenke, Martin; Claus, Rainer; Müller, Albrecht M.

    2015-01-01

    Cord blood hematopoietic stem cells (CB-HSCs) are an outstanding source for transplantation approaches. However, the amount of cells per donor is limited and culture expansion of CB-HSCs is accompanied by a loss of engraftment potential. In order to analyze the molecular mechanisms leading to this impaired potential we profiled global and local epigenotypes during the expansion of human CB hematopoietic stem and progenitor cells (HPSCs). Human CB-derived CD34+ cells were cultured in serum-free medium together with SCF, TPO, FGF, with or without Igfbp2 and Angptl5 (STF/STFIA cocktails). As compared to the STF cocktail, the STFIA cocktail maintains in vivo repopulation capacity of cultured CD34+ cells. Upon expansion, CD34+ cells genome-wide remodel their epigenotype and depending on the cytokine cocktail, cells show different H3K4me3 and H3K27me3 levels. Expanding cells without Igfbp2 and Angptl5 leads to higher global H3K27me3 levels. ChIPseq analyses reveal a cytokine cocktail-dependent redistribution of H3K27me3 profiles. Inhibition of the PRC2 component EZH2 counteracts the culture-associated loss of NOD scid gamma (NSG) engraftment potential. Collectively, our data reveal chromatin dynamics that underlie the culture-associated loss of engraftment potential. We identify PRC2 component EZH2 as being involved in the loss of engraftment potential during the in vitro expansion of HPSCs. PMID:26198814

  15. Smooth muscle progenitor cells from peripheral blood promote the neovascularization of endothelial colony-forming cells

    SciTech Connect

    Joo, Hyung Joon; Seo, Ha-Rim; Jeong, Hyo Eun; Choi, Seung-Cheol; Park, Jae Hyung; Yu, Cheol Woong; Hong, Soon Jun; Chung, Seok; Lim, Do-Sun

    2014-07-11

    Highlights: • Two distinct vascular progenitor cells are induced from adult peripheral blood. • ECFCs induce vascular structures in vitro and in vivo. • SMPCs augment the in vitro and in vivo angiogenic potential of ECFCs. • Both cell types have synergistic therapeutic potential in ischemic hindlimb model. - Abstract: Proangiogenic cell therapy using autologous progenitors is a promising strategy for treating ischemic disease. Considering that neovascularization is a harmonized cellular process that involves both endothelial cells and vascular smooth muscle cells, peripheral blood-originating endothelial colony-forming cells (ECFCs) and smooth muscle progenitor cells (SMPCs), which are similar to mature endothelial cells and vascular smooth muscle cells, could be attractive cellular candidates to achieve therapeutic neovascularization. We successfully induced populations of two different vascular progenitor cells (ECFCs and SMPCs) from adult peripheral blood. Both progenitor cell types expressed endothelial-specific or smooth muscle-specific genes and markers, respectively. In a protein array focused on angiogenic cytokines, SMPCs demonstrated significantly higher expression of bFGF, EGF, TIMP2, ENA78, and TIMP1 compared to ECFCs. Conditioned medium from SMPCs and co-culture with SMPCs revealed that SMPCs promoted cell proliferation, migration, and the in vitro angiogenesis of ECFCs. Finally, co-transplantation of ECFCs and SMPCs induced robust in vivo neovascularization, as well as improved blood perfusion and tissue repair, in a mouse ischemic hindlimb model. Taken together, we have provided the first evidence of a cell therapy strategy for therapeutic neovascularization using two different types of autologous progenitors (ECFCs and SMPCs) derived from adult peripheral blood.

  16. Humoral Activity of Cord Blood-Derived Stem/Progenitor Cells: Implications for Stem Cell-Based Adjuvant Therapy of Neurodegenerative Disorders

    PubMed Central

    Paczkowska, Edyta; Kaczyńska, Katarzyna; Pius-Sadowska, Ewa; Rogińska, Dorota; Kawa, Miłosz; Ustianowski, Przemysław; Safranow, Krzysztof; Celewicz, Zbigniew; Machaliński, Bogusław

    2013-01-01

    Background Stem/progenitor cells (SPCs) demonstrate neuro-regenerative potential that is dependent upon their humoral activity by producing various trophic factors regulating cell migration, growth, and differentiation. Herein, we compared the expression of neurotrophins (NTs) and their receptors in specific umbilical cord blood (UCB) SPC populations, including lineage-negative, CD34+, and CD133+ cells, with that in unsorted, nucleated cells (NCs). Methods and Results The expression of NTs and their receptors was detected by QRT-PCR, western blotting, and immunofluorescent staining in UCB-derived SPC populations (i.e., NCs vs. lineage-negative, CD34+, and CD133+ cells). To better characterize, global gene expression profiles of SPCs were determined using genome-wide RNA microarray technology. Furthermore, the intracellular production of crucial neuro-regenerative NTs (i.e., BDNF and NT-3) was assessed in NCs and lineage-negative cells after incubation for 24, 48, and 72 h in both serum and serum-free conditions. We discovered significantly higher expression of NTs and NT receptors at both the mRNA and protein level in lineage-negative, CD34+, and CD133+ cells than in NCs. Global gene expression analysis revealed considerably higher expression of genes associated with the production and secretion of proteins, migration, proliferation, and differentiation in lineage-negative cells than in CD34+ or CD133+ cell populations. Notably, after short-term incubation under serum-free conditions, lineage-negative cells and NCs produced significantly higher amounts of BDNF and NT-3 than under steady-state conditions. Finally, conditioned medium (CM) from lineage-negative SPCs exerted a beneficial impact on neural cell survival and proliferation. Conclusions Collectively, our findings demonstrate that UCB-derived SPCs highly express NTs and their relevant receptors under steady-state conditions, NT expression is greater under stress-related conditions and that CM from SPCs

  17. The Acute Exposure Effects of Inhaled Nickel Nanoparticles on Murine Endothelial Progenitor Cells

    PubMed Central

    Liberda, Eric N; Cuevas, Azita K; Qu, Qingshan; Chen, Lung Chi

    2014-01-01

    Introduction The discovery of endothelial progenitor cells (EPCs) may help to explain observed cardiovascular effects associated with inhaled nickel nanoparticle exposures such as increases in vascular inflammation, generate reactive oxygen species, alter vasomotor tone, and potentiated atherosclerosis in murine species. Methods Following an acute whole body inhalation exposure to 500μg/m3 of nickel nanoparticles for 5 hrs, bone marrow EPCs from C57BL/6 mice were isolated. EPCs were harvested for their RNA or used in a variety of assays including chemotaxis, tube formation, and proliferation. Gene expression was assessed for important receptors involved in EPC mobilization and homing using RT-PCR methods. EPCs, circulating endothelial progenitor cells (CEPCs), circulating endothelial cells (CECs), and endothelial microparticles (EMPs) were quantified on a BD FACSCalibur to examine endothelial damage and repair associated with the exposure. Results and Conclusions Acute exposure to inhaled nickel nanoparticles significantly increased both bone marrow EPCs as well as their levels in circulation (CEPCs). CECs were significantly elevated indicating that endothelial damage occurred due to the exposure. There was no significant difference in EMPs between the two groups. Tube formation and chemotaxis, but not proliferation, of bone marrow EPCs was impaired in the nickel nanoparticle exposed group. These results coincided with a decrease in the mRNA of receptors involved in EPC mobilization and homing. This data provides new insight into how an acute nickel nanoparticle exposure to half of the current Occupational Safety & Health Administration permissible exposure limit may adversely affect EPCs. PMID:25144474

  18. Progress and prospects of endothelial progenitor cell therapy in coronary stent implantation.

    PubMed

    Lan, Hualin; Wang, Yi; Yin, Tieyin; Wang, Yazhou; Liu, Wanqian; Zhang, Xiaojuan; Yu, Qinsong; Wang, Zhaoxu; Wang, Guixue

    2016-08-01

    Drug-eluting stents (DES) have been widely used to treat coronary artery disease (CAD) since their clinical use has significantly reduced the occurrence of in-stent restenosis (ISR) as compared with the initially applied bare-metal stents (BMS). However, analyses of long-term clinical outcome have raised concerns about the serious safety problem of DES, such as ISR caused by late or very late thrombosis. Various studies showed that those complications were associated with vascular endothelial injury/dysfunction or endothelialization delaying. Recently, through biological characterization of endothelial progenitor cells (EPCs), mechanistic understanding of rapid re-endothelialization of the vascular injury sites after coronary stenting has become possible and is a new research hotspot in the prevention of ISR and late/very late stent thrombosis. It has been well recognized that the formation of a functional endothelial layer from EPCs requires a coordinated sequence of multistep and signaling events, which includes cell mobilization, adhesion, migration and finally the differentiation to vascular endothelial cells (VECs). In this review, we summarize and discuss the currently relevant information about EPCs, the mechanism of DES interfering with the natural vascular healing process in preventing or delaying the formation of a functional endothelial layer, and EPCs-mediated acceleration of re-endothelialization at vascular injury sites. © 2015 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 104B: 1237-1247, 2016. PMID:26059710

  19. Androgen Modulates Functions of Endothelial Progenitor Cells through Activated Egr1 Signaling

    PubMed Central

    Ye, Yizhou; Li, Xizhe; Zhang, You; Shen, Zhenya; Yang, Junjie

    2016-01-01

    Researches show that androgens have important effects on migration of endothelial cells and endothelial protection in coronary heart disease. Endothelial progenitor cells (EPCs) as a progenitor cell type that can differentiate into endothelial cells, have a critical role in angiogenesis and endothelial protection. The relationship between androgen and the functions of EPCs has animated much interest and controversy. In this study, we investigated the angiogenic and migratory functions of EPCs after treatment by dihydrotestosterone (DHT) and the molecular mechanisms as well. We found that DHT treatment enhanced the incorporation of EPCs into tubular structures formed by HUVECs and the migratory activity of EPCs in the transwell assay dose dependently. Moreover, microarray analysis was performed to explore how DHT changes the gene expression profiles of EPCs. We found 346 differentially expressed genes in androgen-treated EPCs. Angiogenesis-related genes like Egr-1, Vcan, Efnb2, and Cdk2ap1 were identified to be regulated upon DHT treatment. Furthermore, the enhanced angiogenic and migratory abilities of EPCs after DHT treatment were inhibited by Egr1-siRNA transfection. In conclusion, our findings suggest that DHT markedly enhances the vessel forming ability and migration capacity of EPCs. Egr1 signaling may be a possible pathway in this process. PMID:26697079

  20. Dysregulation of Vascular Endothelial Progenitor Cells Lung-Homing in Subjects with COPD

    PubMed Central

    Salter, Brittany M.; Manzoor, Fizza; Beaudin, Suzanne; Kjarsgaard, Melanie; Nair, Parameswaran; Gauvreau, Gail M.; Sehmi, Roma

    2016-01-01

    Chronic obstructive pulmonary disease (COPD) is characterized by fixed airflow limitation and progressive decline of lung function and punctuated by occasional exacerbations. The disease pathogenesis may involve activation of the bone marrow stimulating mobilization and lung-homing of progenitor cells. We investigated the hypothesis that lower circulating numbers of vascular endothelial progenitor cells (VEPCs) are a consequence of increased lung-sequestration in COPD. Nonatopic, current or ex-smokers with diagnosed COPD and nonatopic, nonsmoking normal controls were enrolled. Blood and induced sputum extracted primitive hemopoietic progenitors (HPCs) and VEPC were enumerated by flow cytometry. Migration and adhesive responses to fibronectin were assessed. In sputum, VEPC numbers were significantly greater in COPD compared to normal controls. In blood, VEPCs were significantly lower in COPD versus normal controls. There were no differences in HPC levels between the two groups in either compartment. Functionally, there was a greater migrational responsiveness of progenitors from COPD subjects to stromal cell-derived factor-1alpha (SDF-1α) compared to normal controls. This was associated with greater numbers of CXCR4+ progenitors in sputum from COPD. Increased migrational responsiveness of progenitor cells may promote lung-homing of VEPC in COPD which may disrupt maintenance and repair of the airways and contribute to COPD disease pathogenesis. PMID:27445517

  1. Hydrogel Surfaces to Promote Attachment and Spreading of Endothelial Progenitor Cells

    PubMed Central

    Camci-Unal, Gulden; Nichol, Jason William; Bae, Hojae; Tekin, Halil; Bischoff, Joyce; Khademhosseini, Ali

    2011-01-01

    Endothelialization of artificial vascular grafts is a challenging process in cardiovascular tissue engineering. Functionalized biomaterials could be promising candidates to promote endothelialization in repair of cardiovascular injuries. The purpose of this study was to synthesize hyaluronic acid (HA) and heparin based hydrogels that could promote adhesion and spreading of endothelial progenitor cells (EPCs). We report that the addition of heparin into HA-based hydrogels provides an attractive surface for EPCs promoting spreading and the formation of an endothelial monolayer on the hydrogel surface. To increase EPC adhesion and spreading, we covalently immobilized CD34 antibody (Ab) on HA-heparin hydrogels using standard EDC/NHS amine coupling strategies. We found that EPC adhesion and spreading on CD34 Ab immobilized HA-heparin hydrogels was significantly higher than their nonmodified analogs. Once adhered, EPCs spread and formed an endothelial layer on both nonmodified and CD34 Ab modified HA-heparin hydrogels after 3 days of culture. We did not observe significant adhesion and spreading when heparin was not included in the control hydrogels. In addition to EPCs, we also used human umbilical cord vein endothelial cells (HUVECs), which adhered and spread on HA-heparin hydrogels. Macrophages exhibited significantly less adhesion compared to EPCs on the same hydrogels. This composite material could possibly be used to develop surface coatings for artificial cardiovascular implants, due to its specificity for EPC and endothelial cells on an otherwise non-thrombogenic surface. PMID:22223475

  2. Hydrogel surfaces to promote attachment and spreading of endothelial progenitor cells.

    PubMed

    Camci-Unal, Gulden; Nichol, Jason William; Bae, Hojae; Tekin, Halil; Bischoff, Joyce; Khademhosseini, Ali

    2013-05-01

    Endothelialization of artificial vascular grafts is a challenging process in cardiovascular tissue engineering. Functionalized biomaterials could be promising candidates to promote endothelialization in repair of cardiovascular injuries. The purpose of this study was to synthesize hyaluronic acid (HA) and heparin-based hydrogels that could promote adhesion and spreading of endothelial progenitor cells (EPCs). We report that the addition of heparin into HA-based hydrogels provides an attractive surface for EPCs promoting spreading and the formation of an endothelial monolayer on the hydrogel surface. To increase EPC adhesion and spreading, we covalently immobilized CD34 antibody (Ab) on HA-heparin hydrogels, using standard EDC/NHS amine-coupling strategies. We found that EPC adhesion and spreading on CD34 Ab-immobilized HA-heparin hydrogels was significantly higher than their non-modified analogues. Once adhered, EPCs spread and formed an endothelial layer on both non-modified and CD34 Ab-modified HA-heparin hydrogels after 3 days of culture. We did not observe significant adhesion and spreading when heparin was not included in the control hydrogels. In addition to EPCs, we also used human umbilical cord vein endothelial cells (HUVECs), which adhered and spread on HA-heparin hydrogels. Macrophages exhibited significantly less adhesion compared to EPCs on the same hydrogels. This composite material could possibly be used to develop surface coatings for artificial cardiovascular implants, due to its specificity for EPC and endothelial cells on an otherwise non-thrombogenic surface. PMID:22223475

  3. Autologous Endothelial Progenitor Cell-Seeding Technology and Biocompatibility Testing For Cardiovascular Devices in Large Animal Model

    PubMed Central

    Jantzen, Alexandra E.; Lane, Whitney O.; Gage, Shawn M.; Haseltine, Justin M.; Galinat, Lauren J.; Jamiolkowski, Ryan M.; Lin, Fu-Hsiung; Truskey, George A.; Achneck, Hardean E.

    2011-01-01

    Implantable cardiovascular devices are manufactured from artificial materials (e.g. titanium (Ti), expanded polytetrafluoroethylene), which pose the risk of thromboemboli formation1,2,3. We have developed a method to line the inside surface of Ti tubes with autologous blood-derived human or porcine endothelial progenitor cells (EPCs)4. By implanting Ti tubes containing a confluent layer of porcine EPCs in the inferior vena cava (IVC) of pigs, we tested the improved biocompatibility of the cell-seeded surface in the prothrombotic environment of a large animal model and compared it to unmodified bare metal surfaces5,6,7 (Figure 1). This method can be used to endothelialize devices within minutes of implantation and test their antithrombotic function in vivo. Peripheral blood was obtained from 50 kg Yorkshire swine and its mononuclear cell fraction cultured to isolate EPCs4,8. Ti tubes (9.4 mm ID) were pre-cut into three 4.5 cm longitudinal sections and reassembled with heat-shrink tubing. A seeding device was built, which allows for slow rotation of the Ti tubes. We performed a laparotomy on the pigs and externalized the intestine and urinary bladder. Sharp and blunt dissection was used to skeletonize the IVC from its bifurcation distal to the right renal artery proximal. The Ti tubes were then filled with fluorescently-labeled autologous EPC suspension and rotated at 10 RPH x 30 min to achieve uniform cell-coating9. After administration of 100 USP/ kg heparin, both ends of the IVC and a lumbar vein were clamped. A 4 cm veinotomy was performed and the device inserted and filled with phosphate-buffered saline. As the veinotomy was closed with a 4-0 Prolene running suture, one clamp was removed to de-air the IVC. At the end of the procedure, the fascia was approximated with 0-PDS (polydioxanone suture), the subcutaneous space closed with 2-0 Vicryl and the skin stapled closed. After 3 - 21 days, pigs were euthanized, the device explanted en-block and fixed. The Ti

  4. Comparative Evaluation for Potential Differentiation of Endothelial Progenitor Cells and Mesenchymal Stem Cells into Endothelial-Like Cells

    PubMed Central

    Sabry, Dina; Noh, Olfat; Samir, Mai

    2016-01-01

    Understanding the mechanisms of vascular remodeling could lead to more effective treatments for ischemic conditions. We aimed to compare between the abilities of both human Wharton jelly derived mesenchymal stem cells (hMSCs) and human cord blood endothelial progenitor cells (hEPCs) and CD34+ to induce angiogenesis in vitro. hMSCs, hEPCs, and CD34+ were isolated from human umbilical cord blood using microbead (MiniMacs). The cells characterization was assessed by flow cytometry following culture and real-time PCR for vascular endothelial growth factor receptor 2 (VEGFR2) and von Willebrand factor (vWF) to prove stem cells differentiation. The study revealed successful isolation of hEPCs, CD34+, and hMSCs. The hMSCs were identified by gaining CD29+ and CD44+ using FACS analysis. The hEPCs were identified by having CD133+, CD34+, and KDR. The potential ability of hEPCs and CD34+ to differentiate into endothelial-like cells was more than hMSCs. This finding was assessed morphologically in culture and by higher significant VEGFR2 and vWF genes expression (p<0.05) in differentiated hEPCs and CD34+ compared to differentiated hMSCs. hEPCs and CD34+ differentiation into endothelial-like cells were much better than that of hMSCs. PMID:27426085

  5. Recent Progress in Endothelial Progenitor Cell Culture Systems: Potential for Stroke Therapy

    PubMed Central

    TAKIZAWA, Shunya; NAGATA, Eiichiro; NAKAYAMA, Taira; MASUDA, Haruchika; ASAHARA, Takayuki

    2016-01-01

    Endothelial progenitor cells (EPCs) participate in endothelial repair and angiogenesis due to their abilities to differentiate into endothelial cells and to secrete protective cytokines and growth factors. Consequently, there is considerable interest in cell therapy with EPCs isolated from peripheral blood to treat various ischemic injuries. Quality and quantity-controlled culture systems to obtain mononuclear cells enriched in EPCs with well-defined angiogenic and anti-inflammatory phenotypes have recently been developed, and increasing evidence from animal models and clinical trials supports the idea that transplantation of EPCs contributes to the regenerative process in ischemic organs and is effective for the therapy of ischemic cerebral injury. Here, we briefly describe the general characteristics of EPCs, and we review recent developments in culture systems and applications of EPCs and EPC-enriched cell populations to treat ischemic stroke. PMID:27041632

  6. Vascular endothelial growth factor-A signaling in bone marrow-derived endothelial progenitor cells exposed to hypoxic stress.

    PubMed

    Hoffmann, Brian R; Wagner, Jordan R; Prisco, Anthony R; Janiak, Agnieszka; Greene, Andrew S

    2013-11-01

    Bone marrow-derived endothelial progenitor cells (BM-EPCs) are stimulated by vascular endothelial growth factor-A (VEGF-A) and other potent proangiogenic factors. During angiogenesis, an increase in VEGF-A expression stimulates BM-EPCs to enhance endothelial tube formation and contribute to an increase in microvessel density. Hypoxia is known to produce an enhanced angiogenic response and heightened levels of VEGF-A have been seen in oxygen deprived epithelial and endothelial cells, yet the pathways for VEGF-A signaling in BM-EPCs have not been described. This study explores the influence of hypoxia on VEGF-A signaling in rat BM-EPCs utilizing a novel proteomic strategy to directly identify interacting downstream components of the combined VEGF receptor(s) signaling pathways, gene expression analysis, and functional phenotyping. VEGF-A signaling network analysis following liquid chromatographic separation and tandem mass spectrometry revealed proteins related to inositol/calcium signaling, nitric oxide signaling, cell survival, cell migration, and inflammatory responses. Alterations in BM-EPC expression of common angiogenic genes and tube formation in response to VEGF-A during hypoxia were measured and combined with the proteomic analysis to enhance and support the signaling pathways detected. BM-EPC tube formation assays in response to VEGF-A exhibited little tube formation; however, a cell projection/migratory phenotype supported the signaling data. Additionally, a novel assay measuring BM-EPC incorporation into preformed endothelial cell tubes indicated a significant increase of incorporated BM-EPCs after pretreatment with VEGF-A during hypoxia. This study verifies known VEGF-A pathway components and reveals several unidentified mechanisms of VEGF-A signaling in BM-EPCs during hypoxia that may be important for migration to sites of vascular regeneration. PMID:24022223

  7. Vascular endothelial growth factor-A signaling in bone marrow-derived endothelial progenitor cells exposed to hypoxic stress

    PubMed Central

    Hoffmann, Brian R.; Wagner, Jordan R.; Prisco, Anthony R.; Janiak, Agnieszka

    2013-01-01

    Bone marrow-derived endothelial progenitor cells (BM-EPCs) are stimulated by vascular endothelial growth factor-A (VEGF-A) and other potent proangiogenic factors. During angiogenesis, an increase in VEGF-A expression stimulates BM-EPCs to enhance endothelial tube formation and contribute to an increase in microvessel density. Hypoxia is known to produce an enhanced angiogenic response and heightened levels of VEGF-A have been seen in oxygen deprived epithelial and endothelial cells, yet the pathways for VEGF-A signaling in BM-EPCs have not been described. This study explores the influence of hypoxia on VEGF-A signaling in rat BM-EPCs utilizing a novel proteomic strategy to directly identify interacting downstream components of the combined VEGF receptor(s) signaling pathways, gene expression analysis, and functional phenotyping. VEGF-A signaling network analysis following liquid chromatographic separation and tandem mass spectrometry revealed proteins related to inositol/calcium signaling, nitric oxide signaling, cell survival, cell migration, and inflammatory responses. Alterations in BM-EPC expression of common angiogenic genes and tube formation in response to VEGF-A during hypoxia were measured and combined with the proteomic analysis to enhance and support the signaling pathways detected. BM-EPC tube formation assays in response to VEGF-A exhibited little tube formation; however, a cell projection/migratory phenotype supported the signaling data. Additionally, a novel assay measuring BM-EPC incorporation into preformed endothelial cell tubes indicated a significant increase of incorporated BM-EPCs after pretreatment with VEGF-A during hypoxia. This study verifies known VEGF-A pathway components and reveals several unidentified mechanisms of VEGF-A signaling in BM-EPCs during hypoxia that may be important for migration to sites of vascular regeneration. PMID:24022223

  8. Donor origin of circulating endothelial progenitors after allogeneic bone marrow transplantation.

    PubMed

    Ikpeazu, C; Davidson, M K; Halteman, D; Browning, P J; Brandt, S J

    2000-01-01

    Endothelial cell precursors circulate in blood and express antigens found on hematopoietic stem cells, suggesting that such precursors might be subject to transplantation. To investigate, we obtained adherence-depleted peripheral blood mononuclear cells from 3 individuals who had received a sex-mismatched allogeneic bone marrow transplant (BMT) and cultured the cells on fibronectin-coated plates with endothelial growth factors. The phenotype of the spindle-shaped cells that emerged in culture was characterized by immunofluorescent staining, and the origin of the cells was determined using a polymerase chain reaction (PCR)-based assay for polymorphic short tandem repeats (STRs). The cells manifested a number of endothelial characteristics-such as von Wlllebrand factor, CD31, and Flk-1/KDR expression; Bandeiraea simplicifolia lectin 1 binding; and acetylated low-density lipoprotein uptake-but lacked expression of certain markers of activation or differentiation, including intercellular adhesion molecule-1, vascular cell adhesion molecule-1, and the epitope for the anti-endothelial cell antibody P1H12. For each patient and at all time points studied (ranging from 5 to 52 months after transplantation), STR-PCR analysis showed that cultured cells and nucleated blood cells came exclusively from the bone marrow donor. These results demonstrate that circulating endothelial progenitors are both transplantable and capable of long-term repopulation of human allogeneic BMT recipients. PMID:10905767

  9. Sonic hedgehog derived from human pancreatic cancer cells augments angiogenic function of endothelial progenitor cells.

    PubMed

    Yamazaki, Madoka; Nakamura, Kazumasa; Mizukami, Yusuke; Ii, Masaaki; Sasajima, Junpei; Sugiyama, Yoshiaki; Nishikawa, Tomoya; Nakano, Yasuhiro; Yanagawa, Nobuyuki; Sato, Kazuya; Maemoto, Atsuo; Tanno, Satoshi; Okumura, Toshikatsu; Karasaki, Hidenori; Kono, Toru; Fujiya, Mikihiro; Ashida, Toshifumi; Chung, Daniel C; Kohgo, Yutaka

    2008-06-01

    Hedgehog signaling is important in the pathogenesis of pancreatic cancer. Several recent observations suggest the involvement of sonic hedgehog (SHH) in postnatal neovascularization. We identified a novel role for SHH in tumor-associated angiogenesis in pancreatic cancer. Immunohistochemical analysis revealed that patched homolog 1 (PTCH1), both a receptor for and transcriptional target of hedgehog signaling, was expressed in a small fraction of endothelial cells within pancreatic cancer, but not in normal pancreatic tissue. When endothelial progenitor cells (EPC) isolated from human peripheral blood were cultured with supernatant from SHH-transfected 293 cells or pancreatic cancer cells, mRNA levels of vascular endothelial growth factor (VEGF), stromal cell-derived factor-1 and angiopoietin-1 were significantly increased, whereas no such induction was observed in human umbilical vein endothelial cell (HUVEC) and human dermal microvascular endothelial cell (HMVEC). HUVEC tube formation was stimulated when cocultured with EPC, and preconditioning EPC with supernatant from KP-1 N pancreatic cancer cells highly expressing SHH significantly enhanced the effect. The effect was partially attenuated by specific inhibition of SHH with cyclopamine or a neutralizing antibody. These findings suggest that tumor-derived SHH can induce angiogenesis, and this is mediated by its effects on EPC specifically. Targeting SHH would be a novel therapeutic approach that can inhibit not only proliferation of cancer cells but also EPC-mediated angiogenesis. PMID:18422746

  10. Lineage tracking of mesenchymal and endothelial progenitors in BMP-induced bone formation.

    PubMed

    Kolind, Mille; Bobyn, Justin D; Matthews, Brya G; Mikulec, Kathy; Aiken, Alastair; Little, David G; Kalajzic, Ivo; Schindeler, Aaron

    2015-12-01

    To better understand the relative contributions of mesenchymal and endothelial progenitor cells to rhBMP-2 induced bone formation, we examined the distribution of lineage-labeled cells in Tie2-Cre:Ai9 and αSMA-creERT2:Col2.3-GFP:Ai9 reporter mice. Established orthopedic models of ectopic bone formation in the hind limb and spine fusion were employed. Tie2-lineage cells were found extensively in the ectopic bone and spine fusion masses, but co-staining was only seen with tartrate-resistant acid phosphatase (TRAP) activity (osteoclasts) and CD31 immunohistochemistry (vascular endothelial cells), and not alkaline phosphatase (AP) activity (osteoblasts). To further confirm the lack of a functional contribution of Tie2-lineage cells to BMP-induced bone, we developed conditional knockout mice where Tie2-lineage cells are rendered null for key bone transcription factor osterix (Tie2-cre:Osx(fx/fx) mice). Conditional knockout mice showed no difference in BMP-induced bone formation compared to littermate controls. Pulse labeling of mesenchymal cells with Tamoxifen in mice undergoing spine fusion revealed that αSMA-lineage cells contributed to the osteoblastic lineage (Col2.3-GFP), but not to endothelial cells or osteoclast populations. These data indicate that the αSMA+ and Tie2+ progenitor lineages make distinct cellular contributions to bone formation, angiogenesis, and resorption/remodeling. PMID:26141839

  11. The Effects of Inhaled Nickel Nanoparticles on Murine Endothelial Progenitor Cells

    NASA Astrophysics Data System (ADS)

    Liberda, Eric N.

    Introduction. Particulate air pollution, specifically nickel found on or in particulate matter, has been associated with an increased risk of mortality in human population studies and can cause increases in vascular inflammation, generate reactive oxygen species, alter vasomotor tone, and potentiate atherosclerosis in murine exposures. With the discovery of endothelial progenitor cells (EPCs), a door has been opened which may explain these observed cardiovascular effects associated with inhaled air particles and nickel exposure. In order to further quantify the effects of inhaled nickel nanoparticles and attempt to elucidate how the observed findings from other studies may occur, several whole body inhalation exposure experiments to nickel nanoparticles were performed. Methods. Following whole body exposure to approximately 500mug/m3 of nickel nanoparticles for 5 hrs, bone marrow EPCs from C57BL/6 mice were isolated. EPCs were harvested for their RNA or used in a variety of assays including chemotaxis, tube formation, and proliferation. Gene expression was assessed for important receptors involved in EPC mobilization and homing using RT-PCR methods. EPCs, circulating endothelial progenitor cells, circulating endothelial cells (CECs), and endothelial microparticles (EMPs) were quantified on a BD FACSCalibur to examine endothelial damage and repair associated with the inhalation exposure. Plasma proteins were assessed using the 2D DIGE proteomic approach and commercially available ELISAs. Results and Conclusions. Exposure to inhaled nickel nanoparticles significantly increased both bone marrow EPCs as well as their levels in circulation. CECs were significantly upregulated suggesting that endothelial damage occurred due to the exposure. There was no significant difference in EMPs between the two groups. Tube formation and chemotaxis, but not proliferation, of bone marrow EPCs was impaired in the nickel nanoparticle exposed group. This decrease in EPC function

  12. Role of endothelial nitric oxide in bone marrow-derived progenitor cell mobilization.

    PubMed

    de Resende, M Monterio; Huw, L-Y; Qian, H-S; Kauser, K

    2007-01-01

    Mobilization and recruitment of bone marrow-derived progenitor cells (BMDPCs) play an important role in postischemic tissue repair. Patients with coronary artery disease (CAD) or peripheral vascular disease (PVD) exhibit endothelial dysfunction, and as a result are likely to have a reduced number of progenitor cells mobilized in their peripheral circulation following ischemic injury. Identification of eNOS independent pathways for BMDPC mobilization may have important therapeutic value in this patient population. To identify such mechanisms we investigated the effect of granulocyte-colony stimulating factor (GCSF) and stem cell factor (SCF) in eNOS-KO mice with and without surgical hind-limb ischemia. Our results suggest that BMDPC mobilization can be achieved via activation of NO-independent pathways. PMID:17554503

  13. Resident Endothelial Progenitor Cells From Human Placenta Have Greater Vasculogenic Potential Than Circulating Endothelial Progenitor Cells From Umbilical Cord Blood

    PubMed Central

    Rapp, Brian M.; Saadatzedeh, M. Reza; Ofstein, Richard H.; Bhavsar, Janak R.; Tempel, Zachary S.; Moreno, Oscar; Morone, Peter; Booth, Dana A.; Traktuev, Dmitry O.; Dalsing, Michael C.; Ingram, David A.; Yoder, Mervin C.; March, Keith L.; Murphy, Michael P.

    2012-01-01

    Endothelial colony-forming cells (ECFCs) isolated from umbilical cord blood (CBECFCs) are highly proliferative and form blood vessels in vivo. The purpose of this investigation was to isolate and characterize a population of resident ECFCs from the chorionic villi of term human placenta and provide a comparative analysis of their proliferative and vasculogenic potential with CBECFCs. ECFCs were isolated from umbilical cord blood and chorionic villi from placentas obtained by caesarean deliveries. Placental ECFCs (PECFCs) expressed CD144, CD31, CD105, and KDR and were negative for CD45 and CD34, consistent with other ECFC phenotypes. PECFCs were capable of 28.6 ± 6.0 population doublings before reaching senescence (vs. 47.4 ± 3.2 for CBECFCs, p < 0.05, n = 4). In single cell assays, 46.5 ± 1.2% underwent at least one division (vs. 51.0 ± 1.8% of CBECFCs, p = 0.07, n = 6), and of those dividing PECFCs, 71.8 ± 0.9% gave rise to colonies of >500 cells (highly proliferative potential clones) over 14 days (vs. 69.4 ± 0.7% of CBECFCs, p = 0.07, n = 9). PECFCs formed 5.2 ± 0.8 vessels/mm2 in collagen/fibronectin plugs implanted into non-obese diabetic/severe combined immunodeficient (NOD/SCID) mice, whereas CBECFCs formed only 1.7 ± 1.0 vessels/mm2 (p < 0.05, n = 4). This study demonstrates that circulating CBECFCs and resident PECFCs are identical phenotypically and contain equivalent quantities of high proliferative potential clones. However, PECFCs formed significantly more blood vessels in vivo than CBECFCs, indicating that differences in vasculogenic potential between circulating and resident ECFCs exist. PMID:27004134

  14. Functional characterization of late outgrowth endothelial progenitor cells in patients with end-stage renal failure

    PubMed Central

    Zhao, Jing; Bolton, Eleanor M; Randle, Lucy; Bradley, John Andrew; Lever, Andrew M L

    2014-01-01

    Renal transplantation is potentially curative in renal failure, but long-term efficacy is limited by untreatable chronic rejection. Endothelial damage contributes to chronic rejection and is potentially repairable by circulating endothelial progenitor cells (EPC). The frequency and function of EPC are variably influenced by end-stage renal failure (ESRF). Here, we isolated and functionally characterized the late outgrowth EPC (LO-EPC) from ESRF patients to investigate their potential for endothelial repair. Patients with ESRF generated more LO-EPC colonies than healthy controls and had higher plasma levels of IL-1rα, IL-16, IL-6, MIF, VEGF, Prolactin, and PLGF. Patients' LO-EPC displayed normal endothelial cell morphology, increased secretion of PLGF, MCP-1, and IL-1β, and normal network formation in vitro and in vivo. They demonstrated decreased adhesion to extracellular matrix. Integrin gene profiles and protein expression were comparable in patients and healthy volunteers. In some patients, mesenchymal stem cells (MSC) were co-isolated and could be differentiated into adipocytes and osteocytes in vitro. This is the first study to characterize LO-EPC from patients with ESRF. Their behavior in vitro reflects the presence of elevated trophic factors; their ability to proliferate in vitro and angiogenic function makes them candidates for prevention of chronic rejection. Their impaired adhesion and the presence of MSC are areas for potential therapeutic intervention. PMID:24471420

  15. Protective effects of tanshinone IIA on endothelial progenitor cells injured by tumor necrosis factor-α

    PubMed Central

    WANG, XING-XIANG; YANG, JIN-XIU; PAN, YAN-YUN; ZHANG, YE-FEI

    2015-01-01

    Tanshinone IIA (Tan IIA) is a Traditional Chinese Medicine commonly used in Asian and Western countries for the prevention and treatment of cardiovascular disorders, such as atherosclerosis. Endothelial dysfunction and associated inflammatory processes have a critical role in the development of atherosclerosis. Endothelial progenitor cells (EPCs) have been demonstrated to be involved in certain aspects of the endothelial repair process. The present study aimed to investigate the putative protective effects of Tan IIA on EPCs injured by tumor necrosis factor-α (TNF-α). The potential effects of Tan IIA on TNF-α-stimulated EPC proliferation, migration, adhesion, in vitro tube formation ability and paracrine activity were investigated in the current study. The results indicated that TNF-α impaired EPC proliferation, migration, adhesion capacity and vasculogenesis ability in vitro as well as promoted EPC secretion of inflammatory cytokines, including monocyte chemoattractant protein-1 (MCP-1), interleukin-6 (IL-6) and soluble CD40 ligand (sCD40L). However, Tan IIA was able to reverse these effects. In conclusion, these findings demonstrated that Tan IIA may have the potential to protect EPCs against damage induced by TNF-α. Therefore, these results may provide evidence for the pharmacological basis of Tan IIA and its potential use in the prevention and treatment of early atherosclerosis associated with EPC and endothelial damage. PMID:26095681

  16. TNFα-Damaged-HUVECs Microparticles Modify Endothelial Progenitor Cell Functional Activity

    PubMed Central

    Luna, Carlos; Carmona, Andrés; Alique, Matilde; Carracedo, Julia; Ramirez, Rafael

    2015-01-01

    Endothelial progenitor cells (EPCs) have an important role in the maintenance of vascular integrity and homeostasis. While there are many studies that explain EPCs mechanisms action, there are few studies that demonstrate how they interact with other emerging physiological elements such as Endothelial Microparticles (EMPs). EMPs are membranous structures with a size between 100 and 1000 nm that act as molecular information transporter in biological systems and are known as an important elements in develop different pathologies; moreover a lot of works explains that are novel biomarkers. To elucidate these interactions, we proposed an in vitro model of endothelial damage mediated by TNFalpha, in which damaged EMPs and EPCs are in contact to assess EPCs functional effects. We have observed that damaged EMPs can modulate several EPCs classic factors as colony forming units (CFUs), contribution to repair a physically damaged endothelium (wound healing), binding to mature endothelium, and co-adjuvants to the formation of new vessels in vitro (angiogenesis). All of these in a dose-dependent manner. Damaged EMPs at a concentration of 103 MPs/ml have an activating effect of these capabilities, while at concentrations of 105 MPs/ml these effects are attenuated or reduced. This in vitro model helps explain that in diseases where there is an imbalance between these two elements (EPCs and damaged EMPs), the key cellular elements in the regeneration and maintenance of vascular homeostasis (EPCs) are not fully functional, and could explain, at least in part, endothelial dysfunction associated in various pathologies. PMID:26733886

  17. Disrupted Endothelial Cell Layer and Exposed Extracellular Matrix Proteins Promote Capture of Late Outgrowth Endothelial Progenitor Cells

    PubMed Central

    Mitrofan, Claudia-Gabriela; Appleby, Sarah L.; Morrell, Nicholas W.; Lever, Andrew M. L.

    2016-01-01

    Late outgrowth endothelial progenitor cells (LO-EPC) possess a high proliferative potential, differentiate into vascular endothelial cells (EC), and form networks, suggesting they play a role in vascular repair. However, due to their scarcity in the circulation there is a requirement for ex vivo expansion before they could provide a practical cell therapy and it is currently unclear if they would home and engraft to an injury site. Using an in vitro flow system we studied LO-EPC under simulated injury conditions including EC activation, ischaemia, disrupted EC integrity, and exposed basement membrane. Perfused LO-EPC adhered to discontinuous EC paracellularly at junctional regions between adjacent cells under shear stress 0.7 dyn/cm2. The interaction was not adhesion molecule-dependent and not enhanced by EC activation. LO-EPC expressed high levels of the VE-Cadherin which may explain these findings. Ischaemia reperfusion injury decreased the interaction with LO-EPC due to cell retraction. LO-EPC interacted with exposed extracellular matrix (ECM) proteins, fibronectin and vitronectin. The interaction was mediated by integrins α5β3, αvβ1, and αvβ3. This study has demonstrated that an injured local environment presents sufficient adhesive signals to capture flow perfused LO-EPC in vitro and that LO-EPC have properties consistent with their potential role in vascular repair. PMID:27413378

  18. Disrupted Endothelial Cell Layer and Exposed Extracellular Matrix Proteins Promote Capture of Late Outgrowth Endothelial Progenitor Cells.

    PubMed

    Zhao, Jing; Mitrofan, Claudia-Gabriela; Appleby, Sarah L; Morrell, Nicholas W; Lever, Andrew M L

    2016-01-01

    Late outgrowth endothelial progenitor cells (LO-EPC) possess a high proliferative potential, differentiate into vascular endothelial cells (EC), and form networks, suggesting they play a role in vascular repair. However, due to their scarcity in the circulation there is a requirement for ex vivo expansion before they could provide a practical cell therapy and it is currently unclear if they would home and engraft to an injury site. Using an in vitro flow system we studied LO-EPC under simulated injury conditions including EC activation, ischaemia, disrupted EC integrity, and exposed basement membrane. Perfused LO-EPC adhered to discontinuous EC paracellularly at junctional regions between adjacent cells under shear stress 0.7 dyn/cm(2). The interaction was not adhesion molecule-dependent and not enhanced by EC activation. LO-EPC expressed high levels of the VE-Cadherin which may explain these findings. Ischaemia reperfusion injury decreased the interaction with LO-EPC due to cell retraction. LO-EPC interacted with exposed extracellular matrix (ECM) proteins, fibronectin and vitronectin. The interaction was mediated by integrins α5β3, αvβ1, and αvβ3. This study has demonstrated that an injured local environment presents sufficient adhesive signals to capture flow perfused LO-EPC in vitro and that LO-EPC have properties consistent with their potential role in vascular repair. PMID:27413378

  19. Endothelial Progenitor Cells Combined with Cytosine Deaminase-Endostatin for Suppression of Liver Carcinoma.

    PubMed

    Chen, Rong; Yu, Hui; An, Yan-Li; Chen, Hua-Jun; Jia, ZhenYu; Teng, Gao-Jun

    2016-06-01

    Transplantation of gene transfected endothelial progenitor cells (EPCs) provides a novel method for treatment of human tumors. To study treatment of hepatocellular carcinoma using cytosine deaminase (CD)- and endostatin (ES)-transfected endothelial progenitor cells (EPCs), mouse bone marrow-derived EPCs were cultured and transfected with Lenti6.3-CD-EGFP and Lenti6.3-ES-Monomer-DsRed labeled with superparamagnetic iron oxide (SPIO) nanoparticles. DiD (lipophilic fluorescent dye)-labeled EPCs were injected into normal mice and mice with liver carcinoma. The EPCs loaded with CD-ES were infused into the mice through caudal veins and tumor volumes were measured. The tumor volumes in the EPC + SPIO + CD/5-Fc + ES group were found to be smaller as a result and grew more slowly than those from the EPC + SPIO + LV (lentivirus, empty vector control) group. Survival times were also measured after infusion of the cells into the mice. The median survival time was found to be longer in the EPC + SPIO + CD/5-Fc + ES group than in the others. In conclusion, the EPCs transfected with CD-ES suppressed the liver carcinoma cells in vitro, migrated primarily to the carcinoma, inhibited tumor growth, and also extended the median survival time for the mice with liver carcinoma. PMID:27319212

  20. Cathepsin L is required for endothelial progenitor cell-induced neovascularization

    SciTech Connect

    Urbich, Carmen; Heeschen, Christopher; Aicher, Alexandra; Sasaki, Ken-ichiro; Bruhl, Thomas; Hofmann, Wolf K.; Peters, Christoph; Reinheckel, Thomas; Pennacchio, Len A.; Abolmaali, Nasreddin D.; Chavakis, Emmanouil; Zeiher, Andreas M.; Dimmeler, Stefanie

    2004-01-15

    Infusion of endothelial progenitor cells (EPCs), but not of mature endothelial cells (ECs), promotes neovascularization after ischemia. We performed a gene expression profiling of EPCs and ECs to identify genes, which might be important for the neovascularization capacity of EPCs. Intriguingly, the protease cathepsin L (CathL) was highly expressed in EPCs as opposed to ECs and is essential for matrix degradation and invasion by EPCs in vitro. CathL deficient mice showed impaired functional recovery after hind limb ischemia supporting the concept for an important role of CathL in postnatal neovascularization. Infused CathL deficient progenitor cells failed to home to sites of ischemia and to augment neovascularization. In contrast, over expression of CathL in mature ECs significantly enhanced their invasive activity and induced their neovascularization capacity in vivo. Taken together, CathL plays a crucial role for the integration of circulating EPCs into the ischemic tissue and is required for neovascularization mediated by EPCs.

  1. Endothelial Progenitor Cells Predict Long-Term Mortality in Hemodialysis Patients

    PubMed Central

    Lu, Chien-Lin; Leu, Jyh-Gang; Liu, Wen-Chih; Zheng, Cai-Mei; Lin, Yuh-Feng; Shyu, Jia-Fwu; Wu, Chia-Chao; Lu, Kuo-Cheng

    2016-01-01

    Background: The endothelial progenitor cells (EPCs) dysfunction is a critical event in the initiation of atherosclerotic plaque development and the level of circulating EPCs can be considered a biomarker of cardiovascular events. The level and functional change in EPCs has been investigated in hemodialysis patients, but the effect of absolute number of EPCs on risk of death has not yet been explored. We hypothesized that the number of EPCs predicted death from cardiovascular and all-cause mortality in hemodialysis patients. Methods: We evaluate the association between endothelial progenitor cells and clinical outcome in 154 patients on maintenance hemodialysis. The blood sample was drawn at the time of patient enrollment and EPCs were identified by flow cytometry using triple staining for CD34/CD133/KDR. Results: The median duration of follow-up was 4.19 years. There were 79 (51.3%) deaths during the follow-up period, 41 of whom died due to a confirmed cardiovascular cause. The cumulative survival was greater in the high-EPC group than the low-EPC group for all-cause and cardiovascular mortality. Decreased EPCs levels were associated with a significant increase in the risk of cardiovascular and all-cause mortality after adjusting for age, gender, current smokers, diabetes mellitus, and hypertension. Conclusions: The level of circulating EPCs independently predicts the clinical outcome in patients on maintenance hemodialysis. Thus, the EPCs levels may be a useful predictive tool for evaluating the risk of death in maintenance hemodialysis patients. PMID:26941585

  2. 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

  3. Systemic influences contribute to prolonged microvascular rarefaction after brain irradiation: a role for endothelial progenitor cells

    PubMed Central

    Ashpole, Nicole M.; Warrington, Junie P.; Mitschelen, Matthew C.; Yan, Han; Sosnowska, Danuta; Gautam, Tripti; Farley, Julie A.; Csiszar, Anna; Ungvari, Zoltan

    2014-01-01

    Whole brain radiation therapy (WBRT) induces profound cerebral microvascular rarefaction throughout the hippocampus. Despite the vascular loss and localized cerebral hypoxia, angiogenesis fails to occur, which subsequently induces long-term deficits in learning and memory. The mechanisms underlying the absence of vessel recovery after WBRT are unknown. We tested the hypotheses that vascular recovery fails to occur under control conditions as a result of loss of angiogenic drive in the circulation, chronic tissue inflammation, and/or impaired endothelial cell production/recruitment. We also tested whether systemic hypoxia, which is known to promote vascular recovery, reverses these chronic changes in inflammation and endothelial cell production/recruitment. Ten-week-old C57BL/6 mice were subjected to a clinical series of fractionated WBRT: 4.5-Gy fractions 2 times/wk for 4 wk. Plasma from radiated mice increased in vitro endothelial cell proliferation and adhesion compared with plasma from control mice, indicating that WBRT did not suppress the proangiogenic drive. Analysis of cytokine levels within the hippocampus revealed that IL-10 and IL-12(p40) were significantly increased 1 mo after WBRT; however, systemic hypoxia did not reduce these inflammatory markers. Enumeration of endothelial progenitor cells (EPCs) in the bone marrow and circulation indicated that WBRT reduced EPC production, which was restored with systemic hypoxia. Furthermore, using a bone marrow transplantation model, we determined that bone marrow-derived endothelial-like cells home to the hippocampus after systemic hypoxia. Thus, the loss of production and homing of EPCs have an important role in the prolonged vascular rarefaction after WBRT. PMID:25038144

  4. The angiogenic gene profile of circulating endothelial progenitor cells from ischemic stroke patients

    PubMed Central

    2013-01-01

    Background The identification of circulating endothelial progenitor cells (EPCs) has introduced new possibilities for cell-based treatments for stroke. We tested the angiogenic gene expression of outgrowth endothelial cells (OECs), an EPC subtype capable to shape vessel structures. Methods OECs (at colony or mature stages) from ischemic stroke patients (n=8) were characterized using the RT2 ProfilerTM human angiogenesis PCR Array, and human microvascular endothelial cells (hCMEC/D3) were used as an expression reference of endothelial cells. Results Colony-OECs showed higher expression of CCL2, ID3, IGF-1, MMP9, TGFBR1, TNFAIP2, TNF and TGFB1. However, BAI-1, NRP2, THBS1, MMP2 and VEGFC expression was increased in mature-OECs (p<0.05). ID3 (p=0.008) and TGFBR1 (p=0.03) genes remained significantly overexpressed in colony-OECs compared to mature-OECs or hCMEC/D3. MMP9 levels were significantly increased in colony-OECs (p=0.025) compared to mature-OECs. Moreover, MMP-2, VEGF-C, THBS1 and NRP-2 gene expression was also significantly increased in mature-OECs compared to hCMEC/D3 (p<0.05). Some of these genes were positively validated by RT-PCR. Conclusion Our study shows that OECs from stroke patients present higher levels of pro-angiogenic factors at early stages, decreasing in mature OECs when they become more similar to mature microvascular endothelial cells. PMID:23388410

  5. [Roles of aberrant endothelial progenitor cells in pathogenesis of systemic sclerosis].

    PubMed

    Kuwana, Masataka

    2013-01-01

    Systemic sclerosis (SSc) is a multisystem connective tissue disease characterized by excessive fibrosis and microvasculopathy, along with poor vascular formation and repair. The maintenance of the postnatal vascular system requires constant remodeling through vasculogenesis, which is mediated by the de novo differentiation of mature endothelial cells from endothelial progenitor cells (EPCs). However, a great deal of controversy about EPCs and their roles in postnatal vascular formation has arisen because of discrepancies in how EPCs are defined. The current consensus is that EPCs are heterogeneous cell population containing an extremely small count of "true EPCs", and pro-angiogenic hematopoietic cells (PHCs) that promotes vascular formation and repair through secretion of pro-angiogenic factors, and differentiation into endothelial cells and mural cells. In 2004, we reported a reduced number and impaired function of circulating CD34(+)CD133(+)CD309(+)CD45(dim)CD14(-) EPCs, which are now regarded as an immature subset of PHCs, in patients with SSc, and proposed a theory that defective vascular repair machinery as one of important mechanisms contributing to SSc vasculopathy. In addition, we showed that in SSc patients, circulating monocytic PHCs were increased and have enhanced angiogenic potency and differentiation potential to fibroblast-like cells. In summary, EPCs are involved in the pathogenesis of SSc by participating in two major pathological features, microvasculopathy and excessive fibrosis. Understanding the roles of EPCs in disease process of SSc may be key to dissecting its pathogenesis and to developing novel therapeutic strategies for this intractable condition. PMID:23445728

  6. Surface-modified hyaluronic acid hydrogels to capture endothelial progenitor cells.

    PubMed

    Camci-Unal, Gulden; Aubin, Hug; Ahari, Amirhossein Farajzadeh; Bae, Hojae; Nichol, Jason William; Khademhosseini, Ali

    2010-10-21

    A major challenge to the effective treatment of injured cardiovascular tissues is the promotion of endothelialization of damaged tissues and implanted devices. For this reason, there is a need for new biomaterials that promote endothelialization to enhance vascular repair. The goal of this work was to develop antibody-modified polysaccharide-based hydrogels that could selectively capture endothelial progenitor cells (EPCs). We showed that CD34 antibody immobilization on hyaluronic acid (HA) hydrogels provides a suitable surface to capture EPCs. The effect of CD34 antibody immobilization on EPC adhesion was found to be dependent on antibody concentration. The highest level of EPC attachment was found to be 52.2 cells per mm(2) on 1% HA gels modified with 25 μg mL(-1) antibody concentration. Macrophages did not exhibit significant attachment on these modified hydrogel surfaces compared to the EPCs, demonstrating the selectivity of the system. Hydrogels containing only HA, with or without immobilized CD34, did not allow for spreading of EPCs 48 h after cell seeding, even though the cells were adhered to the hydrogel surface. To promote spreading of EPCs, 2% (w/v) gelatin methacrylate (GelMA) containing HA hydrogels were synthesized and shown to improve cell spreading and elongation. This strategy could potentially be useful to enhance the biocompatibility of implants such as artificial heart valves or in other tissue engineering applications where formation of vascular structures is required. PMID:22368689

  7. Hypoxia-Induced Endothelial Progenitor Cell Function Is Blunted in Angiotensinogen Knockout Mice

    PubMed Central

    Choi, Jin-Hwa; Nguyen, Minh-Phuong; Lee, Dongjin; Oh, Goo-Taeg; Lee, You-Mie

    2014-01-01

    Angiotensinogen (AGT), the precursor of angiotensin I, is known to be involved in tumor angiogenesis and associated with the pathogenesis of coronary atherosclerosis. This study was undertaken to determine the role played by AGT in endothelial progenitor cells (EPCs) in tumor progression and metastasis. It was found that the number of EPC colonies formed by AGT heterozygous knockout (AGT+/−) cells was less than that formed by wild-type (WT) cells, and that the migration and tube formation abilities of AGT+/− EPCs were significantly lower than those of WT EPCs. In addition, the gene expressions of vascular endothelial growth factor (VEGF), Flk1, angiopoietin (Ang)-1, Ang-2, Tie-2, stromal derived factor (SDF)-1, C-X-C chemokine receptor type 4 (CXCR4), and of endothelial nitric oxide synthase (eNOS) were suppressed in AGT+/− EPCs. Furthermore, the expressions of hypoxia-inducible factor (HIF)-1α and -2α were downregulated in AGT+/− early EPCs under hypoxic conditions, suggesting a blunting of response to hypoxia. Moreover, the activation of Akt/eNOS signaling pathways induced by VEGF, epithelial growth factor (EGF), or SDF-1α were suppressed in AGT+/− EPCs. In AGT+/− mice, the incorporation of EPCs into the tumor vasculature was significantly reduced, and lung tumor growth and melanoma metastasis were attenuated. In conclusion, AGT is required for hypoxia-induced vasculogenesis. PMID:24938229

  8. Changes in Caspase-3, B Cell Leukemia/Lymphoma-2, Interleukin-6, Tumor Necrosis Factor-α and Vascular Endothelial Growth Factor Gene Expression after Human Umbilical Cord Blood Derived Mesenchymal Stem Cells Transfusion in Pulmonary Hypertension Rat Models

    PubMed Central

    Kim, Kwan Chang; Lee, Jae Chul; Lee, Hyeryon; Cho, Min-Sun; Choi, Soo Jin

    2016-01-01

    Background and Objectives Failure of vascular smooth muscle apoptosis and inflammatory response in pulmonary arterial hypertension (PAH) is a current research focus. The goals of this study were to determine changes in select gene expressions in monocrotaline (MCT)-induced PAH rat models after human umbilical cord blood derived mesenchymal stem cells (hUCB-MSCs) transfusion. Materials and Methods The rats were separated into 3 groups i.e., control group (C group), M group (MCT 60 mg/kg), and U group (hUCB-MSCs transfusion) a week after MCT injection. Results TUNEL assay showed that the U group had significantly lowered positive apoptotic cells in the lung tissues, as compared with the M group. mRNA of caspase-3, B cell leukemia/lymphoma (Bcl)-2, interleukin (IL)-6, tumor necrosis factor (TNF)-α and vascular endothelial growth factor (VEGF) in the lung tissues were greatly reduced at week 4 in the U group. Immunohistochemical staining of the lung tissues also demonstrated a similar pattern, with the exception of IL-6. The protein expression of caspase-3, Bcl-2 VEGF, IL-6, TNF-α and brain natriuretic peptide in the heart tissues were significantly lower in the U group, as compared with the M group at week 2. Furthermore, the protein expression of VEGF, IL-6 and BNP in the heart tissues were significantly lower in the U group at week 4. Collagen content in the heart tissues was significantly lower in the U group, as compared with M group at weeks 2 and 4, respectively. Conclusion hUCB-MSCs could prevent inflammation, apoptosis and remodeling in MCT-induced PAH rat models. PMID:26798389

  9. Circulating Endothelial Progenitor Cells and Clinical Outcome in Patients with Aortic Stenosis

    PubMed Central

    Shimoni, Sara; Bar, Iris; Meledin, Valery; Derazne, Estela; Gandelman, Gera; George, Jacob

    2016-01-01

    Background Aortic stenosis (AS) is the most common valvular disease. Endothelial progenitor cells (EPCs) have a role in the repair of endothelial surfaces after injury. Reduced numbers of EPCs are associated with endothelial dysfunction and adverse clinical events, suggesting that endothelial injury in the absence of sufficient repair by circulating EPCs promotes the progression of vascular and possibly valvular disorders. The aim of this study was to assess EPC number in patients with AS and to study the predictive value of their circulating levels on prognosis. Methods The number of EPCs was determined by flow cytometry in 241 patients with AS and a control group of 73 pts. Thirty-eight, 52 and 151 patients had mild, moderate and severe AS, respectively. We evaluated the association between baseline levels of EPCs and death from cardiovascular causes during follow up. Results EPC level was significantly higher in patients with AS compared to the control group (p = 0.017). Two hundred and three patients with moderate and severe AS were followed for a median of 20 months. One hundred and twenty patients underwent an intervention. Thirty four patients died during follow up, 20 patients died due to cardiac causes. Advanced age, the presence of coronary artery disease, AS severity index (combination of high NYHA class, smaller aortic valve area and elevated pulmonary artery pressure) and a low EPC number were predictors of cardiac death in the univariate analysis. Multivariate logistic regression model identified low EPCs number and AS severity index as associated with cardiac death during follow up (p = 0.026 and p = 0.037, respectively). Conclusions EPC number is increased in patients with AS. However, in patients with moderate or severe AS a relatively low number of EPCs is associated with cardiac death at follow up. These results may help to identify AS patients at increased cardiovascular risk. PMID:26913741

  10. CD34+ Cells Represent Highly Functional Endothelial Progenitor Cells in Murine Bone Marrow

    PubMed Central

    Yang, Junjie; Ii, Masaaki; Kamei, Naosuke; Alev, Cantas; Kwon, Sang-Mo; Kawamoto, Atsuhiko; Akimaru, Hiroshi; Masuda, Haruchika; Sawa, Yoshiki; Asahara, Takayuki

    2011-01-01

    Background Endothelial progenitor cells (EPCs) were shown to have angiogenic potential contributing to neovascularization. However, a clear definition of mouse EPCs by cell surface markers still remains elusive. We hypothesized that CD34 could be used for identification and isolation of functional EPCs from mouse bone marrow. Methodology/Principal Findings CD34+ cells, c-Kit+/Sca-1+/Lin− (KSL) cells, c-Kit+/Lin− (KL) cells and Sca-1+/Lin− (SL) cells were isolated from mouse bone marrow mononuclear cells (BMMNCs) using fluorescent activated cell sorting. EPC colony forming capacity and differentiation capacity into endothelial lineage were examined in the cells. Although CD34+ cells showed the lowest EPC colony forming activity, CD34+ cells exhibited under endothelial culture conditions a more adherent phenotype compared with the others, demonstrating the highest mRNA expression levels of endothelial markers vWF, VE-cadherin, and Flk-1. Furthermore, a dramatic increase in immediate recruitment of cells to the myocardium following myocardial infarction and systemic cell injection was observed for CD34+ cells comparing with others, which could be explained by the highest mRNA expression levels of key homing-related molecules Integrin β2 and CXCR4 in CD34+ cells. Cell retention and incorporation into the vasculature of the ischemic myocardium was also markedly increased in the CD34+ cell-injected group, giving a possible explanation for significant reduction in fibrosis area, significant increase in neovascularization and the best cardiac functional recovery in this group in comparison with the others. Conclusion These findings suggest that mouse CD34+ cells may represent a functional EPC population in bone marrow, which could benefit the investigation of therapeutic EPC biology. PMID:21655289

  11. Altered SDF-1-mediated differentiation of bone marrow-derived endothelial progenitor cells in diabetes mellitus

    PubMed Central

    De Falco, Elena; Avitabile, Daniele; Totta, Pierangela; Straino, Stefania; Spallotta, Francesco; Cencioni, Chiara; Torella, Anna Rita; Rizzi, Roberto; Porcelli, Daniele; Zacheo, Antonella; Vito, Luca Di; Pompilio, Giulio; Napolitano, Monica; Melillo, Guido; Capogrossi, Maurizio C; Pesce, Maurizio

    2009-01-01

    In diabetic patients and animal models of diabetes mellitus (DM), circulating endothelial progenitor cell (EPC) number is lower than in normoglycaemic conditions and EPC angiogenic properties are inhibited. Stromal cell derived factor-1 (SDF-1) plays a key role in bone marrow (BM) c-kit+ stem cell mobilization into peripheral blood (PB), recruitment from PB into ischemic tissues and differentiation into endothelial cells. The aim of the present study was to examine the effect of DM in vivo and in vitro, on murine BM-derived c-kit+ cells and on their response to SDF-1. Acute hindlimb ischemia was induced in streptozotocin-treated DM and control mice; circulating c-kit+ cells exhibited a rapid increase followed by a return to control levels which was significantly faster in DM than in control mice. CXCR4 expression by BM c-kit+ cells as well as SDF-1 protein levels in the plasma and in the skeletal muscle, both before and after the induction of ischemia, were similar between normoglycaemic and DM mice. However, BM-derived c-kit+ cells from DM mice exhibited an impaired differentiation towards the endothelial phenotype in response to SDF-1; this effect was associated with diminished protein kinase phosphorylation. Interestingly, SDF-1 ability to induce differentiation of c-kit+ cells from DM mice was restored when cells were cultured under normoglycaemic conditions whereas c-kit+ cells from normoglycaemic mice failed to differentiate in response to SDF-1 when they were cultured in hyperglycaemic conditions. These results show that DM diminishes circulating c-kit+ cell number following hindlimb ischemia and inhibits SDF-1-mediated AKT phosphorylation and differentiation towards the endothelial phenotype of BM-derived c-kit+ cells. PMID:20196780

  12. Transplantation of vascular endothelial growth factor 165-transfected endothelial progenitor cells for the treatment of limb ischemia

    PubMed Central

    WANG, SHENG; CHEN, ZHONG; TANG, XIAOBIN; LIU, HUI; YANG, LIAO; WANG, YANYANG

    2015-01-01

    The present study aimed to investigate the effects of neovascularization in rabbits with limb ischemia transplanted with vascular endothelial growth factor (VEGF)165-transfected endothelial progenitor cells (EPC). Bone marrow mononuclear cells were isolated by gradient centrifugation, cultured in M199 culture medium and induced into EPCs using VEGF, basic fibroblast growth factor, and insulin-like growth factor-1, and subsequently identified. The EPCs were transfected with Adv-green fluorescent protein-VEGF165 and the proliferation potential of the cells was determined using an MTT assay. The protein expression levels of VEGF were measured by detecting its concentration levels in the supernatant using an ABC-ELISA assay. A rabbit hind limb ischemic model was established and randomly divided into three groups: (A) Control group, (B) EPC-transplanted group, and (C) Ad-VEGF165/EPCs-transplanted group. The effects of transplantation and the levels of recanalization were detected. Incorporation of the transplanted cells into the ischemic region was confirmed by 5-bromodeoxyuridine staining, and the levels of recanalization were measured by computer tomography ateriography and immunohistochemical staining. Bone marrow-derived EPCs were induced, cultivated, and successfully identified. The results of the present study determined the optimum transfection ratio that promoted the growth of EPCs. The EPCs were successfully transfected with VEGF165, and EPC proliferation was not affected by the transfection. The supernatant protein concentration levels of VEGF were markedly higher in the VEGF165-transfected group, as compared with those of the control group. Introduction of the transplanted cells into the ischemic region of group C occurred more efficiently, as compared with groups A and B. The recanalization capillary density in group C was significantly higher, as compared with groups A and B. VEGF gene transfection was able to improve the quality of EPCs, and the response

  13. Are Endothelial Progenitor Cells the Real Solution for Cardiovascular Diseases? Focus on Controversies and Perspectives

    PubMed Central

    Balistreri, Carmela R.; Buffa, Silvio; Pisano, Calogera; Lio, Domenico; Ruvolo, Giovanni; Mazzesi, Giuseppe

    2015-01-01

    Advanced knowledge in the field of stem cell biology and their ability to provide a cue for counteracting several diseases are leading numerous researchers to focus their attention on “regenerative medicine” as possible solutions for cardiovascular diseases (CVDs). However, the lack of consistent evidence in this arena has hampered the clinical application. The same condition affects the research on endothelial progenitor cells (EPCs), creating more confusion than comprehension. In this review, this aspect is discussed with particular emphasis. In particular, we describe biology and physiology of EPCs, outline their clinical relevance as both new predictive, diagnostic, and prognostic CVD biomarkers and therapeutic agents, discuss advantages, disadvantages, and conflicting data about their use as possible solutions for vascular impairment and clinical applications, and finally underline a very crucial aspect of EPCs “characterization and definition,” which seems to be the real cause of large heterogeneity existing in literature data on this topic. PMID:26509164

  14. Are Endothelial Progenitor Cells the Real Solution for Cardiovascular Diseases? Focus on Controversies and Perspectives.

    PubMed

    Balistreri, Carmela R; Buffa, Silvio; Pisano, Calogera; Lio, Domenico; Ruvolo, Giovanni; Mazzesi, Giuseppe

    2015-01-01

    Advanced knowledge in the field of stem cell biology and their ability to provide a cue for counteracting several diseases are leading numerous researchers to focus their attention on "regenerative medicine" as possible solutions for cardiovascular diseases (CVDs). However, the lack of consistent evidence in this arena has hampered the clinical application. The same condition affects the research on endothelial progenitor cells (EPCs), creating more confusion than comprehension. In this review, this aspect is discussed with particular emphasis. In particular, we describe biology and physiology of EPCs, outline their clinical relevance as both new predictive, diagnostic, and prognostic CVD biomarkers and therapeutic agents, discuss advantages, disadvantages, and conflicting data about their use as possible solutions for vascular impairment and clinical applications, and finally underline a very crucial aspect of EPCs "characterization and definition," which seems to be the real cause of large heterogeneity existing in literature data on this topic. PMID:26509164

  15. Overexpression of LOXIN Protects Endothelial Progenitor Cells From Apoptosis Induced by Oxidized Low Density Lipoprotein.

    PubMed

    Veas, Carlos; Jara, Casandra; Willis, Naomi D; Pérez-Contreras, Karen; Gutierrez, Nicolas; Toledo, Jorge; Fernandez, Paulina; Radojkovic, Claudia; Zuñiga, Felipe A; Escudero, Carlos; Aguayo, Claudio

    2016-04-01

    Human endothelial progenitor cells (hEPC) are adult stem cells located in the bone marrow and peripheral blood. Studies have indicated that hEPC play an important role in the recovery and repair of injured endothelium, however, their quantity and functional capacity is reduced in several diseases including hypercholesterolemia. Recently, it has been demonstrated that hEPC express lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) and its activation by oxidized low-density lipoprotein (ox-LDL) induces cellular dysfunction and apoptosis. This study aimed to investigate whether overexpression of LOXIN, a truncated isoform of LOX-1 that acts as a dominant negative, plays a protective role against ox-LDL-induced apoptosis in hEPC. Human endothelial progenitor cells exposed to ox-LDL showed a significant increase in LOX-1 expression, and apoptosis began at ox-LDL concentrations above 50 μg/mL. All hEPC apoptosed at 200 μg/mL ox-LDL. High LOXIN expression was generated using adenoviral systems in hEPC and SiHa cells transduced with 100 colony-forming units per cell. Transduced LOXIN localized to the plasma membrane and blocked ox-LDL uptake mediated by LOX-1. Overexpression of LOXIN protected hEPC from ox-LDL-induced apoptosis, and therefore maybe a novel way of improving hEPC function and quantity. These results suggest that adenoviral vectors of LOXIN may provide a possible treatment for diseases related to ox-LDL and vascular endothelium dysfunction, including atherosclerosis. PMID:26771151

  16. Particle Radiation-Induced Nontargeted Effects in Bone-Marrow-Derived Endothelial Progenitor Cells.

    PubMed

    Sasi, Sharath P; Park, Daniel; Muralidharan, Sujatha; Wage, Justin; Kiladjian, Albert; Onufrak, Jillian; Enderling, Heiko; Yan, Xinhua; Goukassian, David A

    2015-01-01

    Bone-marrow- (BM-) derived endothelial progenitor cells (EPCs) are critical for endothelial cell maintenance and repair. During future space exploration missions astronauts will be exposed to space irradiation (IR) composed of a spectrum of low-fluence protons ((1)H) and high charge and energy (HZE) nuclei (e.g., iron-(56)Fe) for extended time. How the space-type IR affects BM-EPCs is limited. In media transfer experiments in vitro we studied nontargeted effects induced by (1)H- and (56)Fe-IR conditioned medium (CM), which showed significant increase in the number of p-H2AX foci in nonirradiated EPCs between 2 and 24 h. A 2-15-fold increase in the levels of various cytokines and chemokines was observed in both types of IR-CM at 24 h. Ex vivo analysis of BM-EPCs from single, low-dose, full-body (1)H- and (56)Fe-IR mice demonstrated a cyclical (early 5-24 h and delayed 28 days) increase in apoptosis. This early increase in BM-EPC apoptosis may be the effect of direct IR exposure, whereas late increase in apoptosis could be a result of nontargeted effects (NTE) in the cells that were not traversed by IR directly. Identifying the role of specific cytokines responsible for IR-induced NTE and inhibiting such NTE may prevent long-term and cyclical loss of stem and progenitors cells in the BM milieu. PMID:26074973

  17. Reactive astrocytes promote adhesive interactions between brain endothelium and endothelial progenitor cells via HMGB1 and beta-2 integrin signaling

    PubMed Central

    Hayakawa, Kazuhide; Pham, Loc-Duyen D.; Arai, Ken; Lo, Eng H.

    2014-01-01

    Endothelial progenitor cells (EPCs) may contribute to neurovascular repair after stroke and neurodegeneration. A key step in this process should involve adhesive interactions between EPCs and the targeted cerebral endothelium. Here, we tested the hypothesis that reactive astrocytes may play a critical role in enhancing adhesive interactions and transmigration of EPCs across cerebral endothelial cells. Transiently seeding EPCs onto a monolayer of RBE.4 rat brain endothelial cells resulted in a time-dependent adherence between the two cell types. Blocking β2 integrins on EPCs or blocking the receptor for advanced glycation endproducts (RAGE) on endothelial cells significantly decreased EPC-endothelial adherence. Next, we tested whether reactive astrocytes can enhance this process by growing EPCs, brain endothelial cells and astrocytes together in a transwell co-culture system. The presence of reactive astrocytes in the lower chamber significantly promoted adherence between EPCs and endothelial cells in the upper chamber. This process involved the release of soluble HMGB1 from reactive astrocytes that then upregulated endothelial expression of RAGE via Egr1 signaling. Directly adding HMGB1 to the transwell system also promoted EPC-endothelial adhesion and accelerated EPC transmigration into the lower chamber. These initial findings provide proof-of-concept that reactive astrocytes promote crosstalk between cerebral endothelium and EPCs. Further investigation of this phenomenon may lead to a better understanding of cell-cell interactions required for neurovascular recovery after stroke. PMID:24480450

  18. Bone marrow-derived endothelial progenitor cells are involved in aneurysm repair in rabbits.

    PubMed

    Fang, Xinggen; Zhao, Rui; Wang, Kuizhong; Li, Zifu; Yang, Penfei; Huang, Qinghai; Xu, Yi; Hong, Bo; Liu, Jianmin

    2012-09-01

    Endothelial progenitor cells (EPC) are believed to be involved in aneurysmal repair and remodeling. The aim of this study was to test this hypothesis and, if true, explore how EPC contribute to aneurysm repair in a rabbit model of elastase-induced carotid aneurysm. Rabbits were divided randomly into an in situ carotid EPC transfusion group (ISCT group, n=5), and an intravenous EPC transfusion group (IVT group, n=5). Autologous EPC were double-labeled with Hoechst 33342 and 5,6-carboxyfluorescein diacetate succinimidyl ester before injection into the animals in either the carotid artery (ISCT group) or marginal ear veins (IVT group). Three weeks later, labeled cells in the aneurysms were observed with respect to location, adhesion, and growth to detect signs of aneurysm repair. Labeled EPC were detected within the neointima in all five aneurysms in the ISCT group and in three of the five aneurysms in the IVT group, but there was no endothelial growth in the aneurysmal neointima in either group. These results show that bone marrow-derived EPC are involved in the process of aneurysm repair in this rabbit model. PMID:22789632

  19. Correlation between increased circulating endothelial progenitor cells and stage of non-Hodgkin lymphoma.

    PubMed

    Yu, Dan-dan; Liu, Hong-li; Bai, Yun-lin; Wu, Bian; Chen, Wei-hong; Ren, Jing-hua; Zhang, Tao; Yang, Kun-yu; Wu, Gang

    2013-04-01

    This study aims to examine the levels of circulating endothelial progenitor cells (cEPCs) in the peripheral blood of patients with non-Hodgkin lymphoma (NHL) and their correlation with the tumor stage. Forty-one patients with biopsy-proven NHL and 16 healthy individuals were recruited. Peripheral blood mononuclear cells (PBMCs) were isolated by density gradient centrifugation, and cEPCs were characterized by triple staining using antibodies against CD133, CD34 and vascular endothelial growth factor receptor-2 (VEGFR-2, CD309) and quantified by flow cytometry. In NHL patients, the number of cEPCs was significantly greater than in control group (P=0.000). The cEPCs counts in patients with NHL of stage III-IV were significantly greater than in stage I-II (P=0.010). FACS analysis revealed that the number of cEPCs in NHL patients had no correlation with the gender (P=0.401) or the pathological category (P=0.852). It was suggested that the over-expression of cEPCs in NHL patients may serve as a novel biomarker for disease progression in NHL. PMID:23592145

  20. Endothelial Progenitor Cells in Tumor Angiogenesis: Another Brick in the Wall

    PubMed Central

    Marçola, Marina; Rodrigues, Camila Eleuterio

    2015-01-01

    Until 15 years ago, vasculogenesis, the formation of new blood vessels from undifferentiated cells, was thought to occur only during embryonic development. The discovery of circulating cells that are able to promote vascular regeneration and repair—the so-called endothelial progenitor cells (EPCs)—changed that, and EPCs have since been studied extensively. It is already known that EPCs include many subtypes of cells that play a variety of roles in promoting vascular growth. Some EPCs are destined to differentiate into endothelial cells, whereas others are capable of promoting and sustaining angiogenesis through paracrine mechanisms. Vasculogenesis and angiogenesis might constitute complementary mechanisms for postnatal neovascularization, and EPCs could be at the core of this process. Although the formation of new blood vessels from preexisting vasculature plays a beneficial role in many physiological processes, such as wound healing, it also contributes to tumor growth and metastasis. However, many aspects of the role played by EPCs in tumor angiogenesis remain unclear. This review aims to address the main aspects of EPCs differentiation and certain characteristics of their main function, especially in tumor angiogenesis, as well as the potential clinical applications. PMID:26000021

  1. Endothelial progenitor cells in tumor angiogenesis: another brick in the wall.

    PubMed

    Marçola, Marina; Rodrigues, Camila Eleuterio

    2015-01-01

    Until 15 years ago, vasculogenesis, the formation of new blood vessels from undifferentiated cells, was thought to occur only during embryonic development. The discovery of circulating cells that are able to promote vascular regeneration and repair-the so-called endothelial progenitor cells (EPCs)-changed that, and EPCs have since been studied extensively. It is already known that EPCs include many subtypes of cells that play a variety of roles in promoting vascular growth. Some EPCs are destined to differentiate into endothelial cells, whereas others are capable of promoting and sustaining angiogenesis through paracrine mechanisms. Vasculogenesis and angiogenesis might constitute complementary mechanisms for postnatal neovascularization, and EPCs could be at the core of this process. Although the formation of new blood vessels from preexisting vasculature plays a beneficial role in many physiological processes, such as wound healing, it also contributes to tumor growth and metastasis. However, many aspects of the role played by EPCs in tumor angiogenesis remain unclear. This review aims to address the main aspects of EPCs differentiation and certain characteristics of their main function, especially in tumor angiogenesis, as well as the potential clinical applications. PMID:26000021

  2. A Pilot Study of Circulating Endothelial and Hematopoietic Progenitor Cells in Children With Sarcomas.

    PubMed

    Pradhan, Kamnesh R; Mund, Julie A; Claussen, Heather L; Gosiengfiao, Yasmin C; Radulescu, Vlad C; Ballard, Jennifer J; Liu, Ziyue; Vik, Terry A; Case, Jamie

    2015-08-01

    Utilizing a multiparametric flow cytometry protocol, we assessed various cell types implicated in tumor angiogenesis that were found circulating in the peripheral blood of children with sarcomas (cases) based on their cell surface antigen expression. Circulating endothelial cells (CECs), endothelial colony-forming cells (ECFCs), and the ratio of 2 distinct populations of circulating hematopoietic stem and progenitor cells (CHSPCs), the proangiogenic CHSPCs (pCHSPCs) and nonangiogenic CHSPCs (nCHSPCs) were enumerated. Multiparametric flow cytometry was analyzed in cases at baseline and at 4 additional timepoints until the end of treatment and levels compared with each other and with healthy controls. At all timepoints, cases had significantly lower levels of CECs, but elevated ECFCs and a pCHSPC:nCHSPC ratio compared with controls (all P-values <0.05). There was no significant difference in any of the cell types analyzed based on tumor histology, stage (localized vs. metastatic), or tumor size. After treatment, only the CECs among the complete responders were significantly lower at end of therapy (P<0.01) compared with nonresponders, whereas the ECFCs among all cases significantly increased (P<0.05) compared with baseline. No decline in the pCHSPC:nCHSPC ratio was observed despite tumor response. On the basis of these results, a validation of CECs as prognostic biomarker is now warranted. PMID:26115508

  3. Butein Inhibits Angiogenesis of Human Endothelial Progenitor Cells via the Translation Dependent Signaling Pathway

    PubMed Central

    Chung, Ching-Hu; Chang, Chien-Hsin; Chen, Shiou-Sheng; Wang, Hsueh-Hsiao; Yen, Juei-Yu; Hsiao, Che-Jen; Wu, Nan-Lin; Chen, Yen-Ling; Huang, Tur-Fu; Wang, Po-Chuan; Yeh, Hung-I; Wang, Shih-Wei

    2013-01-01

    Compelling evidence indicates that bone marrow-derived endothelial progenitor cells (EPCs) can contribute to postnatal neovascularization and tumor angiogenesis. EPCs have been shown to play a “catalytic” role in metastatic progression by mediating the angiogenic switch. Understanding the pharmacological functions and molecular targets of natural products is critical for drug development. Butein, a natural chalcone derivative, has been reported to exert potent anticancer activity. However, the antiangiogenic activity of butein has not been addressed. In this study, we found that butein inhibited serum- and vascular endothelial growth factor- (VEGF-) induced cell proliferation, migration, and tube formation of human EPCs in a concentration dependent manner without cytotoxic effect. Furthermore, butein markedly abrogated VEGF-induced vessels sprouting from aortic rings and suppressed microvessel formation in the Matrigel implant assay in vivo. In addition, butein concentration-dependently repressed the phosphorylation of Akt, mTOR, and the major downstream effectors, p70S6K, 4E-BP1, and eIF4E in EPCs. Taken together, our results demonstrate for the first time that butein exhibits the antiangiogenic effect both in vitro and in vivo by targeting the translational machinery. Butein is a promising angiogenesis inhibitor with the potential for treatment of cancer and other angiogenesis-related diseases. PMID:23840271

  4. Irisin Increased the Number and Improved the Function of Endothelial Progenitor Cells in Diabetes Mellitus Mice

    PubMed Central

    Wang, Jinxiang; Song, Mingbao; Zhou, Fang; Fu, Dagan; Ruan, Guangping; Zhu, Xiangqing; Bai, Yinyin; Huang, Lan; Pang, Rongqing; Kang, Huali

    2016-01-01

    Abstract: The dysfunction of endothelial progenitor cells (EPCs) was found to be associated with vascular complications in diabetes mellitus (DM) patients. Previous studies found that regular exercise could improve the function of EPCs in DM patients, but the underling mechanism was unclear. Irisin, a newly identified myokine, was induced by exercise and has been demonstrated to mediate some of the positive effects of exercise. In this study, we hypothesize that irisin may have direct effects on EPC function in DM mice. These data showed for the first time that irisin increased the number of EPCs in peripheral blood of DM mice and improved the function of EPCs derived from DM mice bone marrow. The mechanism for the effect of irisin is related to the PI3K/Akt/eNOS pathway. Furthermore, irisin was demonstrated to improve endothelial repair in DM mice that received EPC transplants after carotid artery injury. The results of this study indicate a novel effect of irisin in regulating the number and function of EPCs via the PI3K/Akt/eNOS pathway, suggesting a potential for the administration of exogenous irisin as a succedaneum to improve EPC function in diabetic patients who fail to achieve such improvements through regular exercise. PMID:27002278

  5. Immortalized Functional Endothelial Progenitor Cell Lines from Umbilical Cord Blood for Vascular Tissue Engineering

    PubMed Central

    Sobhan, Praveen K.; Seervi, Mahendra; Joseph, Jeena; Varghese, Saneesh; Pillai, Prakash Rajappan; Sivaraman, Divya Mundackal; James, Jackson; George, Roshin Elizabeth; Elizabeth, K.E.; Pillai, M. Radhakrishna

    2012-01-01

    Endothelial progenitor cells (EPCs) play a significant role in multiple biological processes such as vascular homeostasis, regeneration, and tumor angiogenesis. This makes them a promising cell of choice for studying a variety of biological processes, toxicity assays, biomaterial–cell interaction studies, as well as in tissue-engineering applications. In this study, we report the generation of two clones of SV40-immortalized EPCs from umbilical cord blood. These cells retained most of the functional features of mature endothelial cells and showed no indication of senescence after repeated culture for more than 240 days. Extensive functional characterization of the immortalized cells by western blot, flow cytometry, and immunofluorescence studies substantiated that these cells retained their ability to synthesize nitric oxide, von Willebrand factor, P-Selectin etc. These cells achieved unlimited proliferation potential subsequent to inactivation of the cyclin-dependent kinase inhibitor p21, but failed to form colonies on soft agar. We also show their enhanced growth and survival on vascular biomaterials compared to parental cultures in late population doubling. These immortalized EPCs can be used as a cellular model system for studying the biology of these cells, gene manipulation experiments, cell–biomaterial interactions, as well as a variety of tissue-engineering applications. PMID:22889128

  6. Quercetin protects against high glucose-induced damage in bone marrow-derived endothelial progenitor cells.

    PubMed

    Zhao, Li-Rong; Du, Yu-Jun; Chen, Lei; Liu, Zhi-Gang; Pan, Yue-Hai; Liu, Jian-Feng; Liu, Bin

    2014-10-01

    Endothelial progenitor cells (EPCs), a group of bone marrow-derived pro-angiogenic cells, contribute to vascular repair after damage. EPC dysfunction exists in diabetes and results in poor wound healing in diabetic patients with trauma or surgery. The aim of the present study was to determine the effect of quercetin, a natural flavonoid on high glucose‑induced damage in EPCs. Treatment with high glucose (40 mM) decreased cell viability and migration, and increased oxidant stress, as was evidenced by the elevated levels of reactive oxygen species (ROS), malondialdehyde (MDA) and superoxide dismutase in bone marrow-derived EPCs. Moreover, high glucose reduced the levels of endothelial nitric oxide synthase (eNOS) phosphorylation, nitric oxide (NO) production and intracellular cyclic guanosine monophosphate (cGMP). Quercetin supplement protected against high glucose‑induced impairment in cell viability, migration, oxidant stress, eNOS phosphorylation, NO production and cGMP levels. Quercetin also increased Sirt1 expression in EPCs. Inhibition of Sirt1 by a chemical antagonist sirtinol abolished the protective effect of quercetin on eNOS phosphorylation, NO production and cGMP levels following high glucose stress. To the best of our knowledge, the results provide the first evidence that quercetin protects against high glucose‑induced damage by inducing Sirt1-dependent eNOS upregulation in EPCs, and suggest that quercetin is a promising therapeutic agent for diabetic patients undergoing surgery or other invasive procedures. PMID:25197782

  7. Absence of Correlation between Changes in the Number of Endothelial Progenitor Cell Subsets

    PubMed Central

    Attar, Armin; Parsanezhad, Mohammad Ebrahim; Namavar Jahromi, Bahia; Habibagahi, Mojtaba

    2015-01-01

    Background and Objectives Previously, various methodologies were used to enumerate the endothelial progenitor cells (EPCs). We now know that these methodologies enumerate at least three different EPC subsets: circulating angiogenic cells (CACs), colony-forming unit endothelial cells (CFU-ECs), and endothelial colony-forming cells (ECFCs). It is not clear whether there is a correlation between changes in the number of these subsets. The aim of the current study is to find an answer to this question. Materials and Methods The number of all EPC subsets was quantified in the peripheral blood of nine pregnant women in their first and third trimesters of pregnancy. We enumerated 14 cell populations by quantitative flow-cytometry using various combinations of the markers, CD34, CD133, CD309, and CD45, to cover most of the reported phenotypes of CACs and ECFCs. Culturing technique was used to enumerate the CFU-ECs. Changes in the number of cells were calculated by subtracting the number of cells in the first trimester peripheral blood from the number of cells in the third trimester peripheral blood, and correlations between these changes were analyzed. Results The number of CFU-ECs did not correlate with the number of ECFCs and CACs. Also, CACs and ECFCs showed independent behaviors. However, the number of CACs showed a strong correlation with the number of CD133+CD309+ cells (p=0.001) and a moderate correlation with the number of CD34+CD309+ cells (p=0.042). Also, the number of ECFCs was correlated with the number of CD309+CD45- cells (p=0.029) and CD34+CD45- cells (p=0.03). Conclusion Our study showed that the three commonly used methods for quantifying EPC subsets represent different cells with independent behaviors. Also, any study that measured the number of EPCs using the flow cytometry method with a marker combination that lacks CD309 may be inaccurate. PMID:26240587

  8. Prognostic relevance of circulating endothelial progenitor cells in patients with chronic heart failure.

    PubMed

    Koller, Lorenz; Hohensinner, Philipp; Sulzgruber, Patrick; Blum, Steffen; Maurer, Gerald; Wojta, Johann; Hülsmann, Martin; Niessner, Alexander

    2016-08-01

    Novel strategies for a tailored risk prediction in chronic heart failure (CHF) are crucial to identify patients at very high risk for an improved patient management and to specify treatment regimens. Endothelial progenitor cells (EPCs) are an important endogenous repair mechanism with the ability to counteract endothelial injury and the possibility of new vessel formation. We hypothesised that exhaustion of circulating EPCs may be a suitable prognostic biomarker in patients with CHF. EPCs, defined as CD34+CD45dimKDR+ cells, were analysed using fluorescence-activated cell sorting. EPCs were measured in 185 patients with CHF including 87 (47 %) patients with ischaemic aetiology and 98 (53 %) patients with non-ischaemic CHF and followed for a median time of 2.7 years. During this period, 34.7 % of patients experienced the primary study endpoint all-cause mortality. EPC count was a significant and independent inverse predictor of mortality with an hazard ratio hazard ratio (HR) per increase of one standard deviation (1-SD) of 0.47 (95 % confidence interval [CI]: 0.35-0.61; p<0.001) and remained significant after multivariable adjustment for a comprehensive set of cardiovascular risk factors and potential confounders with a HR per 1-SD of 0.54 (95 % CI: 0.4-0.73; p<0.001). EPCs further demonstrated additional prognostic information indicated by improvements in C-statistic, net reclassification index and integrated discrimination increment. In conclusion, in our study circulating EPCs turned out as strong and independent inverse predictors of mortality underlining the importance of an impaired endothelial repair mechanism in the pathophysiology and progression of CHF. PMID:27412580

  9. Application of anodized titanium for enhanced recruitment of endothelial progenitor cells

    PubMed Central

    2012-01-01

    Objectives To study the efficacy of an effective anodized titanium surface with enhanced attachment of endothelial progenitor cell (EPC). Background In-stent restenosis is a major obstacle for vascular patency after catheter-based intravascular interventions. Recently, stents that capture EPCs have been paid attention in order to make a functional endothelialized layer at the site of stent-induced endothelial denudation. Anodized titanium has been shown to enhance stem cell attachment. Anodization is a quick and inexpensive method, which can provide suitable stent surface. Methods Surface topography was examined by high-resolution scanning electron microscopy (SEM). Substrates were co-cultured with EPCs at second passage in 24-well culture plates. Evaluation of cell growth, proliferation, viability, surface cytotoxicity and cell adhesion was performed using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) test and 4,6-diamidino-2-phenylindole dihydrochloride staining. For platelet attachment, platelets added to substrates were evaluated under SEM. Results The average MTT values for tissue culture polystyrene plate, unanodized and anodized titanium with nanostructure were equal to 0.49, 0.16 and 0.72, respectively (P < 0.05). The surface had no cytotoxic effects on cells. The average cell attachment results showed that 9,955 ± 461.18, 3,300 ± 197.98 and 11,359 ± 458.10 EPCs were attached per well of tissue culture polystyrene plate, unanodized and anodized titanium surfaces, respectively (P < 0.05). Conclusions Anodized titanium surfaces can be potentially applied for devices that need enhanced recruitment of EPCs. This unique property makes these anodized surfaces good and cheap candidates for designing cardiovascular medical devices as endovascular stents. PMID:22676440

  10. Pancreatic-carcinoma-cell-derived pro-angiogenic factors can induce endothelial-cell differentiation of a subset of circulating CD34+ progenitors

    PubMed Central

    2013-01-01

    Background CD34+ progenitor cells comprise both hematopoietic and endothelial progenitor cells. Recent studies suggest that circulating endothelial progenitor cells are recruited into the angiogenic vascular system of several cancers, including pancreatic carcinoma, and that they correlate with clinical progress. However, whether endothelial progenitor cell mobilization occurs in response to cytokine release by tumor cells is still unclear. Methods The chemotactic- and/or differentiating-activities of the poorly-differentiated pancreatic carcinoma cell line PT45, and of the immortal H6c7 cell line, a line of near-normal pancreatic duct epithelial cells, on endothelial progenitor cells were investigated in vitro using circulating CD34+ as model. Results The study showed that Vascular Endothelial Growth Factor produced by PT45 cells and, at lesser extent, by H6c7 cells, predominantly chemoattract peripheral blood CD34+ expressing the type 2 relative receptor. Addition of PT45-conditioned medium to CD34+ cells, cultured under conditions supporting myeloid cell development, diverted the differentiation of a subset of these progenitor cells into cells expressing endothelial cell markers, such as CD146, CD105, VE-cadherin and von Willebrand Factor-related antigen. Moreover, these endothelial-like cells formed capillary networks in vitro, chiefly through the release of Angiopoietin-1 by PT45 cells. Conclusions The results demonstrate that pancreatic-carcinoma cells potentially attract circulating endothelial progenitor cells to the tumor site, by releasing high levels of pro-angiogenic factors such as Vascular Endothelial Growth Factor and Angiopoietin-1, and may direct the differentiation of these cell subsets of the CD34+ cell population into endothelial cells; the latter cells may become a component of the newly-formed vessels, contributing to angiogenesis-mediated tumor growth and metastasis. PMID:24341512

  11. Myocardial regeneration by transplantation of modified endothelial progenitor cells expressing SDF-1 in a rat model

    PubMed Central

    Schuh, Alexander; Kroh, Andreas; Konschalla, Simone; Liehn, Elisa A; Sobota, Radoslav M; Biessen, Erik AL; Bot, Ilze; Sönmez, Tolga Taha; Schober, Andreas; Marx, Nikolaus; Weber, Christian; Sasse, Alexander

    2012-01-01

    Cell based therapy has been shown to attenuate myocardial dysfunction after myocardial infarction (MI) in different acute and chronic animal models. It has been further shown that stromal-cell derived factor-1α (SDF-1α) facilitates proliferation and migration of endogenous progenitor cells into injured tissue. The aim of the present study was to investigate the role of exogenously applied and endogenously mobilized cells in a regenerative strategy for MI therapy. Lentivirally SDF-1α-infected endothelial progenitor cells (EPCs) were injected after 90 min. of ligation and reperfusion of the left anterior descending artery (LAD) intramyocardial and intracoronary using a new rodent catheter system. Eight weeks after transplantation, echocardiography and isolated heart studies revealed a significant improvement of LV function after intramyocardial application of lentiviral with SDF-1 infected EPCs compared to medium control. Intracoronary application of cells did not lead to significant differences compared to medium injected control hearts. Histology showed a significantly elevated rate of apoptotic cells and augmented proliferation after transplantation of EPCs and EPCs + SDF-1α in infarcted myocardium. In addition, a significant increased density of CD31+ vessel structures, a lower collagen content and higher numbers of inflammatory cells after transplantation of SDF-1 transgenic cells were detectable. Intramyocardial application of lentiviral-infected EPCs is associated with a significant improvement of myocardial function after infarction, in contrast to an intracoronary application. Histological results revealed a significant augmentation of neovascularization, lower collagen content, higher numbers of inflammatory cells and remarkable alterations of apoptotic/proliferative processes in infarcted areas after cell transplantation. PMID:22288686

  12. Transplantation of endothelial progenitor cells in treating rats with IgA nephropathy

    PubMed Central

    2014-01-01

    Background Therapeutic options in IgAN are still limited. The aim of this study is to explore the feasibility of using endothelial progenitor cell to treat IgAN in rat model. Methods Rat bone marrow mononuclear cells (BM-MNCs) obtained with density gradient centrifugation were cultured in vitro, and induced into endothelial progenitor cells (EPCs). EPCs were identified by surface marker CD34, CD133 and VEGFR2 (FLK-1) and by Dil-Ac-LDL/FITC-UEA-1 double staining. EPCs were labeled with PKH26 prior to transplantation. Rat model of IgAN was established by oral administration of bovine serum albumin together with lipopolysaccharide via the caudal vein and subcutaneous injection of CCL4. Kidney paraffin sections were stained by H&E and PAS. Immunofluorescence was used to assess IgA deposition in the glomeruli. Peritubular capillary (PTC) density was determined by CD31 staining. Monocyte chemoattrant protein-1 (MCP-1), hypoxia-inducible factor-1α (HIF-1α) and CD105 were also measured by immunohistochemistry, western blotting and real-time fluorescent quantitative PCR. Results The transplanted BM-EPCs were successfully located in IgAN rat kidney. After transplantation, Urinary red blood cell, urine protein, BUN, Scr and IgA serum level were significantly decreased, so were the areas of glomerular extracellular matrix and the IgA deposition in the glomeruli. In addition, PTC density was elevated. And the expression levels of HIF-1α and MCP-1 were significantly down-regulated, while the expression of CD105 was up-regulated. All these changes were not observed in control groups. Conclusion The BM-EPCs transplantation significantly decreases the expansion of glomerular extracellular matrix and the deposition of IgA in the glomeruli; lowers the expression of inflammatory factors; increases PTC density; improves ischemic-induced renal tissue hypoxia, all of which improves the renal function and slows the progress of IgA nephropathy. PMID:25012471

  13. Impaired Endothelial Repair Capacity of Early Endothelial Progenitor Cells in Hypertensive Patients With Primary Hyperaldosteronemia: Role of 5,6,7,8-Tetrahydrobiopterin Oxidation and Endothelial Nitric Oxide Synthase Uncoupling.

    PubMed

    Chen, Long; Ding, Mei-Lin; Wu, Fang; He, Wen; Li, Jin; Zhang, Xiao-Yu; Xie, Wen-Li; Duan, Sheng-Zhong; Xia, Wen-Hao; Tao, Jun

    2016-02-01

    Although hyperaldosteronemia exerts detrimental impacts on vascular endothelium in addition to elevating blood pressure, the effects and molecular mechanisms of hyperaldosteronemia on early endothelial progenitor cell (EPC)-mediated endothelial repair after arterial damage are yet to be determined. The aim of this study was to investigate the endothelial repair capacity of early EPCs from hypertensive patients with primary hyperaldosteronemia (PHA). In vivo endothelial repair capacity of early EPCs from PHAs (n=20), age- and blood pressure-matched essential hypertension patients (n=20), and age-matched healthy subjects (n=20) was evaluated by transplantation into a nude mouse carotid endothelial denudation model. Endothelial function was evaluated by flow-mediated dilation of brachial artery in human subjects. In vivo endothelial repair capacity of early EPCs and flow-mediated dilation were impaired both in PHAs and in essential hypertension patients when compared with age-matched healthy subjects; however, the early EPC in vivo endothelial repair capacity and flow-mediated dilation of PHAs were impaired more severely than essential hypertension patients. Oral spironolactone improved early EPC in vivo endothelial repair capacity and flow-mediated dilation of PHAs. Increased oxidative stress, oxidative 5,6,7,8-tetrahydrobiopterin degradation, endothelial nitric oxide synthase uncoupling and decreased nitric oxide production were found in early EPCs from PHAs. Nicotinamide adenine dinucleotide phosphate oxidase subunit p47(phox) knockdown or 5,6,7,8-tetrahydrobiopterin supplementation attenuated endothelial nitric oxide synthase uncoupling and enhanced in vivo endothelial repair capacity of early EPCs from PHAs. In conclusion, PHAs exhibited more impaired endothelial repair capacity of early EPCs than did essential hypertension patients independent of blood pressure, which was associated with mineralocorticoid receptor-dependent oxidative stress and subsequently 5

  14. Statins Enhance Clonal Growth of Late Outgrowth Endothelial Progenitors and Increase Myocardial Capillary Density in the Chronically Ischemic Heart

    PubMed Central

    Wang, Wen; Lang, Jennifer K.; Suzuki, Gen; Canty, John M.; Cimato, Thomas

    2011-01-01

    Background Coronary artery disease and ischemic heart disease are leading causes of heart failure and death. Reduced blood flow to heart tissue leads to decreased heart function and symptoms of heart failure. Therapies to improve heart function in chronic coronary artery disease are important to identify. HMG-CoA reductase inhibitors (statins) are an important therapy for prevention of coronary artery disease, but also have non-cholesterol lowering effects. Our prior work showed that pravastatin improves contractile function in the chronically ischemic heart in pigs. Endothelial progenitor cells are a potential source of new blood vessels in ischemic tissues. While statins are known to increase the number of early outgrowth endothelial progenitor cells, their effects on late outgrowth endothelial progenitor cells (LOEPCs) and capillary density in ischemic heart tissue are not known. We hypothesized that statins exert positive effects on the mobilization and growth of late outgrowth EPCs, and capillary density in ischemic heart tissue. Methodology/Principal Findings We determined the effects of statins on the mobilization and growth of late outgrowth endothelial progenitor cells from pigs. We also determined the density of capillaries in myocardial tissue in pigs with chronic myocardial ischemia with or without treatment with pravastatin. Pravastatin therapy resulted in greater than two-fold increase in CD31+ LOEPCs versus untreated animals. Addition of pravastatin or simvastatin to blood mononuclear cells increased the number of LOEPCs greater than three fold in culture. Finally, in animals with chronic myocardial ischemia, pravastatin increased capillary density 46%. Conclusions Statins promote the derivation, mobilization, and clonal growth of LOEPCs. Pravastatin therapy in vivo increases myocardial capillary density in chronically ischemic myocardium, providing an in vivo correlate for the effects of statins on LOEPC growth in vitro. Our findings provide

  15. Adiponectin Pretreatment Counteracts the Detrimental Effect of a Diabetic Environment on Endothelial Progenitors

    PubMed Central

    Leicht, Simon F.; Schwarz, Theresa M.; Hermann, Patrick C.; Seissler, Jochen; Aicher, Alexandra; Heeschen, Christopher

    2011-01-01

    OBJECTIVE It has been shown that vascular progenitors from patients with diabetes are dysfunctional. However, therapeutic strategies to counteract their reduced functional capacity are still lacking. Because adiponectin has reported salutary effects on endothelial function, we investigated the functional effects of globular adiponectin (gAcrp), the active domain of adiponectin, on isolated endothelial colony-forming cells (ECFC). RESEARCH DESIGN AND METHODS ECFC were isolated from peripheral blood of type 2 diabetic patients (dmECFC) and compared with ECFC of healthy young volunteers (yECFC) and nondiabetic age-matched control subjects (hECFC). Cells were treated with gAcrp for 48 h followed by assessment of cell counts, cell cycle analysis, and migration capacity. For in vivo evaluation, human ECFC were injected into normoglycemic or streptozotocin-induced hyperglycemic nu/nu mice after hind limb ischemia. RESULTS Whereas dmECFC were functionally impaired compared with yECFC and hECFC, gAcrp significantly enhanced their in vitro proliferation and migratory activity. In vitro effects were significantly stronger in hECFC compared with dmECFC and were mediated through the cyclooxygenase-2 pathway. Most important, however, we observed a profound and sustained increase of the in vivo neovascularization in mice receiving gAcrp-pretreated dmECFC compared with untreated dmECFC under both normoglycemic and hyperglycemic conditions. CONCLUSIONS Pretreatment of ECFC with gAcrp enhanced the functional capacity of ECFC in vitro and in vivo in normoglycemic and hyperglycemic environments. Therefore, preconditioning of dmECFC with gAcrp may be a novel approach to counteract their functional impairment in diabetes. PMID:21270275

  16. High-mobility group box 1 activates integrin-dependent homing of endothelial progenitor cells.

    PubMed

    Chavakis, Emmanouil; Hain, Andreas; Vinci, Maria; Carmona, Guillaume; Bianchi, Marco E; Vajkoczy, Peter; Zeiher, Andreas M; Chavakis, Triantafyllos; Dimmeler, Stefanie

    2007-02-01

    Endothelial progenitor cells (EPCs) are recruited to ischemic regions and improve neovascularization. Integrins contribute to EPC homing. High-mobility group box 1 (HMGB1) is a nuclear protein that is released extracellularly on cell necrosis and tissue damage, eliciting a proinflammatory response and stimulating tissue repair. In the present study, we investigated the effects of HMGB1 on EPC homing. EPCs express the HMGB1 receptors RAGE (receptor for advanced glycation end products) and TLR2 (Toll-like receptor 2). EPC migration was stimulated by HMGB1 in a RAGE-dependent manner. In addition, the HMGB1-induced migration of EPCs on fibronectin and fibrinogen was significantly inhibited by antibodies against beta1 and beta2 integrins, respectively. Short-term prestimulation of EPCs with HMGB1 also increased EPC adhesion to endothelial cell monolayers, and this effect was blocked by antibodies to beta2 integrins or RAGE. HMGB1 increased EPC adhesion to the immobilized integrin ligands intercellular adhesion molecule-1 and fibronectin in a RAGE-dependent manner. Strikingly, HMGB1 rapidly increased integrin affinity and induced integrin polarization. Using intravital microscopy in a tumor model of neovascularization, prestimulation of EPCs with HMGB1 enhanced the initial in vivo adhesion of EPCs to microvessels and the recruitment of EPCs in the tumor tissue. In addition, prestimulation of EPCs with HMGB1 increased the homing of EPCs to ischemic muscles. In conclusion, these data represent a link between HMGB1 and integrin functions of EPCs and demonstrate that HMGB1 stimulates EPC homing to ischemic tissues. These results may provide a platform for the development of novel therapeutic approaches to improve EPC homing. PMID:17218606

  17. NOTCH4 signaling controls EFNB2-induced endothelial progenitor cell dysfunction in preeclampsia.

    PubMed

    Liu, Xiaoxia; Luo, Qingqing; Zheng, Yanfang; Liu, Xiaoping; Hu, Ying; Liu, Weifang; Luo, Minglian; Zhao, Yin; Zou, Li

    2016-07-01

    Preeclampsia is a serious complication of pregnancy and is closely related to endothelial dysfunction, which can be repaired by endothelial progenitor cells (EPCs). The DLL4/NOTCH-EFNB2 (ephrinB2) cascade may be involved in the pathogenesis of preeclampsia by inhibiting the biological activity of EPCs. In addition, both NOTCH1 and NOTCH4, which are specific receptors for DLL4/NOTCH, play critical roles in the various steps of angiogenesis. However, it has not been determined which receptor (NOTCH1, NOTCH4, or both) is specific for the DLL4/NOTCH-EFNB2 cascade. Accordingly, we performed a series of investigations to evaluate it. EFNB2 expression was examined when NOTCH4 or NOTCH1 was downregulated, with or without DLL4 treatment. Then, the effects of NOTCH4 on EPC function were detected. Additionally, we analyzed NOTCH4 and EFNB2 expression in the EPCs from preeclampsia and normal pregnancies. Results showed that NOTCH4 downregulation led to decreased expression of EFNB2, which maintained the same level in the presence of DLL4/NOTCH activation. By contrast, NOTCH1 silencing resulted in a moderate increase in EFNB2 expression, which further increased in the presence of DLL4/NOTCH activation. The downregulation of NOTCH4 resulted in an increase of EPC biological activity, which was similar to EFNB2 silencing. NOTCH4 expression, consistent with the EFNB2 level, increased notably in preeclampsia EPCs compared with the controls. These findings suggest that NOTCH4, not NOTCH1, is the specific receptor for the DLL4/NOTCH-EFNB2 cascade. Blockade of this cascade may enhance the angiogenic property of EPCs, and act as a potential target to promote angiogenesis in patients with preeclampsia. PMID:27069008

  18. Postischemic microvasculopathy and endothelial progenitor cell-based therapy in ischemic AKI: update and perspectives.

    PubMed

    Patschan, D; Kribben, A; Müller, G A

    2016-08-01

    Acute kidney injury (AKI) dramatically increases mortality of hospitalized patients. Incidences have been increased in recent years. The most frequent cause is transient renal hypoperfusion or ischemia which induces significant tubular cell dysfunction/damage. In addition, two further events take place: interstitial inflammation and microvasculopathy (MV). The latter evolves within minutes to hours postischemia and may result in permanent deterioration of the peritubular capillary network, ultimately increasing the risk for chronic kidney disease (CKD) in the long term. In recent years, our understanding of the molecular/cellular processes responsible for acute and sustained microvasculopathy has increasingly been expanded. The methodical approaches for visualizing impaired peritubular blood flow and increased vascular permeability have been optimized, even allowing the depiction of tissue abnormalities in a three-dimensional manner. In addition, endothelial dysfunction, a hallmark of MV, has increasingly been recognized as an inductor of both vascular malfunction and interstitial inflammation. In this regard, so-called regulated necrosis of the endothelium could potentially play a role in postischemic inflammation. Endothelial progenitor cells (EPCs), represented by at least two major subpopulations, have been shown to promote vascular repair in experimental AKI, not only in the short but also in the long term. The discussion about the true biology of the cells continues. It has been proposed that early EPCs are most likely myelomonocytic in nature, and thus they may simply be termed proangiogenic cells (PACs). Nevertheless, they reliably protect certain types of tissues/organs from ischemia-induced damage, mostly by modulating the perivascular microenvironment in an indirect manner. The aim of the present review is to summarize the current knowledge on postischemic MV and EPC-mediated renal repair. PMID:27194716

  19. Enhanced angiogenic potency of monocytic endothelial progenitor cells in patients with systemic sclerosis

    PubMed Central

    2010-01-01

    Introduction Microvasculopathy is one of the characteristic features in patients with systemic sclerosis (SSc), but underlying mechanisms still remain uncertain. In this study, we evaluated the potential involvement of monocytic endothelial progenitor cells (EPCs) in pathogenic processes of SSc vasculopathy, by determining their number and contribution to blood vessel formation through angiogenesis and vasculogenesis. Methods Monocytic EPCs were enriched and enumerated using a culture of peripheral blood mononuclear cells and platelets on fibronectin in 23 patients with SSc, 22 patients with rheumatoid arthritis (RA), and 21 healthy controls. To assess the capacity of monocytic EPCs to promote vascular formation and the contribution of vasculogenesis to this process, we used an in vitro co-culture system with human umbilical vein endothelial cells (HUVECs) on Matrigel® and an in vivo murine tumor neovascularization model. Results Monocytic EPCs were significantly increased in SSc patients than in RA patients or healthy controls (P = 0.01 for both comparisons). Monocytic EPCs derived from SSc patients promoted tubular formation in Matrigel® cultures more than those from healthy controls (P = 0.007). Transplantation of monocytic EPCs into immunodeficient mice resulted in promotion of tumor growth and blood vessel formation, and these properties were more prominent in SSc than healthy monocytic EPCs (P = 0.03 for both comparisons). In contrast, incorporation of SSc monocytic EPCs into the tubular structure was less efficient in vitro and in vivo, compared with healthy monocytic EPCs. Conclusions SSc patients have high numbers of aberrant circulating monocytic EPCs that exert enhanced angiogenesis but are impaired in vasculogenesis. However, these cells apparently cannot overcome the anti-angiogenic environment that characterizes SSc-affected tissues. PMID:21050433

  20. IFNα Serum Levels Are Associated with Endothelial Progenitor Cells Imbalance and Disease Features in Rheumatoid Arthritis Patients

    PubMed Central

    Rodríguez-Carrio, Javier; de Paz, Banesa; López, Patricia; Prado, Catuxa; Alperi-López, Mercedes; Ballina-García, Francisco Javier; Suárez, Ana

    2014-01-01

    Introduction IFNα has been largely implicated in the ethiopathogenesis of autoimmune diseases but only recently it has been linked to endothelial damage and accelerated atherosclerosis in autoimmunity. In addition, proinflammatory conditions are supposed to be implicated in the cardiovascular status of these patients. Since a role for IFNα in endothelial damage and impaired Endothelial Progenitor Cell (EPC) number and function has been reported in other diseases, we aimed to evaluate the potential associations of IFNα serum levels on EPC populations and cytokine profiles in Rheumatoid Arthritis (RA) patients. Methods pre-EPC, EPC and mature EPC (mEPC) populations were quantified by flow cytometry analyzing their differential CD34, CD133 and VEGFR2 expression in blood samples from 120 RA patients, 52 healthy controls (HC), and 83 systemic lupus erythematosus (SLE) patients as disease control. Cytokine serum levels were measured by immunoassays and clinical and immunological data, including cardiovascular (CV) events and CV risk factors, were retrospectively obtained by reviewing clinical records. Results Long-standing, but not recent onset RA patients displayed a significant depletion of all endothelial progenitor populations, unless high IFNα levels were present. In fact, the IFNhigh RA patient group (n = 40, 33%), showed increased EPC levels, comparable to SLE patients. In addition, high IFNα serum levels were associated with higher disease activity (DAS28), presence of autoantibodies, higher levels of IL-1β, IL-6, IL-10 and MIP-1α, lower amounts of TGF-β, and increased mEPC/EPC ratio, thus suggesting higher rates of endothelial damage and an endothelial repair failure. Finally, the relationship between high IFNα levels and occurrence of CV events observed in RA patients seems to support this hypothesis. Conclusions IFNα serum marker could be used to identify a group of RA patients with increased disease activity, EPC imbalance, enhanced

  1. Changes of Number and Function of Late Endothelial Progenitor Cells in Peripheral Blood of COPD Patients Combined with Pulmonary Hypertension.

    PubMed

    Liu, Pei; Zhang, Hongmei; Liu, Jianxin; Sheng, Chunfeng; Zhang, Linlin; Zeng, Yanjun

    2016-06-01

    Objective The objective of this study was to investigate the changes of number and function of late endothelial progenitor cells (EPCs) in peripheral blood of chronic obstructive pulmonary disease (COPD) patients combined with pulmonary hypertension. Subjects and Methods The study enrolled 120 cases including 40 non-COPD and pulmonary arterial hypertension (PAH) patients (non-COPD group), 40 COPD non-PAH patients (COPD group), and 40 COPD patients combined with PAH (COPD + PAH group). Peripheral blood mononuclear cells were separated by density gradient centrifugation, cultured for 21 days, and then identified as late endothelial progenitor cells. The cell colonies were counted. MTT assay, modified Boyden chamber assay, and human fibronectin plates were used to measure the proliferation, migration, and adhesion functions of the late endothelial progenitor cells, respectively. Results Compared with non-COPD and COPD groups, the number of peripheral blood late EPCs in COPD + PAH group was significantly reduced, and the proliferation, adhesion, and migration capacities were significantly lowered; the differences were statistically significant (p < 0.05). The number and function of late EPCs decreased with the increase of pulmonary artery pressure (p < 0.05). Conclusion The number of late EPCs in COPD patients combined with pulmonary hypertension was reduced, which implies the impaired cell functions. The changes of number and function were negatively correlated with the severity of pulmonary hypertension. PMID:25226359

  2. Trichostatin A enhances vascular repair by injected human endothelial progenitors through increasing the expression of TAL1-dependent genes.

    PubMed

    Palii, Carmen G; Vulesevic, Branka; Fraineau, Sylvain; Pranckeviciene, Erinija; Griffith, Alexander J; Chu, Alphonse; Faralli, Hervé; Li, Yuhua; McNeill, Brian; Sun, Jie; Perkins, Theodore J; Dilworth, F Jeffrey; Perez-Iratxeta, Carol; Suuronen, Erik J; Allan, David S; Brand, Marjorie

    2014-05-01

    A major goal of cell therapy for vascular diseases is to promote revascularization through the injection of endothelial stem/progenitor cells. The gene regulatory mechanisms that underlie endothelial progenitor-mediated vascular repair, however, remain elusive. Here, we identify the transcription factor TAL1/SCL as a key mediator of the vascular repair function of primary human endothelial colony-forming cells (ECFCs). Genome-wide analyses in ECFCs demonstrate that TAL1 activates a transcriptional program that promotes cell adhesion and migration. At the mechanistic level, we show that TAL1 upregulates the expression of migratory and adhesion genes through recruitment of the histone acetyltransferase p300. Based on these findings, we establish a strategy that enhances the revascularization efficiency of ECFCs after ischemia through ex vivo priming with the histone deacetylase inhibitor TSA. Thus, small molecule epigenetics drugs are effective tools for modifying the epigenome of stem/progenitor cells prior to transplantation as a means to enhance their therapeutic potential. PMID:24792117

  3. Experimental study on apoptosis of TNFR1 receptor pro-endothelial progenitor cells activated by high glucose induced oxidative stress

    PubMed Central

    Liu, Yong; Xei, Fei; Xu, Xiong-Fei; Zeng, Hong; He, Hu-Qiang; Zhang, Lei; Zheng, Ying-Qiang; He, Yan-Zheng

    2015-01-01

    Objective: To investigate whether high glucose in vitro activating TNFR1 and further promote rat marrow endothelial progenitor cells (EPCs) apoptosis. Methods: Rat morrow endothelial progenitor cells were cultured and identified by Confocal Microscopy; then were treated with high glucose (5.5, 15, 30, 60 mmol/L), mannitol (15, 30, 60, 90 mmol/L), high glucose + Tempol and high glucose+ MAB430. Apoptosis rate of the above cells were detected by flow cytometry. ROS and MDA level and anti-O2- were detected by colorimetric technique; the expression level of TNFR1 induced signal pathway related proteins were detected by Western blotting. Results: High glucose can induce endothelial progenitor cells apoptosis, which is mostly in the later stage (72 h-96 h) instead of the earlier stage (24 h-48 h); high glucose can also induce oxidative stress reaction and the produces ROS and MDA increase significantly in the later stage (after 72 h), but anti-O2- decrease significantly. TNF apoptosis signal pathway related protein expression level not increase in the earlier stage (before 24 h) but increase significantly in the later stage (after 72 h). Tempol and MAB430 down-regulate TNF apoptosis signal pathway related protein expression and reduce EPCs apoptosis. Conclusion: High glucose activates the TNFR1 of TPCs through oxidative stress reaction and further induces cell apoptosis. PMID:26884909

  4. Promotion of adhesion and proliferation of endothelial progenitor cells on decellularized valves by covalent incorporation of RGD peptide and VEGF.

    PubMed

    Zhou, Jianliang; Ding, Jingli; Nie, Bin'en; Hu, Shidong; Zhu, Zhigang; Chen, Jia; Xu, Jianjun; Shi, Jiawei; Dong, Nianguo

    2016-09-01

    Tissue engineered heart valve is a promising alternative to current heart valve surgery, for its capability of growth, repair, and remodeling. However, extensive development is needed to ensure tissue compatibility, durability and antithrombotic potential. This study aims to investigate the biological effects of multi-signal composite material of polyethyl glycol-cross-linked decellularized valve on adhesion and proliferation of endothelial progenitor cells. Group A to E was decellularized valve leaflets, composite material of polyethyl glycol-cross-linked decellularized valves leaflets, vascular endothelial growth factor-composite materials, Arg-Gly-Asp peptide-composite materials and multi-signal modified materials of polyethyl glycol-cross-linked decellularized valve leaflets, respectively. The endothelial progenitor cells were seeded for each group, cell adhesion and proliferation were detected and neo-endothelium antithrombotic function of the multi-signal composite materials was evaluated. At 2, 4, and 8 h after the seeding, the cell numbers and 3H-TdR incorporation in group D were the highest. At 2, 4, and 8 days after the seeding, the cell numbers and 3H-TdR incorporation were significantly higher in groups C, D, and E compared with groups A and B (P < 0.05) and cell numbers and the expression of t-PA and eons in the neo-endothelium were quite similar to those in the human umbilical vein endothelial cells at 2, 4, and 8 days after the seeding. The Arg-Gly-Asp- peptides (a sequential peptide composed of arginine (Arg), glycine (Gly) and aspartic acid (Asp)) and VEGF-conjugated onto the composite material of PEG-crosslinked decellularized valve leaflets synergistically promoted the adhesion and proliferation of endothelial progenitor cells on the composite material, which may help in tissue engineering of heart valves. PMID:27541486

  5. Amlodipine Ameliorates Ischemia-Induced Neovascularization in Diabetic Rats through Endothelial Progenitor Cell Mobilization.

    PubMed

    Sun, Jiayin; Xie, Jun; Kang, Lina; Ferro, Albert; Dong, Li; Xu, Biao

    2016-01-01

    Objectives. We investigated whether amlodipine could improve angiogenic responses in a diabetic rat model of acute myocardial infarction (AMI) through improving bone marrow endothelial progenitor cell (EPC) mobilization, in the same way as angiotensin converting enzyme inhibitors. Methods. After induction of AMI by coronary artery ligation, diabetic rats were randomly assigned to receive perindopril (2 mgkg(-1) day(-1)), amlodipine (2.5 mgkg(-1) day(-1)), or vehicle by gavage (n = 20 per group). Circulating EPC counts before ligation and on days 1, 3, 5, 7, 14, and 28 after AMI were measured in each group. Microvessel density, cardiac function, and cardiac remodeling were assessed 4 weeks after treatment. The signaling pathway related to EPC mobilization was also measured. Results. Circulating EPC count in amlodipine- and perindopril-treated rats peaked at day 7, to an obvious higher level than the control group peak which was reached earlier (at day 5). Rats treated with amlodipine showed improved postischemia neovascularization and cardiac function, together with reduced cardiac remodeling, decreased interstitial fibrosis, and cardiomyocyte apoptosis. Amlodipine treatment also increased cardiac SDF-1/CXCR4 expression and gave rise to activation of VEGF/Akt/eNOS signaling in bone marrow. Conclusions. Amlodipine promotes neovascularization by improving EPC mobilization from bone marrow in diabetic rats after AMI, and activation of VEGF/Akt/eNOS signaling may in part contribute to this. PMID:27243031

  6. Amlodipine Ameliorates Ischemia-Induced Neovascularization in Diabetic Rats through Endothelial Progenitor Cell Mobilization

    PubMed Central

    Sun, Jiayin; Xie, Jun; Kang, Lina; Ferro, Albert; Dong, Li; Xu, Biao

    2016-01-01

    Objectives. We investigated whether amlodipine could improve angiogenic responses in a diabetic rat model of acute myocardial infarction (AMI) through improving bone marrow endothelial progenitor cell (EPC) mobilization, in the same way as angiotensin converting enzyme inhibitors. Methods. After induction of AMI by coronary artery ligation, diabetic rats were randomly assigned to receive perindopril (2 mgkg−1 day−1), amlodipine (2.5 mgkg−1 day−1), or vehicle by gavage (n = 20 per group). Circulating EPC counts before ligation and on days 1, 3, 5, 7, 14, and 28 after AMI were measured in each group. Microvessel density, cardiac function, and cardiac remodeling were assessed 4 weeks after treatment. The signaling pathway related to EPC mobilization was also measured. Results. Circulating EPC count in amlodipine- and perindopril-treated rats peaked at day 7, to an obvious higher level than the control group peak which was reached earlier (at day 5). Rats treated with amlodipine showed improved postischemia neovascularization and cardiac function, together with reduced cardiac remodeling, decreased interstitial fibrosis, and cardiomyocyte apoptosis. Amlodipine treatment also increased cardiac SDF-1/CXCR4 expression and gave rise to activation of VEGF/Akt/eNOS signaling in bone marrow. Conclusions. Amlodipine promotes neovascularization by improving EPC mobilization from bone marrow in diabetic rats after AMI, and activation of VEGF/Akt/eNOS signaling may in part contribute to this. PMID:27243031

  7. Thickness-controllable electrospun fibers promote tubular structure formation by endothelial progenitor cells.

    PubMed

    Hong, Jong Kyu; Bang, Ju Yup; Xu, Guan; Lee, Jun-Hee; Kim, Yeon-Ju; Lee, Ho-Jun; Kim, Han Seong; Kwon, Sang-Mo

    2015-01-01

    Controlling the thickness of an electrospun nanofibrous scaffold by altering its pore size has been shown to regulate cell behaviors such as cell infiltration into a three-dimensional (3D) scaffold. This is of great importance when manufacturing tissue-engineering scaffolds using an electrospinning process. In this study, we report the development of a novel process whereby additional aluminum foil layers were applied to the accumulated electrospun fibers of an existing aluminum foil collector, effectively reducing the incidence of charge buildup. Using this process, we fabricated an electrospun scaffold with a large pore (pore size >40 μm) while simultaneously controlling the thickness. We demonstrate that the large pore size triggered rapid infiltration (160 μm in 4 hours of cell culture) of individual endothelial progenitor cells (EPCs) and rapid cell colonization after seeding EPC spheroids. We confirmed that the 3D, but not two-dimensional, scaffold structures regulated tubular structure formation by the EPCs. Thus, incorporation of stem cells into a highly porous 3D scaffold with tunable thickness has implications for the regeneration of vascularized thick tissues and cardiac patch development. PMID:25709441

  8. Increased Endothelial Progenitor Cell Levels are Associated with Good Outcome in Intracerebral Hemorrhage.

    PubMed

    Pías-Peleteiro, Juan; Pérez-Mato, María; López-Arias, Esteban; Rodríguez-Yáñez, Manuel; Blanco, Miguel; Campos, Francisco; Castillo, José; Sobrino, Tomás

    2016-01-01

    Circulating endothelial progenitor cells (EPCs) play a role in the regeneration of damaged brain tissue. However, the relationship between circulating EPC levels and functional recovery in intracerebral hemorrhage (ICH) has not yet been tested. Therefore, our aim was to study the influence of circulating EPCs on the outcome of ICH. Forty-six patients with primary ICH (males, 71.7%; age, 72.7 ± 10.8 years) were prospectively included in the study within 12 hours of symptom onset. The main outcome variable was good functional outcome at 12 months (modified Rankin scale ≤2), considering residual volume at 6 months as a secondary variable. Circulating EPC (CD34(+)/CD133(+)/KDR(+)) levels were measured by flow cytometry from blood samples obtained at admission, 72 hours and day 7. Our results indicate that patients with good outcome show higher EPC numbers at 72 hours and day 7 (all p < 0.001). However, only EPC levels at day 7 were independently associated with good functional outcome at 12 months (OR, 1.15; CI95%, 1.01-1.35) after adjustment by age, baseline stroke severity and ICH volume. Moreover, EPC levels at day 7 were negatively correlated to residual volume (r = -0.525; p = 0.005). In conclusion, these findings suggest that EPCs may play a role in the functional recovery of ICH patients. PMID:27346699

  9. Patient-Derived Endothelial Progenitor Cells Improve Vascular Graft Patency in Rodent Model

    PubMed Central

    Stroncek, JD; Ren, LC; Klitzman, B; Reichert, WM

    2011-01-01

    Late outgrowth endothelial progenitor cells (EPCs) derived from the peripheral blood of patients with significant coronary artery disease were sodded into the lumens of small diameter expanded polytetrafluoroethylene (ePTFE) vascular grafts. Grafts (1 mm inner diameter) were denucleated and sodded either with native EPCs or with EPCs transfected with an adenoviral vector containing the gene for human thrombomodulin (EPC+AdTM). EPC+AdTM was shown to increase the in vitro rate of graft activated protein C (APC) production 4-fold over grafts sodded with untransfected EPCs (p<0.05). Unsodded control and EPC-sodded and EPC+AdTM-sodded grafts were implanted bilaterally into the femoral arteries of athymic rats for 7 or 28 days. Unsodded control grafts, both with and without denucleation treatment, each exhibited 7-day patency rates of 25%. Unsodded grafts showed extensive thrombosis and were not tested for patency over 28 days. In contrast, grafts sodded with untransfected EPCs or EPC+AdTM both had 7-day patency rates of 88-89% and 28-day patency rates of 75-88%. Intimal hyperplasia was observed near both the proximal and distal anastomoses in all sodded graft conditions but did not appear to be the primary occlusive failure event. This in vivo study suggests autologous EPCs derived from the peripheral blood of patients with coronary artery disease may improve the performance of synthetic vascular grafts, although no differences were observed between untransfected EPCs and TM transfected EPCs. PMID:21945828

  10. Endothelial progenitor cells promote tumor growth and progression by enhancing new vessel formation

    PubMed Central

    Zhao, Xin; Liu, Huan-Qiu; Li, Ji; Liu, Xiao-Liang

    2016-01-01

    Tumor growth and progression require new blood vessel formation to deliver nutrients and oxygen for further cell proliferation and to create a neovascular network exit for tumor cell metastasis. Endothelial progenitor cells (EPCs) are a bone marrow (BM)-derived stem cell population that circulates in the peripheral circulation and homes to the tumor bed to participate in new blood vessel formation. In addition to structural support to nascent vessels, these cells can also regulate the angiogenic process by paracrine secretion of a number of proangiogenic growth factors and cytokines, thus playing a crucial role in tumor neovascularization and development. Inhibition of EPC-mediated new vessel formation may be a promising therapeutic strategy in tumor treatment. EPC-mediated neovascularization is a complex process that includes multiple steps and requires a series of cytokines and modulators, thus understanding the underlying mechanisms may provide anti-neovasculogenesis targets that may be blocked for the prevention of tumor development. The present review stresses the process and contribution of EPCs to the formation of new blood vessels in solid tumors, in an attempt to gain an improved understanding of the underlying cellular and molecular mechanisms involved, and to provide a potential effective therapeutic target for cancer treatment. PMID:27446353

  11. Thickness-controllable electrospun fibers promote tubular structure formation by endothelial progenitor cells

    PubMed Central

    Hong, Jong Kyu; Bang, Ju Yup; Xu, Guan; Lee, Jun-Hee; Kim, Yeon-Ju; Lee, Ho-Jun; Kim, Han Seong; Kwon, Sang-Mo

    2015-01-01

    Controlling the thickness of an electrospun nanofibrous scaffold by altering its pore size has been shown to regulate cell behaviors such as cell infiltration into a three-dimensional (3D) scaffold. This is of great importance when manufacturing tissue-engineering scaffolds using an electrospinning process. In this study, we report the development of a novel process whereby additional aluminum foil layers were applied to the accumulated electrospun fibers of an existing aluminum foil collector, effectively reducing the incidence of charge buildup. Using this process, we fabricated an electrospun scaffold with a large pore (pore size >40 μm) while simultaneously controlling the thickness. We demonstrate that the large pore size triggered rapid infiltration (160 μm in 4 hours of cell culture) of individual endothelial progenitor cells (EPCs) and rapid cell colonization after seeding EPC spheroids. We confirmed that the 3D, but not two-dimensional, scaffold structures regulated tubular structure formation by the EPCs. Thus, incorporation of stem cells into a highly porous 3D scaffold with tunable thickness has implications for the regeneration of vascularized thick tissues and cardiac patch development. PMID:25709441

  12. Patient-derived endothelial progenitor cells improve vascular graft patency in a rodent model.

    PubMed

    Stroncek, J D; Ren, L C; Klitzman, B; Reichert, W M

    2012-01-01

    Late outgrowth endothelial progenitor cells (EPCs) derived from the peripheral blood of patients with significant coronary artery disease were sodded into the lumens of small diameter expanded polytetrafluoroethylene (ePTFE) vascular grafts. Grafts (1mm inner diameter) were denucleated and sodded either with native EPCs or with EPCs transfected with an adenoviral vector containing the gene for human thrombomodulin (EPC+AdTM). EPC+AdTM was shown to increase the in vitro rate of graft activated protein C (APC) production 4-fold over grafts sodded with untransfected EPCs (p<0.05). Unsodded control and EPC-sodded and EPC+AdTM-sodded grafts were implanted bilaterally into the femoral arteries of athymic rats for 7 or 28 days. Unsodded control grafts, both with and without denucleation treatment, each exhibited 7 day patency rates of 25%. Unsodded grafts showed extensive thrombosis and were not tested for patency over 28 days. In contrast, grafts sodded with untransfected EPCs or EPC+AdTM both had 7 day patency rates of 88-89% and 28 day patency rates of 75-88%. Intimal hyperplasia was observed near both the proximal and distal anastomoses in all sodded graft conditions but did not appear to be the primary occlusive failure event. This in vivo study suggests autologous EPCs derived from the peripheral blood of patients with coronary artery disease may improve the performance of synthetic vascular grafts, although no differences were observed between untransfected EPCs and TM transfected EPCs. PMID:21945828

  13. Increased Endothelial Progenitor Cell Levels are Associated with Good Outcome in Intracerebral Hemorrhage

    PubMed Central

    Pías-Peleteiro, Juan; Pérez-Mato, María; López-Arias, Esteban; Rodríguez-Yáñez, Manuel; Blanco, Miguel; Campos, Francisco; Castillo, José; Sobrino, Tomás

    2016-01-01

    Circulating endothelial progenitor cells (EPCs) play a role in the regeneration of damaged brain tissue. However, the relationship between circulating EPC levels and functional recovery in intracerebral hemorrhage (ICH) has not yet been tested. Therefore, our aim was to study the influence of circulating EPCs on the outcome of ICH. Forty-six patients with primary ICH (males, 71.7%; age, 72.7 ± 10.8 years) were prospectively included in the study within 12 hours of symptom onset. The main outcome variable was good functional outcome at 12 months (modified Rankin scale ≤2), considering residual volume at 6 months as a secondary variable. Circulating EPC (CD34+/CD133+/KDR+) levels were measured by flow cytometry from blood samples obtained at admission, 72 hours and day 7. Our results indicate that patients with good outcome show higher EPC numbers at 72 hours and day 7 (all p < 0.001). However, only EPC levels at day 7 were independently associated with good functional outcome at 12 months (OR, 1.15; CI95%, 1.01–1.35) after adjustment by age, baseline stroke severity and ICH volume. Moreover, EPC levels at day 7 were negatively correlated to residual volume (r = −0.525; p = 0.005). In conclusion, these findings suggest that EPCs may play a role in the functional recovery of ICH patients. PMID:27346699

  14. Effect of hypoxia on integrin-mediated adhesion of endothelial progenitor cells

    PubMed Central

    Kaiser, Ralf; Friedrich, Denise; Chavakis, Emmanouil; Böhm, Michael; Friedrich, Erik B

    2012-01-01

    Homing of endothelial progenitor cells (EPCs) is crucial for neoangiogenesis, which might be negatively affected by hypoxia. We investigated the influence of hypoxia on fibronectin binding integrins for migration and cell-matrix-adhesion. AMP-activated kinase (AMPK) and integrin-linked kinase (ILK) were examined as possible effectors of hypoxia.Human EPCs were expanded on fibronectin (FN) and integrin expression was profiled by flow cytometry. Cell-matrix-adhesion- and migration-assays on FN were performed to examine the influence of hypoxia and AMPK-activation. Regulation of AMPK and ILK was shown by Western blot analysis. We demonstrate the presence of integrin β1, β2 and α5 on EPCs. Adhesion to FN is reduced by blocking β1 and α5 (49% and 2% of control, P < 0.05) whereas α4-blockade has no effect. Corresponding effects were shown for migration. Hypoxia and AMPK-activation decrease adhesion on FN. Although total AMPK-expression remains unchanged, phospho-AMPK increases eightfold.The EPCs require α5 for adhesion on FN. Hypoxia and AMPK-activation decrease adhesion. As α5 is the major adhesive factor for EPCs on FN, this suggests a link between AMPK and α5-integrins. We found novel evidence for a connection between hypoxia, AMPK-activity and integrin activity. This might affect the fate of EPCs in ischaemic tissue. PMID:22353471

  15. Endothelial Progenitor Cells Correlate with Clinical Outcome of Traumatic Brain Injury

    PubMed Central

    Liu, Li; Wei, Huijie; Chen, Fanglian; Wang, Jinghua; Dong, Jing-fei; Zhang, Jianning

    2012-01-01

    Objective Endothelial progenitor cells (EPCs) play an active role in vascular repair and revascularization of tissue damaged by traumatic, inflammatory, and ischemic injures. We correlate the changes in circulating EPCs with the severity of traumatic brain injury (TBI). The study is designed to investigate the EPC mobilization after injury and a potential use of circulating EPCs as a prognostic marker for evaluating trauma severity and clinical outcomes. Design A prospective cohort study conducted in two neurosurgical intensive care units (NSICU) of Tianjin Medical University General Hospital and Tianjin Huanhu Hospital. Patients Patients with traumatic brain injury and age- and gender-matched healthy controls. Interventions None Measurements and Main Results Changes in the levels of circulating EPCs were monitored for up to 21 days in 84 patients with TBI. Results were correlated with the clinical assessment of injury severity as determined by Glasgow Coma Scale (GCS). The level of circulating EPCs was found to be suppressed 24–48 hrs after injury, but rapidly increased, reaching the highest at day 5–7 posttrauma. Circulating EPCs in patients with improved GCS were significantly higher than those with deteriorated conditions, and remained persistently low in patients who died of trauma. Conclusions The results suggest that the level of circulating EPCs correlates with the clinical severity and outcome of TBI, and may offer potentials as a prognostic marker for TBI. A long-term follow up of these patients is ongoing. PMID:21460712

  16. Ambient Fine Particulate Matter Induces Apoptosis of Endothelial Progenitor Cells Through Reactive Oxygen Species Formation

    PubMed Central

    Cui, Yuqi; Xie, Xiaoyun; Jia, Fengpeng; He, Jianfeng; Li, Zhihong; Fu, Minghuan; Hao, Hong; Liu, Ying; Liu, Jason Z.; Cowan, Peter J.; Zhu, Hua; Sun, Qinghua; Liu, Zhenguo

    2015-01-01

    Background/Aims Bone marrow (BM)-derived endothelial progenitor cells (EPCs) play a critical role in angiogenesis and vascular repair. Some environmental insults, like fine particulate matter (PM) exposure, significantly impair cardiovascular functions. However, the mechanisms for PM-induced adverse effects on cardiovascular system remain largely unknown. The present research was to study the detrimental effects of PM on EPCs and explore the potential mechanisms. Methods PM was intranasal-distilled into male C57BL/6 mice for one month. Flow cytometry was used to measure the number of EPCs, apoptosis level of circulating EPCs and intracellular reactive oxygen species (ROS) formation. Serum TNF-α and IL-1β were measured using ELISA. To determine the role of PM-induced ROS in EPC apoptosis, PM was co-administrated with the antioxidant N-acetylcysteine (NAC) in wild type mice or used in a triple transgenic mouse line (TG) with overexpression of antioxidant enzyme network (AON) composed of superoxide dismutase (SOD)1, SOD3, and glutathione peroxidase (Gpx-1) with decreased in vivo ROS production. Results PM treatment significantly decreased circulating EPC population, promoted apoptosis of EPCs in association with increased ROS production and serum TNF-α and IL-1β levels, which could be effectively reversed by either NAC treatment or overexpression of AON. Conclusion PM exposure significantly decreased circulating EPCs population due to increased apoptosis via ROS formation in mice. PMID:25591776

  17. TNFα Regulates Endothelial Progenitor Cell Migration via CADM1 and NF-kB

    PubMed Central

    Prisco, Anthony R.; Hoffmann, Brian R.; Kaczorowski, Catherine C.; McDermott-Roe, Chris; Stodola, Timothy J.; Exner, Eric C.; Greene, Andrew S.

    2016-01-01

    Shortly after the discovery of endothelial progenitor cells (EPCs) in 1997, many clinical trials were conducted using EPCs as a cellular based therapy with the goal of restoring damaged organ function by inducing growth of new blood vessels (angiogenesis). Results were disappointing, largely because the cellular and molecular mechanisms of EPC-induced angiogenesis were not clearly understood. Following injection, EPCs must migrate to the target tissue and engraft prior to induction of angiogenesis. In this study EPC migration was investigated in response to tumor necrosis factor α (TNFα), a pro-inflammatory cytokine, to test the hypothesis that organ damage observed in ischemic diseases induces an inflammatory signal that is important for EPC homing. In this study, EPC migration and incorporation were modeled in vitro using a co-culture assay where TNFα treated EPCs were tracked while migrating towards vessel-like structures. It was found that TNFα treatment of EPCs increased migration and incorporation into vessel-like structures. Using a combination of genomic and proteomic approaches, NF-kB mediated upregulation of CADM1 was identified as a mechanism of TNFα induced migration. Inhibition of NF-kB or CADM1 significantly decreased migration of EPCs in vitro suggesting a role for TNFα signaling in EPC homing during tissue repair. PMID:26867147

  18. Knockdown of stromal interaction molecule 1 attenuates hepatocyte growth factor-induced endothelial progenitor cell proliferation.

    PubMed

    Shi, Yankun; Song, Mingbao; Guo, Ruiwei; Wang, Hong; Gao, Pan; Shi, Weibin; Huang, Lan

    2010-03-01

    Increased Ca(2+) entry through store-operated Ca(2+) channels (SOCCs) plays an essential role in the regulation of hepatocyte growth factor (HGF)-induced cell proliferation. Stromal interaction molecule 1 (STIM1) is thought to transmit endoplasmic reticulum (ER) Ca(2+) store depletion signals to the plasma membrane (PM), causing the opening of SOCCs in the PM. However, the relationship between HGF and STIM1 in endothelial progenitor cell (EPC) proliferation remains uncharacterized. The objective of this study was to evaluate the potential involvement of STIM1 in HGF-induced EPC proliferation. For this purpose, we used cultured rat bone marrow-derived EPCs and found that HGF-induced EPC proliferation at low concentrations. Store-operated Ca(2+) entry (SOCE) was elevated in HGF-treated EPCs, and the SOCC inhibitors 2-aminoethoxydiphenyl borate (2-APB) and BTP-2 inhibited the HGF-induced proliferation response. Moreover, STIM1 mRNA and protein expression levels were increased in response to HGF stimulation and knockdown of STMI1 decreased SOCE and prevented HGF-induced EPC proliferation. In conclusion, our data suggest that HGF-induced EPC proliferation is mediated partly via activation of STIM1. PMID:20404049

  19. Circulating endothelial progenitor cells in women with gestational alterations of glucose tolerance.

    PubMed

    Penno, Giuseppe; Pucci, Laura; Lucchesi, Daniela; Lencioni, Cristina; Iorio, Maria Carla; Vanacore, Renato; Storti, Eugenia; Resi, Veronica; Di Cianni, Graziano; Del Prato, Stefano

    2011-07-01

    Endothelial progenitor cells (EPCs) play a role in angiogenesis during pregnancy. The aim of this study was to evaluate circulating EPCs in pregnant women with gestational alterations of glucose tolerance. Glucose tolerance, insulin sensitivity and β-cell function were derived from oral glucose tolerance tests in 23 women with normal glucose tolerance (NGT), 18 with gestational impaired glucose tolerance (GIGT) and 24 with gestational diabetes mellitus (GDM). Circulating cells expressing CD34 in combination with CD133, kinase insert domain receptor (KDR) or both were quantified by flow cytometry. Women with GIGT and GDM had lower CD34(+)KDR(+) and CD34(+)CD133( +)KDR(+) cells at 27±3.2 weeks' gestation compared with NGT (ANOVA p<0.02 for both). CD34(+)KDR(+) and CD34(+)CD133(+)KDR(+) cells were inversely correlated with the area-under-the-glucose-curve (p<0.005, for both) and positively to insulin secretion-sensitivity index (p<0.05, for both). Alterations of glucose tolerance during pregnancy are associated with a decrease in EPCs. Hyperglycaemia might exert a direct effect on depletion of EPCs. PMID:21653675

  20. Macrophages commit postnatal endothelium-derived progenitors to angiogenesis and restrict endothelial to mesenchymal transition during muscle regeneration

    PubMed Central

    Zordan, P; Rigamonti, E; Freudenberg, K; Conti, V; Azzoni, E; Rovere-Querini, P; Brunelli, S

    2014-01-01

    The damage of the skeletal muscle prompts a complex and coordinated response that involves the interactions of many different cell populations and promotes inflammation, vascular remodeling and finally muscle regeneration. Muscle disorders exist in which the irreversible loss of tissue integrity and function is linked to defective neo-angiogenesis with persistence of tissue necrosis and inflammation. Here we show that macrophages (MPs) are necessary for efficient vascular remodeling in the injured muscle. In particular, MPs sustain the differentiation of endothelial-derived progenitors to contribute to neo-capillary formation, by secreting pro-angiogenic growth factors. When phagocyte infiltration is compromised endothelial-derived progenitors undergo a significant endothelial to mesenchymal transition (EndoMT), possibly triggered by the activation of transforming growth factor-β/bone morphogenetic protein signaling, collagen accumulates and the muscle is replaced by fibrotic tissue. Our findings provide new insights in EndoMT in the adult skeletal muscle, and suggest that endothelial cells in the skeletal muscle may represent a new target for therapeutic intervention in fibrotic diseases. PMID:24481445

  1. Effective Mobilization of Very Small Embryonic-Like Stem Cells and Hematopoietic Stem/Progenitor Cells but Not Endothelial Progenitor Cells by Follicle-Stimulating Hormone Therapy

    PubMed Central

    Zbucka-Kretowska, Monika; Eljaszewicz, Andrzej; Lipinska, Danuta; Grubczak, Kamil; Rusak, Malgorzata; Mrugacz, Grzegorz; Dabrowska, Milena; Ratajczak, Mariusz Z.; Moniuszko, Marcin

    2016-01-01

    Recently, murine hematopoietic progenitor stem cells (HSCs) and very small embryonic-like stem cells (VSELs) were demonstrated to express receptors for sex hormones including follicle-stimulating hormone (FSH). This raised the question of whether FSH therapy at clinically applied doses can mobilize stem/progenitor cells in humans. Here we assessed frequencies of VSELs (referred to as Lin−CD235a−CD45−CD133+ cells), HSPCs (referred to as Lin−CD235a−CD45+CD133+ cells), and endothelial progenitor cells (EPCs, identified as CD34+CD144+, CD34+CD133+, and CD34+CD309+CD133+ cells) in fifteen female patients subjected to the FSH therapy. We demonstrated that FSH therapy resulted in statistically significant enhancement in peripheral blood (PB) number of both VSELs and HSPCs. In contrast, the pattern of responses of EPCs delineated by different cell phenotypes was not uniform and we did not observe any significant changes in EPC numbers following hormone therapy. Our data indicate that FSH therapy mobilizes VSELs and HSPCs into peripheral blood that on one hand supports their developmental origin from germ lineage, and on the other hand FSH can become a promising candidate tool for mobilizing HSCs and stem cells with VSEL phenotype in clinical settings. PMID:26635885

  2. Effective Mobilization of Very Small Embryonic-Like Stem Cells and Hematopoietic Stem/Progenitor Cells but Not Endothelial Progenitor Cells by Follicle-Stimulating Hormone Therapy.

    PubMed

    Zbucka-Kretowska, Monika; Eljaszewicz, Andrzej; Lipinska, Danuta; Grubczak, Kamil; Rusak, Malgorzata; Mrugacz, Grzegorz; Dabrowska, Milena; Ratajczak, Mariusz Z; Moniuszko, Marcin

    2016-01-01

    Recently, murine hematopoietic progenitor stem cells (HSCs) and very small embryonic-like stem cells (VSELs) were demonstrated to express receptors for sex hormones including follicle-stimulating hormone (FSH). This raised the question of whether FSH therapy at clinically applied doses can mobilize stem/progenitor cells in humans. Here we assessed frequencies of VSELs (referred to as Lin(-)CD235a(-)CD45(-)CD133(+) cells), HSPCs (referred to as Lin(-)CD235a(-)CD45(+)CD133(+) cells), and endothelial progenitor cells (EPCs, identified as CD34(+)CD144(+), CD34(+)CD133(+), and CD34(+)CD309(+)CD133(+) cells) in fifteen female patients subjected to the FSH therapy. We demonstrated that FSH therapy resulted in statistically significant enhancement in peripheral blood (PB) number of both VSELs and HSPCs. In contrast, the pattern of responses of EPCs delineated by different cell phenotypes was not uniform and we did not observe any significant changes in EPC numbers following hormone therapy. Our data indicate that FSH therapy mobilizes VSELs and HSPCs into peripheral blood that on one hand supports their developmental origin from germ lineage, and on the other hand FSH can become a promising candidate tool for mobilizing HSCs and stem cells with VSEL phenotype in clinical settings. PMID:26635885

  3. Association of endothelial progenitor cells and peptic ulcer treatment in patients with type 2 diabetes mellitus

    PubMed Central

    NIE, ZHIHONG; XU, LIMIN; LI, CHUANYUAN; TIAN, TAO; XIE, PINGPING; CHEN, XIA; LI, BOJING

    2016-01-01

    The present study aimed to investigate the association between endothelial progenitor cells (EPCs) and peptic ulcers in patients with or without type 2 diabetes mellitus (T2DM), in association with the efficiency of peptic ulcer treatment. The study recruited healthy subjects and peptic ulcer patients with or without T2DM. All the ulcer patients, including those with and without T2DM, were administered omeprazole for 8 weeks. Peptic ulcer patients with T2DM were additionally treated with glipizide and novolin. Blood samples were then obtained from the three groups following ulcer treatment. CD133+ cells were isolated from the blood samples using magnetic bead selection, and cultured in complete medium 199. Morphological and quantity changes in EPCs were observed by light and fluorescence microscopy. In addition, flow cytometric analysis was used to quantify the number of vascular endothelial cells. The treatment was partially effective in 7 of the 32 peptic ulcer patients without T2DM and 12 of the 32 peptic ulcer patients with T2DM. However, this treatment was ineffective in 20 of the 32 peptic ulcer patients with T2DM. Notably, 25 peptic ulcer patients without T2DM were defined as completely recovered following treatment. In addition, the number of circulating EPCs as well as their colony forming ability was significantly reduced (P<0.05) in the peptic ulcer patients with T2DM following ulcer treatment, compared with the other groups. Circulating EPC counts were significantly increased in peptic ulcer patients without T2DM, as compared with the healthy controls. With regards to colony formation, peptic ulcer patients without T2DM did not exhibit improved colony formation ability. In conclusion, the number of circulating EPCs and their colony-forming ability was significantly reduced in peptic ulcer patients with T2DM following ulcer treatment when compared with the other groups. This suggests that the poor curative effect of peptic ulcer treatment in these

  4. Circulating endothelial progenitor cells in type 1 diabetic patients with erectile dysfunction.

    PubMed

    Maiorino, Maria Ida; Bellastella, Giuseppe; Petrizzo, Michela; Della Volpe, Elisabetta; Orlando, Rosanna; Giugliano, Dario; Esposito, Katherine

    2015-06-01

    Circulating endothelial progenitor cells (EPCs) are bone marrow-derived stem cells able to migrate to sites of damaged endothelium and differentiate into endothelial cells, thereby contributing to vascular repair. Recent studies demonstrated a reduction of EPCs in patients with diabetes mellitus or erectile dysfunction (ED). The aim of this study was to evaluate the circulating levels of different EPCs phenotypes and their relation with testosterone levels in young type 1 diabetic patients with ED. We studied 118 consecutively type 1 diabetic patients and 60 age-matched healthy controls. Erectile function was assessed by completing the International Index of Erectile Function (IIEF-5) and EPCs levels by flow cytometry. Testosterone concentrations were evaluated in all the study population. We identified 38 diabetic patients with ED (Group 1) and 80 patients without ED (Group 2). CD34+KDR+CD133+ cells were significantly lower in patients in Group 1 as compared with those in Group 2 [median and interquartile range, n/10(6) events, 12 (6-16) vs. 18 (13-22), P < 0.001)]. In all participants in the study, there was a significant correlation between circulating CD34+KDR+CD133+ cells and testosterone levels (r = 0.410, P < 0.001), which was highest in Group 1, intermediate in Group 2, and lowest in Group 3 (controls). There was a significant correlation between IIEF-5 score and both CD34+KDR+ (r = 0.459, P = 0.003) and CD34+KDR+CD133+ (r = 0.316, P = 0.050) cells among patients of Group 1, as well as between testosterone levels and most of the EPCs phenotypes. Finally, multivariate regression analysis identified levels of circulating CD34+KDR+ cells as an independent risk factor for ED (β-coefficient 0.348, P = 0.007). In conclusion, type 1 diabetic patients with ED show reduced levels of CD34+KDR+CD133+ cells, whose number correlates with IIEF. Further studies are needed to fully understand the exact mechanisms by which testosterone regulates vascular homeostasis. PMID

  5. Circulating Endothelial Progenitor Cells in Castration Resistant Prostate Cancer: A Randomized, Controlled, Biomarker Study

    PubMed Central

    Fuereder, Thorsten; Wacheck, Volker; Strommer, Sabine; Horak, Peter; Gerschpacher, Marion; Lamm, Wolfgang; Kivaranovic, Danijel; Krainer, Michael

    2014-01-01

    Background Endothelial progenitor cells (CEPs) and circulating endothelial cells (CECs) are potential biomarkers of response to anti-angiogenic treatment regimens. In the current study, we investigated the effect of docetaxel and sunitinib on CEP/CEC kinetics and clinical response in castration resistant prostate cancer (CRPC) patients. Patients and methods Chemonaive patients with CRPC were enrolled in this study to receive either sunitinib (37.5 mg/d), in combination with docetaxel (75 mg/m2) or docetaxel alone. CEP and CEC kinetics were analyzed for every cycle. The primary objective was to compare CEP/CEC pharmacodynamics between both treatment arms. We also investigated if CEC/CEP spikes, induced by MTD docetaxel, are suppressed by sunitinib in patients treated with docetaxel/sunitinib relative to docetaxel monotherapy. Results A total of 27 patients were enrolled. We observed a significant increase of CEP/CEC (total/viable) counts over time within each cycle (coefficients 0.29233, 0.22092 and 0.26089, respectively; p<0.001). However, no differences between the treatment groups, in terms of CEP and CEC kinetics, were detected. In the docetaxel monotherapy arm 4 (30%) patients responded to therapy with a 50% PSA decline, while 9 (64%) patients showed a PSA decline in the combination group (n.s.). The median PFS in the docetaxel monotherapy group was 3.1 months (2.6–3.6 months, 95% CI) and 6.2 months (4.9–7.4 months, 95% CI; p = 0.062) in the combination arm. Sunitinib/docetaxel was reasonably well tolerated and toxicity manageable. Conclusion In summary, no significant differences in CEC and CEP kinetics between the treatment arms were observed, although a highly significant increase of CEPs/CECs within each cycle over time was detected. These results mirror the challenge we have to face when employing anti-angiogenic strategies in CRPC. Additional preclinical research is needed to elucidate the underlying molecular mechanisms. However, docetaxel

  6. Endothelial Progenitor Cells Are Related to Glycemic Control in Children With Type 1 Diabetes Over Time

    PubMed Central

    Hörtenhuber, Thomas; Rami-Mehar, Birgit; Satler, Miriam; Nagl, Katrin; Höbaus, Clemens; Höllerl, Florian; Koppensteiner, Renate; Schernthaner, Guntram; Schober, Edith; Schernthaner, Gerit-Holger

    2013-01-01

    OBJECTIVE The risk of cardiovascular death before the age of 40 is 20-fold higher in patients with type 1 diabetes mellitus (T1DM). Endothelial progenitor cells (EPCs) predict cardiovascular morbidity and mortality in patients without diabetes. We hypothesized that EPCs are modified in children with T1DM and are related to characteristics of T1DM such as glycemic control. RESEARCH DESIGN AND METHODS Children (n = 190; 156 T1DM subjects and 34 control subjects) were included in an observational cohort study and matched for age and sex. EPCs were enumerated by flow cytometry at the beginning (cross-sectional) and 1 year later (longitudinal). To analyze changes of variables during the observation, Δ values were calculated. RESULTS EPCs were significantly reduced in T1DM children versus control subjects (609 ± 359 vs. 1,165 ± 484, P < 0.001). Multivariate regression modeling revealed that glycated hemoglobin A1c (HbA1c) was the strongest independent predictor of EPCs (β = −0.355, P < 0.001). Overall glycemic control at the beginning and end of study did not differ (7.8 ± 1.2 vs. 7.8 ± 1.2 relative %, P = NS), but we observed individual HbA1c changes of −4.30/+3.10 relative %. The strongest EPC increase was observed in the patients with the most favorable HbA1c lowering during the 1-year follow-up. Accordingly, the strongest EPC decrease was demonstrated in the patients with the strongest HbA1c worsening during the time period. CONCLUSIONS This is the first prospective study demonstrating diminished EPCs in children with T1DM. The association of better glycemic control with an increase in EPC numbers within 1 year suggests that a reduction of the high cardiovascular disease burden might be mediated likewise. PMID:23340890

  7. Relation between endothelial progenitor cells and arterial stiffness in patients with psoriasis.

    PubMed

    Liu, Ju-Hua; Chen, Yan; Zhen, Zhe; Yeung, Chi-Keung; Chan, Johnny; Chan, Henry H; Tse, Hung-Fat; Yiu, Kai-Hang

    2016-08-01

    Patients with psoriasis are prone to premature atherosclerosis. We hypothesize that depletion of circulating endothelial progenitor cells (EPC) is related to patients with psoriasis and can contribute to the development of atherosclerosis. Thirty-five plaque-type psoriasis patients (41.9 ± 5.5 years, 30 men) and 20 age- and sex-matched controls were studied. Four subpopulations of EPC, namely, CD34(+) EPC, CD133(+) EPC, CD34(+) /kinase insert domain-containing receptor (KDR)(+) EPC and CD133(+) /KDR(+) EPC were measured by flow cytometry. Arterial stiffness in psoriasis patients was assessed by heart to ankle pulse wave velocity (haPWV), augmentation index (AI) and carotid intima media thickness (IMT). Patients with psoriasis had a lower level of CD34(+) EPC (7.85 ± 2.49% vs 6.26 ± 2.13%, P = 0.02) compared with healthy controls. In patients with psoriasis, level of CD34(+) EPC was negatively related with haPWV (r = -0.43 P = 0.01) and Psoriasis Area and Severity Index (r = -0.39 P = 0.02). Multivariate regression analysis further demonstrated that haPWV was independently associated with level of CD34(+) EPC. Each percentage decrease in CD34(+) EPC accounted for an increase in haPWV of +0.02 m/s. The result demonstrated that patients with psoriasis had reduced CD34(+) EPC compared with controls. Importantly, CD34(+) EPC was independently related with haPWV in these patients. This finding suggests that EPC reduction is associated with the development of arterial stiffness in patients with psoriasis. PMID:26704131

  8. Autologous endothelial progenitor cells improve allograft survival in porcine lung transplantation with prolonged ischemia

    PubMed Central

    Yen, Yi-Ting; Roan, Jun-Neng; Fang, Shih-Yuan; Chang, Shi-Wei; Tseng, Yau-Lin

    2016-01-01

    Background As endothelial progenitor cells (EPCs) attenuated acute lung injury (ALI) in rabbit model, we hypothesized that autologous EPCs preserved lung graft function during the acute reperfusion period of lung transplantation and tested the therapeutic potential of EPCs in a porcine model of lung transplantation with prolonged graft ischemia. Methods Day-7 EPCs isolated from the recipient subjects or plain culture media were administered into the left pulmonary artery immediately before restoration of pulmonary blood flow in a porcine lung allotransplantation model, with the transplantation surgeons blinded to the content of injection. Hemodynamics and arterial blood gas were recorded, and the right pulmonary artery was occluded 30 min after reperfusion to evaluate the lung graft function. The lung grafts were sectioned for histological examination at the end of experiments. The total ischemic time for lung graft was approximately 14 h. Results All animals receiving plain medium died within 40 min after reperfusion, but 3 out of 5 (60%) piglets receiving EPCs survived up to 4 h after diversion of the entire cardiac output into the lung graft (P<0.01). The donor body weight, recipient body weight, cold ischemic time, and time for anastomosis were comparable between the EPC and control group (P=0.989, 0.822, 0.843, and 0.452, respectively). The mean aortic pressure decreased, and the cardiac output and mean pulmonary artery pressure elevated after right pulmonary artery occlusion. All these parameters were gradually compensated in the EPC group but decompensated in the control group. Better preservation of gas exchange function, reduced thrombi formation in the terminal pulmonary arterioles, and attenuated interstitial hemorrhage of the lung graft were observed in the EPC group. Conclusions We concluded autologous EPCs significantly enhanced the function of lung allograft and improved survival in a porcine model of lung transplantation with prolonged ischemia

  9. Endothelial progenitors encapsulated in bioartificial niches are insulated from systemic cytotoxicity and are angiogenesis competent.

    PubMed

    Ratliff, B B; Ghaly, T; Brudnicki, P; Yasuda, K; Rajdev, M; Bank, M; Mares, J; Hatzopoulos, A K; Goligorsky, M S

    2010-07-01

    Intrinsic stem cells (SC) participate in tissue remodeling and regeneration in various diseases and following toxic insults. Failure of tissue regeneration is in part attributed to lack of SC protection from toxic stress of noxious stimuli, thus prompting intense research efforts to develop strategies for SC protection and functional preservation for in vivo delivery. One strategy is creation of artificial SC niches in an attempt to mimic the requirements of endogenous SC niches by generating scaffolds with properties of extracellular matrix. Here, we investigated the use of hyaluronic acid (HA) hydrogels as an artificial SC niche and examined regenerative capabilities of encapsulated embryonic endothelial progenitor cells (eEPC) in three different in vivo models. Hydrogel-encapsulated eEPC demonstrated improved resistance to toxic insult (adriamycin) in vitro, thus prompting in vivo studies. Implantation of HA hydrogels containing eEPC to mice with adriamycin nephropathy or renal ischemia resulted in eEPC mobilization to injured kidneys (and to a lesser extent to the spleen) and improvement of renal function, which was equal or superior to adoptively transferred EPC by intravenous infusion. In mice with hindlimb ischemia, EPC encapsulated in HA hydrogels dramatically accelerated the recovery of collateral circulation with the efficacy superior to intravenous infusion of EPC. In conclusion, HA hydrogels protect eEPC against adriamycin cytotoxicity and implantation of eEPC encapsulated in HA hydrogels supports renal regeneration in ischemic and cytotoxic (adriamycin) nephropathy and neovascularization of ischemic hindlimb, thus establishing their functional competence and superior capabilities to deliver stem cells stored in and released from this bioartificial niche. PMID:20410213

  10. Nicotine improves the functional activity of late endothelial progenitor cells via nicotinic acetylcholine receptors.

    PubMed

    Yu, Min; Liu, Qian; Sun, Jing; Yi, Kaihong; Wu, Libiao; Tan, Xuerui

    2011-08-01

    The aim of this study is to investigate whether nicotinic acetylcholine receptors (nAChRs) are involved in the modulation of functional activity of late endothelial progenitor cells (EPCs) induced by nicotine. Total mononuclear cells (MNCs) were isolated from human umbilical cord blood by Ficoll density gradient centrifugation, and then the cells were plated on fibronectin-coated culture plates. Late EPCs were positive for 1,1-dioctadecyl-3,3,3,3-tetramethylindocarbocyanine-labeled acetylated low-density lipoprotein (DiI-acLDL) uptake and fluorescein-isothiocyanate-conjugated Ulex europaeus agglutinin lectin (UEA-1) binding. Expression of von Willbrand factor (vWF), kinase insert domain receptor (KDR), and α7 nAChR was detected by indirect immunofluorescence staining. Late EPCs of 3-5 passages were treated for 32 h with either vehicle or nicotine with or without pre-incubation of nAChR antagonism, mecamylamine, or α-bungarotoxin. The viability, migration, and in vitro vasculogenesis activity of late EPCs were assayed with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, modified Boyden chamber assay, and in vitro angiogenesis assay, respectively. Late EPCs adhesion assay was performed by replating cells on fibronectin-coated plates, and then adherent cells were counted. Incubation with 10 nmol/L nicotine enhanced viable, migratory, adhesive, and in vitro vasculogenesis capacity of late EPCs. The effect of nicotine on late EPCs can be attenuated by mecamylamine or α-bungarotoxin. In conclusion, nicotine improves the functional activity of late EPCs via nAChRs. PMID:21774635

  11. ENDOTHELIAL PROGENITOR CELL ADHESION, GROWTH AND CHARACTERIZATION ON TRABECULAR TITANIUM AND TRABECULAR TITANIUM COATED WITH COLLAGEN OR DECELLULARIZED ECM.

    PubMed

    Gastaldi, G; Caliogna, L; Botta, L; Ghiara, M; Benazzo, F

    2015-01-01

    Adequate blood supply is essential for prosthesis osteointegration and bone healing as it supplies oxygen, nutrition and progenitor cells. The bone healing process and vascularization depend upon the endothelial cells, which speed up implant osteointegration. Endothelial Progenitor Cells (EPC) are a population of stem cells that can reproduce, migrate and acquire mature endothelial phenotype. Their recruitment occurs in the tissue lesion to enhance neovascularization. Trabecular TitaniumTM (TTTM) is a new biomaterial with very interesting biomechanical characteristics and fast osteointegration. This study has investigated adhesion, proliferation and characteristics of EPC on three types of biomaterial: unmodified trabecular titanium, trabecular titanium coated with the ECM deposited by human mesenchymal stem cells isolated from subcutaneous adipose tissue and decellularized and trabecular titanium coated with type I collagen (control scaffold). MTT assay showed similar percentages of EPCs seeded on the different kinds of scaffold: 67% on TT, 70% on decellularized scaffolds and 82% on collagen-coated scaffolds. There were no statistically significant differences between the three groups. We therefore conclude that TTTM allows EPC adhesion and proliferation and, consequently, by permitting vascularization, it favours prosthesis osteointegration. PMID:26652487

  12. Epigenetic Changes in Endothelial Progenitors as a Possible Cellular Basis for Glycemic Memory in Diabetic Vascular Complications

    PubMed Central

    Rajasekar, Poojitha; O'Neill, Christina L.; Eeles, Lydia; Stitt, Alan W.; Medina, Reinhold J.

    2015-01-01

    The vascular complications of diabetes significantly impact the quality of life and mortality in diabetic patients. Extensive evidence from various human clinical trials has clearly established that a period of poor glycemic control early in the disease process carries negative consequences, such as an increase in the development and progression of vascular complications that becomes evident many years later. Importantly, intensive glycemic control established later in the disease process cannot reverse or slow down the onset or progression of diabetic vasculopathy. This has been named the glycemic memory phenomenon. Scientists have successfully modelled glycemic memory using various in vitro and in vivo systems. This review emphasizes that oxidative stress and accumulation of advanced glycation end products are key factors driving glycemic memory in endothelial cells. Furthermore, various epigenetic marks have been proposed to closely associate with vascular glycemic memory. In addition, we comment on the importance of endothelial progenitors and their role as endogenous vasoreparative cells that are negatively impacted by the diabetic milieu and may constitute a “carrier” of glycemic memory. Considering the potential of endothelial progenitor-based cytotherapies, future studies on their glycemic memory are warranted to develop epigenetics-based therapeutics targeting diabetic vascular complications. PMID:26106624

  13. Insulin-Like Growth Factor Binding Protein-2 Promotes Adhesion of Endothelial Progenitor Cells to Endothelial Cells via Integrin α5β1.

    PubMed

    Feng, Nianping; Zhang, Zhuo; Wang, Zhengfei; Zheng, Haihong; Qu, Fujun; He, Xijun; Wang, Chunlai

    2015-11-01

    The contribution of endothelial progenitor cells (EPCs) to new vessel formation has been studied in different physiological and pathological conditions for decades. As previously suggested, insulin-like growth factor binding protein-2 (IGFBP-2) may interact with integrins and promote cell migration. However, the role of IGFBP-2 in regulation of EPC functions remains largely unknown. In this present study, we found that overexpression of IGFBP-2 in human umbilical vein endothelial cells (HUVECs) promoted EPC-endothelial adhesion. Conversely, siRNA-mediated depletion of IGFBP-2 inhibited oxygen-glucose deprivation (OGD)-induced EPC-endothelial adhesion. Further, we demonstrated that the arginine-glycine-aspartic acid (RGD) motif in its C-domain is required for interaction with integrin α5β1. In addition, treatment with IGFBP-2 significantly enhanced incorporation of EPCs into tubule networks formed by HUVECs. Thus, our findings suggest that exogenous administration of IGFBP-2 may facilitate neovascularization and improve treatment of ischemic conditions. PMID:26076738

  14. Influence of Rho kinase inhibitor Fasudil on late endothelial progenitor cells in peripheral blood of COPD patients with pulmonary artery hypertension.

    PubMed

    Liu, Pei; Zhang, Hongmei; Tang, Yijun; Sheng, Chunfeng; Liu, Jianxin; Zeng, Yanjun

    2014-02-01

    The objective of our work was to investigate the influence of Fasudil, a Rho inhibitor on the number and function of the late endothelial progenitor cells in peripheral blood of chronic obstructive pulmonary diseases (COPD) patients with pulmonary artery hypertension. Eighty COPD patients with pulmonary artery hypertension were selected and divided into two groups: the treatment group and the control group, which had 40 patients respectively. The control group received routine treatment, including oxygen uptake, anti-infection and phlegm dissolving. The treatment group received the Fasudil in addition to the routine treatment. The changes on the number and function of the late endothelial progenitor cells in peripheral blood of the patients before and after the treatment were compared between the two groups. The changes on the pulmonary artery pressure were also compared. The number of the late endothelial progenitor cells in peripheral blood of the treatment group increased and the function was enhanced. The pulmonary artery pressure was reduced. The difference before and after the treatment and with the control group was statistically significant (p<0.05). The changes on the number and function of the late endothelial progenitor cells in peripheral blood and the pulmonary artery pressure before and after the treatment of the control group were not statistically significant (p>0.05). The Rho-kinase inhibitor Fasudil increased the number and enhanced the function of the late endothelial progenitor cells in peripheral blood of COPD patients with pulmonary artery hypertension. PMID:24579970

  15. Vascular Endothelial Growth Factor (VEGF) Bioavailability Regulates Angiogenesis and Intestinal Stem and Progenitor Cell Proliferation during Postnatal Small Intestinal Development

    PubMed Central

    Holoyda, Kathleen A.; Hou, Xiaogang; Fowler, Kathryn L.; Grikscheit, Tracy C.

    2016-01-01

    Background Vascular endothelial growth factor (VEGF) is a highly conserved, master regulatory molecule required for endothelial cell proliferation, organization, migration and branching morphogenesis. Podocoryne carnea and drosophila, which lack endothelial cells and a vascular system, express VEGF homologs, indicating potential roles beyond angiogenesis and vasculogenesis. The role of VEGF in the development and homeostasis of the postnatal small intestine is unknown. We hypothesized regulating VEGF bioavailability in the postnatal small intestine would exhibit effects beyond the vasculature and influence epithelial cell stem/progenitor populations. Methods VEGF mutant mice were created that overexpressed VEGF in the brush border of epithelium via the villin promotor following doxycycline treatment. To decrease VEGF bioavailability, sFlt-1 mutant mice were generated that overexpressed the soluble VEGF receptor sFlt-1 upon doxycycline administration in the intestinal epithelium. Mice were analyzed after 21 days of doxycycline administration. Results Increased VEGF expression was confirmed by RT-qPCR and ELISA in the intestine of the VEGF mutants compared to littermates. The VEGF mutant duodenum demonstrated increased angiogenesis and vascular leak as compared to littermate controls. The VEGF mutant duodenum revealed taller villi and increased Ki-67-positive cells in the transit-amplifying zone with reduced Lgr5 expression. The duodenum of sFlt-1 mutants revealed shorter villi and longer crypts with reduced proliferation in the transit-amplifying zone, reduced expression of Dll1, Bmp4 and VE-cadherin, and increased expression of Sox9 and EphB2. Conclusions Manipulating VEGF bioavailability leads to profound effects on not only the intestinal vasculature, but epithelial stem and progenitor cells in the intestinal crypt. Elucidation of the crosstalk between VEGF signaling in the vasculature, mesenchyme and epithelial stem/progenitor cell populations may direct future

  16. Moderate Hypoxia Exhibits Increased Endothelial Progenitor Vessel-forming Ability However Gestational Diabetes Caused to Impede Compensatory Defense Reaction

    PubMed Central

    Dincer, U. Deniz

    2016-01-01

    Endothelium represents a defense barrier and responds and integrates neuro humoral stimulus which describes as a compensatory mechanism. Endothelium formed with endothelial cells (ECs) and their progenitors. Endothelial progenitor cells (EPCs) represent minor subpopulation of mononuclear cells in the blood. During acute hypoxia, larger amount of EPCs mobilize into the peripheral blood and they directly contribute revascularization process. One of the subtypes of EPC is termed endothelial colony forming cells (ECFCs) which they possess de novo vessel-forming ability. The present study aims to investigate the role of hypoxia in EPCs functional and vessel-forming ability. Furthermore, it was investigated whether fetal exposure to a diabetic intrauterine environment influence EPCs adaptation ability. Human umbilical cord blood (HUCB) derived ECFCs were selected in all experimental procedures obtained from normal and gestational diabetes mellitus (GDM) subjects via in vitro cell culture methods. Early passage (<5) HUCB ECFCs obtain from GDM (n; 5) and control (n; 5) subjects were cultured with plates pre-coated with collagen in vitro 72 h hypoxic as well as normoxic condition. Endothelial, angiogenic and hypoxia associated gene specific primers designed to perform Real-time PCR. Senescenes assay conducted onto HUCB ECFCs to investigate their functional clonogenic ability. To quantify their vessel forming ability matrigel assay was applied. These data demonstrates that moderate hypoxia results increased vessel-forming ability and VEGFA expression in HUCB ECFCs obtained from control subjects. However, GDM caused to impede compensatory defense reaction against hypoxia which observed in control subjects. Thus, it illuminates beneficial information related future therapeutic modalities. PMID:27426097

  17. The Effects of Smoking on Levels of Endothelial Progenitor Cells and Microparticles in the Blood of Healthy Volunteers

    PubMed Central

    Mobarrez, Fariborz; Antoniewicz, Lukasz; Bosson, Jenny A.; Kuhl, Jeanette; Pisetsky, David S.; Lundbäck, Magnus

    2014-01-01

    Background Cigarette smoking, both active and passive, is one of the leading causes of morbidity and mortality in cardiovascular disease. To assess the impact of brief smoking on the vasculature, we determined levels of circulating endothelial progenitor cells (EPCs) and circulating microparticles (MPs) following the smoking of one cigarette by young, healthy intermittent smokers. Materials and Methods 12 healthy volunteers were randomized to either smoking or not smoking in a crossover fashion. Blood sampling was performed at baseline, 1, 4 and 24 hours following smoking/not smoking. The numbers of EPCs and MPs were determined by flow cytometry. MPs were measured from platelets, leukocytes and endothelial cells. Moreover, MPs were also labelled with anti-HMGB1 and SYTO 13 to assess the content of nuclear molecules. Results Active smoking of one cigarette caused an immediate and significant increase in the numbers of circulating EPCs and MPs of platelet-, endothelial- and leukocyte origin. Levels of MPs containing nuclear molecules were increased, of which the majority were positive for CD41 and CD45 (platelet- and leukocyte origin). CD144 (VE-cadherin) or HMGB1 release did not significantly change during active smoking. Conclusion Brief active smoking of one cigarette generated an acute release of EPC and MPs, of which the latter contained nuclear matter. Together, these results demonstrate acute effects of cigarette smoke on endothelial, platelet and leukocyte function as well as injury to the vascular wall. PMID:24587320

  18. Endothelial-to-mesenchymal transition in lipopolysaccharide-induced acute lung injury drives a progenitor cell-like phenotype.

    PubMed

    Suzuki, Toshio; Tada, Yuji; Nishimura, Rintaro; Kawasaki, Takeshi; Sekine, Ayumi; Urushibara, Takashi; Kato, Fumiaki; Kinoshita, Taku; Ikari, Jun; West, James; Tatsumi, Koichiro

    2016-06-01

    Pulmonary vascular endothelial function may be impaired by oxidative stress in endotoxemia-derived acute lung injury. Growing evidence suggests that endothelial-to-mesenchymal transition (EndMT) could play a pivotal role in various respiratory diseases; however, it remains unclear whether EndMT participates in the injury/repair process of septic acute lung injury. Here, we analyzed lipopolysaccharide (LPS)-treated mice whose total number of pulmonary vascular endothelial cells (PVECs) transiently decreased after production of reactive oxygen species (ROS), while the population of EndMT-PVECs significantly increased. NAD(P)H oxidase inhibition suppressed EndMT of PVECs. Most EndMT-PVECs derived from tissue-resident cells, not from bone marrow, as assessed by mice with chimeric bone marrow. Bromodeoxyuridine-incorporation assays revealed higher proliferation of capillary EndMT-PVECs. In addition, EndMT-PVECs strongly expressed c-kit and CD133. LPS loading to human lung microvascular endothelial cells (HMVEC-Ls) induced reversible EndMT, as evidenced by phenotypic recovery observed after removal of LPS. LPS-induced EndMT-HMVEC-Ls had increased vasculogenic ability, aldehyde dehydrogenase activity, and expression of drug resistance genes, which are also fundamental properties of progenitor cells. Taken together, our results demonstrate that LPS induces EndMT of tissue-resident PVECs during the early phase of acute lung injury, partly mediated by ROS, contributing to increased proliferation of PVECs. PMID:27106288

  19. Zoledronate Inhibits Ischemia-Induced Neovascularization by Impairing the Mobilization and Function of Endothelial Progenitor Cells

    PubMed Central

    Tsai, Shih-Hung; Huang, Po-Hsun; Chang, Wei-Chou; Tsai, Hsiao-Ya; Lin, Chih-Pei; Leu, Hsin-Bang; Wu, Tao-Cheng; Chen, Jaw-Wen; Lin, Shing-Jong

    2012-01-01

    Background Bisphosphonates are a class of pharmacologic compounds that are commonly used to treat postmenopausal osteoporosis and malignant osteolytic processes. Studies have shown that bone marrow-derived endothelial progenitor cells (EPCs) play a significant role in postnatal neovascularization. Whether the nitrogen-containing bisphosphonate zoledronate inhibits ischemia-induced neovascularization by modulating EPC functions remains unclear. Methodology/Principal Findings Unilateral hindlimb ischemia was surgically induced in wild-type mice after 2 weeks of treatment with vehicle or zoledronate (low-dose: 30 μg/kg; high-dose: 100 μg/kg). Doppler perfusion imaging demonstrated that the ischemic limb/normal side blood perfusion ratio was significantly lower in wild-type mice treated with low-dose zoledronate and in mice treated with high-dose zoledronate than in controls 4 weeks after ischemic surgery (control vs. low-dose vs. high-dose: 87±7% vs. *61±18% vs. **49±17%, *p<0.01, **p<0.005 compared to control). Capillary densities were also significantly lower in mice treated with low-dose zoledronate and in mice treated with high-dose zoledronate than in control mice. Flow cytometry analysis showed impaired mobilization of EPC-like cells (Sca-1+/Flk-1+) after surgical induction of ischemia in mice treated with zoledronate but normal levels of mobilization in mice treated with vehicle. In addition, ischemic tissue from mice that received zoledronate treatment exhibited significantly lower levels of the active form of MMP-9, lower levels of VEGF, and lower levels of phosphorylated eNOS and phosphorylated Akt than ischemic tissue from mice that received vehicle. Results of the in vitro studies showed that incubation with zoledronate inhibited the viability, migration, and tube-forming capacities of EPC. Conclusions/Significance Zoledronate inhibited ischemia-induced neovascularization by impairing EPC mobilization and angiogenic functions. These findings suggest

  20. Effects of corneal stromal cell- and bone marrow-derived endothelial progenitor cell-conditioned media on the proliferation of corneal endothelial cells

    PubMed Central

    Zhu, Meng-Yu; Yao, Qin-Ke; Chen, Jun-Zhao; Shao, Chun-Yi; Yan, Chen-Xi; Ni, Ni; Fan, Xian-Qun; Gu, Ping; Fu, Yao

    2016-01-01

    AIM To explore the effects of conditioned media on the proliferation of corneal endothelial cells (CECs) and to compare the efficiency of different conditioned media (CM). METHODS Rat CECs, corneal stromal cells (CSCs), bone marrow-derived endothelial progenitor cells (BEPCs), and bone marrow-derived mesenchymal stem cells (BMSCs) were isolated and cultured in vitro. CM was collected from CSCs, BEPCs, and BMSCs. CECs were cultivated in different culture media. Cell morphology was recorded, and gene and protein expression were analyzed. RESULTS After grown in CM for 5d, CECs in each experimental group remained polygonal, in a cobblestone-like monolayer arrangement. Immunocytofluorescence revealed positive expression of Na+/K+-ATP, aquaporin 1 (AQP1), and zonula occludens 1 (ZO-1). Based on quantitative polymerase chain reaction (qPCR) analysis, Na+/K+-ATP expression in CSC-CM was notably upregulated by 1.3-fold (±0.036) (P<0.05, n=3). The expression levels of ZO-1, neuron specific enolase (NSE), Vimentin, paired homebox 6 (PAX6), and procollagen type VIII (COL8A1) were notably upregulated in each experimental group. Each CM had a positive effect on CEC proliferation, and CSC-CM had the strongest effect on proliferation. CONCLUSION CSC-CM, BEPC-CM, and BMSC-CM not only stimulated the proliferation of CECs, but also maintained the characteristic differentiated phenotypes necessary for endothelial functions. CSC-CM had the most notable effect on CEC proliferation. PMID:27158599

  1. Endothelial cell specification in the somite is compromised in Pax3-positive progenitors of Foxc1/2 conditional mutants, with loss of forelimb myogenesis.

    PubMed

    Mayeuf-Louchart, Alicia; Montarras, Didier; Bodin, Catherine; Kume, Tsutomu; Vincent, Stéphane D; Buckingham, Margaret

    2016-03-01

    Pax3 and Foxc2 have been shown genetically to mutually repress each other in the mouse somite. Perturbation of this balance in multipotent cells of the dermomyotome influences cell fate; upregulation of Foxc2 favours a vascular fate, whereas higher levels of Pax3 lead to myogenesis. Foxc1 has overlapping functions with Foxc2. In Foxc1/2 double-mutant embryos, somitogenesis is severely affected, precluding analysis of somite derivatives. We have adopted a conditional approach whereby mutations in Foxc1 and Foxc2 genes were targeted to Pax3-expressing cells. Inclusion of a conditional reporter allele in the crosses made it possible to follow cells that had expressed Pax3. At the forelimb level, endothelial and myogenic cells migrate from adjacent somites into the limb bud. This population of endothelial cells is compromised in the double mutant, whereas excessive production of myogenic cells is observed in the trunk. However, strikingly, myogenic progenitors fail to enter the limbs, leading to the absence of skeletal muscle. Pax3-positive migratory myogenic progenitors, marked by expression of Lbx1, are specified in the somite at forelimb level, but endothelial progenitors are absent. The myogenic progenitors do not die, but differentiate prematurely adjacent to the somite. We conclude that the small proportion of somite-derived endothelial cells in the limb is required for the migration of myogenic limb progenitors. PMID:26839363

  2. [Preparation of Biological Functional Magnetic Nanoparticles and Study on the Effect of Guiding Endothelial Progenitor Cells In Vitro].

    PubMed

    Ma, Baolong; Yan, Wei; Chen, Jialong; Qi, Pengkai; Li, Jianhui; Huang, Nan

    2016-02-01

    Coprecipitation method was used to prepare triiron tetroxide magnetic nanoparticles enclosed in L-DOPA, and then EDC was used to activate the carboxyl group of L-DOPA after the nanoparticles were synthesized. The carboxyl group of L-DOPA formed amide bond with specific amino on the aptamer by dehydration condensation reaction. The surfaces of magnetic nanoparticles were modified with aptamer and L-DOPA. X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FT-IR), nanoparticle size analysis (SEM), magnetic measurement (VSM) and other testing methods were used to detect the magnetic nanoparticles in different stages. The endothelial progeni-tor cells (EPCs) were cocultured with the surface modified magnetic nanoparticles to evaluate cell compatibility and the combination effect of nanoparticles on EPCs in a short period of time. Directional guide of the surface-modified magnetic nanoparticles to endothelial progenitor cells (EPCs) was evaluated under an applied magnetic field and simulated dynamic blood flow condition. The results showed that the prepared magnetic nanoparticles had good magnetic response, good cell compatibility within a certain range of the nanoparticle concentrations. The surface modified nanoparticles could combine with EPCs effectively in a short time, and those nanoparticles combined EPCs can be directionally guided on to a stent surface under the magnetic field in the dynamic flow environment. PMID:27382754

  3. Characterization of Umbilical Cord Blood–Derived Late Outgrowth Endothelial Progenitor Cells Exposed to Laminar Shear Stress

    PubMed Central

    Brown, Melissa A.; Wallace, Charles S.; Angelos, Mathew

    2009-01-01

    Endothelial progenitor cells isolated from umbilical cord blood (CB-EPCs) represent a promising source of endothelial cells for synthetic vascular grafts and tissue-engineered blood vessels since they are readily attainable, can be easily isolated, and possess a high proliferation potential. The objective of this study was to compare the functional behavior of late outgrowth CB-EPCs with human aortic endothelial cells (HAECs). CB-EPCs and HAECs were cultured on either smooth muscle cells in a coculture model of a tissue-engineered blood vessels or fibronectin adsorbed to Teflon-AF™–coated glass slides. Late outgrowth CB-EPCs expressed endothelial cell–specific markers and were negative for the monocytic marker CD14. CB-EPCs have higher proliferation rates than HAECs, but are slightly smaller in size. CB-EPCs remained adherent under supraphysiological shear stresses, oriented and elongated in the direction of flow, and expressed similar numbers of α5β1 and αvβ3 integrins and antithrombotic genes compared to HAECs. There were some differences in mRNA levels of E-selectin and vascular cell adhesion molecule 1 between CB-EPCs and HAECs; however, protein levels were similar on the two cell types, and CB-EPCs did not support adhesion of monocytes in the absence of tumor necrosis factor-α stimulation. Although CB-EPCs expressed significantly less endothelial nitric oxide synthase protein after exposure to flow than HAECs, nitric oxide levels induced by flow were not significantly different. These results suggest that late outgrowth CB-EPCs are functionally similar to HAECs under flow conditions and are a promising cell source for cardiovascular therapies. PMID:19480571

  4. Polyphenol-rich Cranberry Juice has a neutral effect on endothelial function but decreases the fraction of osteocalcin expressing endothelial progenitor cells

    PubMed Central

    Flammer, Andreas J.; Martin, Elizabeth; Gossl, Mario; Widmer, R Jay; Lennon, Ryan; Sexton, Jasmine A.; Loeffler, Darrell; Khosla, Sundeep; Lerman, Lilach O.; Lerman, Amir

    2014-01-01

    Purpose Cranberry juice (CJ) contains a remarkably high concentration of polyphenols, considered to be beneficial for cardiovascular and bone health. The current double-blind, randomized study was designed to test whether daily consumption of double-strength Ocean Spray light CJ (2×230ml) over 4 months has beneficial effects on vascular function and on endothelial progenitor cells (EPCs), EPCs carrying the osteoblastic marker osteocalcin in particular. Methods 84 participants (49.5±16.2yrs.) with peripheral endothelial dysfunction and cardiovascular risk factors were enrolled in this double-blind, randomized, controlled trial (69 completed the four month protocol - 32 in the CJ group and 37 in the placebo group - respectively). Vascular responses to reactive hyperemia were measured non-invasively by peripheral arterial tonometry (EndoPAT). Peripheral blood mononuclear cells were stained for EPC markers, as well as osteocalcin, and counted by flow-cytometry. Results Baseline characteristics were similar in both groups. The effect of CJ on peripheral endothelial function and on circulating EPC counts (CD34+/CD133+/KDR+) did not change during the study. A high percentage of EPCs expressed osteocalcin (59.4±35.7%). CJ, as compared to placebo, induced a decrease in the fraction of EPCs expressing osteocalcin (-8.64±48.98 and 19.13±46.11%, respectively, p=0.019). Systemic levels of the adhesion marker ICAM correlated significantly with the number of EPCs expressing osteocalcin. Conclusions The study demonstrated that long term supplementation of polyphenol-rich CJ did not improve peripheral endothelial function. However, the decrease in the fraction of osteocalcin+ EPCs suggests a potential beneficial effect of polyphenol-rich CJ. PMID:22382203

  5. Inhibition of Store-Operated Calcium Entry Protects Endothelial Progenitor Cells from H2O2-Induced Apoptosis

    PubMed Central

    Wang, Yan-Wei; Zhang, Ji-Hang; Yu, Yang; Yu, Jie; Huang, Lan

    2016-01-01

    Store-operated calcium entry (SOCE), a major mode of extracellular calcium entry, plays roles in a variety of cell activities. Accumulating evidence indicates that the intracellular calcium ion concentration and calcium signaling are critical for the responses induced by oxidative stress. The present study was designed to investigate the potential effect of SOCE inhibition on H2O2-induced apoptosis in endothelial progenitor cells (EPCs), which are the predominant cells involved in endothelial repair. The results showed that H2O2-induced EPC apoptosis was reversed by SOCE inhibition induced either using the SOCE antagonist ML-9 or via silencing of stromal interaction molecule 1 (STIM1), a component of SOCE. Furthermore, SOCE inhibition repressed the increases in intracellular reactive oxygen species (ROS) levels and endoplasmic reticulum (ER) stress and ameliorated the mitochondrial dysfunction caused by H2O2. Our findings provide evidence that SOCE inhibition exerts a protective effect on EPCs in response to oxidative stress induced by H2O2 and may serve as a potential therapeutic strategy against vascular endothelial injury. PMID:27169819

  6. Live imaging of Runx1 expression in the dorsal aorta tracks the emergence of blood progenitors from endothelial cells.

    PubMed

    Lam, Enid Yi Ni; Hall, Christopher J; Crosier, Philip S; Crosier, Kathryn E; Flores, Maria Vega

    2010-08-12

    Blood cells of an adult vertebrate are continuously generated by hematopoietic stem cells (HSCs) that originate during embryonic life within the aorta-gonad-mesonephros region. There is now compelling in vivo evidence that HSCs are generated from aortic endothelial cells and that this process is critically regulated by the transcription factor Runx1. By time-lapse microscopy of Runx1-enhanced green fluorescent protein transgenic zebrafish embryos, we were able to capture a subset of cells within the ventral endothelium of the dorsal aorta, as they acquire hemogenic properties and directly emerge as presumptive HSCs. These nascent hematopoietic cells assume a rounded morphology, transiently occupy the subaortic space, and eventually enter the circulation via the caudal vein. Cell tracing showed that these cells subsequently populated the sites of definitive hematopoiesis (thymus and kidney), consistent with an HSC identity. HSC numbers depended on activity of the transcription factor Runx1, on blood flow, and on proper development of the dorsal aorta (features in common with mammals). This study captures the earliest events of the transition of endothelial cells to a hemogenic endothelium and demonstrates that embryonic hematopoietic progenitors directly differentiate from endothelial cells within a living organism. PMID:20453160

  7. Inhibition of Store-Operated Calcium Entry Protects Endothelial Progenitor Cells from H2O2-Induced Apoptosis.

    PubMed

    Wang, Yan-Wei; Zhang, Ji-Hang; Yu, Yang; Yu, Jie; Huang, Lan

    2016-07-01

    Store-operated calcium entry (SOCE), a major mode of extracellular calcium entry, plays roles in a variety of cell activities. Accumulating evidence indicates that the intracellular calcium ion concentration and calcium signaling are critical for the responses induced by oxidative stress. The present study was designed to investigate the potential effect of SOCE inhibition on H2O2-induced apoptosis in endothelial progenitor cells (EPCs), which are the predominant cells involved in endothelial repair. The results showed that H2O2-induced EPC apoptosis was reversed by SOCE inhibition induced either using the SOCE antagonist ML-9 or via silencing of stromal interaction molecule 1 (STIM1), a component of SOCE. Furthermore, SOCE inhibition repressed the increases in intracellular reactive oxygen species (ROS) levels and endoplasmic reticulum (ER) stress and ameliorated the mitochondrial dysfunction caused by H2O2. Our findings provide evidence that SOCE inhibition exerts a protective effect on EPCs in response to oxidative stress induced by H2O2 and may serve as a potential therapeutic strategy against vascular endothelial injury. PMID:27169819

  8. Detrimental effects of Bartonella henselae are counteracted by L-arginine and nitric oxide in human endothelial progenitor cells.

    PubMed

    Salvatore, Paola; Casamassimi, Amelia; Sommese, Linda; Fiorito, Carmela; Ciccodicola, Alfredo; Rossiello, Raffaele; Avallone, Bice; Grimaldi, Vincenzo; Costa, Valerio; Rienzo, Monica; Colicchio, Roberta; Williams-Ignarro, Sharon; Pagliarulo, Caterina; Prudente, Maria Evelina; Abbondanza, Ciro; Lamberti, Florentia; Baroni, Adone; Buommino, Elisabetta; Farzati, Bartolomeo; Tufano, Maria Antonietta; Ignarro, Louis Joseph; Napoli, Claudio

    2008-07-01

    The recruitment of circulating endothelial progenitor cells (EPCs) might have a beneficial effect on the clinical course of several diseases. Endothelial damage and detachment of endothelial cells are known to occur in infection, tissue ischemia, and sepsis. These detrimental effects in EPCs are unknown. Here we elucidated whether human EPCs internalize Bartonella henselae constituting a circulating niche of the pathogen. B. henselae invades EPCs as shown by gentamicin protection assays and transmission electron microscopy (TEM). Dil-Ac-LDL/lectin double immunostaining and fluorescence-activated cell sorting (FACS) analysis of EPCs revealed EPC bioactivity after infection with B. henselae. Nitric oxide (NO) and its precursor l-arginine (l-arg) exert a plethora of beneficial effects on vascular function and modulation of immune response. Therefore, we tested also the hypothesis that l-arg (1-30 mM) would affect the infection of B. henselae or tumor necrosis factor (TNF) in EPCs. Our data provide evidence that l-arg counteracts detrimental effects induced by TNF or Bartonella infections via NO (confirmed by DETA-NO and L-NMMA experiments) and by modulation of p38 kinase phosphorylation. Microarray analysis indicated several genes involved in immune response were differentially expressed in Bartonella-infected EPCs, whereas these genes returned in steady state when cells were exposed to sustained doses of l-arg. This mechanism may have broad therapeutic applications in tissue ischemia, angiogenesis, immune response, and sepsis. PMID:18595894

  9. Detrimental effects of Bartonella henselae are counteracted by l-arginine and nitric oxide in human endothelial progenitor cells

    PubMed Central

    Salvatore, Paola; Casamassimi, Amelia; Sommese, Linda; Fiorito, Carmela; Ciccodicola, Alfredo; Rossiello, Raffaele; Avallone, Bice; Grimaldi, Vincenzo; Costa, Valerio; Rienzo, Monica; Colicchio, Roberta; Williams-Ignarro, Sharon; Pagliarulo, Caterina; Prudente, Maria Evelina; Abbondanza, Ciro; Lamberti, Florentia; Baroni, Adone; Buommino, Elisabetta; Farzati, Bartolomeo; Tufano, Maria Antonietta; Ignarro, Louis Joseph; Napoli, Claudio

    2008-01-01

    The recruitment of circulating endothelial progenitor cells (EPCs) might have a beneficial effect on the clinical course of several diseases. Endothelial damage and detachment of endothelial cells are known to occur in infection, tissue ischemia, and sepsis. These detrimental effects in EPCs are unknown. Here we elucidated whether human EPCs internalize Bartonella henselae constituting a circulating niche of the pathogen. B. henselae invades EPCs as shown by gentamicin protection assays and transmission electron microscopy (TEM). Dil-Ac-LDL/lectin double immunostaining and fluorescence-activated cell sorting (FACS) analysis of EPCs revealed EPC bioactivity after infection with B. henselae. Nitric oxide (NO) and its precursor l-arginine (l-arg) exert a plethora of beneficial effects on vascular function and modulation of immune response. Therefore, we tested also the hypothesis that l-arg (1–30 mM) would affect the infection of B. henselae or tumor necrosis factor (TNF) in EPCs. Our data provide evidence that l-arg counteracts detrimental effects induced by TNF or Bartonella infections via NO (confirmed by DETA-NO and L-NMMA experiments) and by modulation of p38 kinase phosphorylation. Microarray analysis indicated several genes involved in immune response were differentially expressed in Bartonella-infected EPCs, whereas these genes returned in steady state when cells were exposed to sustained doses of l-arg. This mechanism may have broad therapeutic applications in tissue ischemia, angiogenesis, immune response, and sepsis. PMID:18595894

  10. A Case of Abnormal Lymphatic-Like Differentiation and Endothelial Progenitor Cell Activation in Neovascularization Associated with Hemi-Retinal Vein Occlusion

    PubMed Central

    Loukovaara, Sirpa; Gucciardo, Erika; Repo, Pauliina; Lohi, Jouko; Salven, Petri; Lehti, Kaisa

    2015-01-01

    Purpose Pathological vascular differentiation in retinal vein occlusion (RVO)-related neovessel formation remains poorly characterized. The role of intraocular lymphatic-like differentiation or endothelial progenitor cell activity has not been studied in this disease. Methods Vitrectomy was performed in an eye with hemi-RVO; the neovessel membrane located at the optic nerve head was removed and subjected to immunohistochemistry. Characterization of the neovascular tissue was performed using hematoxylin and eosin, α-smooth muscle actin, and the pan-endothelial cell (EC) adhesion molecule CD31. The expression of lymphatic EC markers was studied by lymphatic vessel endothelial hyaluronan receptor 1 (LYVE-1), podoplanin (PDPN), and prospero-related homeobox protein 1 (Prox-1). Potential vascular stem/progenitor cells were identified by active cellular proliferation (Ki67) and expression of the stem cell marker CD117. Results The specimen contained blood vessels lined by ECs and surrounded by pericytes. Immunoreactivity for LYVE-1 and Prox-1 was detected, with Prox-1 being more widely expressed in the active Ki67-positive lumen-lining cells. PDPN expression was instead found in the cells residing in the extravascular tissue. Expression of the stem cell markers CD117 and Ki67 suggested vascular endothelial progenitor cell activity. Conclusions Intraocular lymphatic-like differentiation coupled with progenitor cell activation may be involved in the pathology of neovessel formation in ischemia-induced human hemi-RVO. PMID:26327908

  11. Analyses of Endothelial Cells and Endothelial Progenitor Cells Released Microvesicles by Using Microbead and Q-dot Based Nanoparticle Tracking Analysis

    PubMed Central

    Wang, Jinju; Zhong, Yun; Ma, Xiaotang; Xiao, Xiang; Cheng, Chuanfang; Chen, Yusen; Iwuchukwu, Ifeanyi; Gaines, Kenneth J.; Bin Zhao; Liu, Shiming; Travers, Jeffrey B.; Bihl, Ji C.; Chen, Yanfang

    2016-01-01

    Accurate analysis of specific microvesicles (MVs) from biofluids is critical and challenging. Here we described novel methods to purify and detect MVs shed from endothelial cells (ECs) and endothelial progenitor cells (EPCs) by combining microbeads with fluorescence quantum dots (Q-dots) coupled nanoparticle tracking analysis (NTA). In the in vitro screening systems, we demonstrated that 1) anti-CD105 (EC marker) and anti-CD34 (EPC marker) conjugated-microbeads had the highest sensitivity and specificity for isolating respective MVs, which were confirmed with negative controls, CD41 and CD235a; 2) anti-CD144 (EC marker) and anti-KDR (EPC marker) conjugated-Q-dots exhibited the best sensitivity and specificity for their respective MV NTA detection, which were confirmed with positive control, anti-Annexin V (MV universal marker). The methods were further validated by their ability to efficiently recover the known amount of EC-MVs and EPC-MVs from particle-depleted plasma, and to detect the dynamical changes of plasma MVs in ischemic stroke patients, as compared with traditional flow cytometry. These novel methods provide ideal approaches for functional analysis and biomarker discovery of ECs- and EPCs- derived MVs. PMID:27094208

  12. Bone Marrow-Derived Endothelial Progenitor Cells Protect Against Scopolamine-Induced Alzheimer-Like Pathological Aberrations.

    PubMed

    Safar, Marwa M; Arab, Hany H; Rizk, Sherine M; El-Maraghy, Shohda A

    2016-04-01

    Vascular endothelial dysfunction plays a key role in the pathogenesis of Alzheimer's disease (AD). Patients with AD have displayed decreased circulating endothelial progenitor cells (EPCs) which repair and maintain the endothelial function. Transplantation of EPCs has emerged as a promising approach for the management of cerebrovascular diseases including ischemic stroke, however, its impact on AD has been poorly described. Thus, the current study aimed at investigating the effects of bone marrow-derived (BM) EPCs transplantation in repeated scopolamine-induced cognitive impairment, an experimental model that replicates biomarkers of AD. Intravenously transplanted BM-EPCs migrated into the brain of rats and improved the learning and memory deficits. Meanwhile, they mitigated the deposition of amyloid plaques and associated histopathological alterations. At the molecular levels, BM-EPCs blunted the increase of hippocampal amyloid beta protein (Aβ), amyloid precursor protein (APP) and reinstated the Aβ-degrading neprilysin together with downregulation of p-tau and its upstream glycogen synthase kinase-3β (GSK-3β). They also corrected the perturbations of neurotransmitter levels including restoration of acetylcholine and associated esterase along with dopamine, GABA, and the neuroexitatory glutamate. Furthermore, BM-EPCs induced behavioral recovery via boosting of vascular endothelial growth factor (VEGF), nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF) and its upstream cAMP response element binding (CREB), suppression of the proinflammatory tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β), and upregulation of interleukin-10 (IL-10). BM-EPCs also augmented Nrf2 and seladin-1. Generally, these actions were analogous to those exerted by adipose tissue-derived mesenchymal stem cells (AT-MSCs) and the reference anti-Alzheimer donepezil. For the first time, these findings highlight the beneficial actions of BM-EPCs against the memory

  13. Vitamin D Prevents Endothelial Progenitor Cell Dysfunction Induced by Sera from Women with Preeclampsia or Conditioned Media from Hypoxic Placenta

    PubMed Central

    Myerski, Ashley C.; von Kaisenberg, Constantin S.; Grundmann, Magdalena; Hubel, Carl A.; von Versen-Höynck, Frauke

    2014-01-01

    Context Placenta-derived circulating factors contribute to the maternal endothelial dysfunction underlying preeclampsia. Endothelial colony forming cells (ECFC), a sub-population of endothelial progenitor cells (EPCs), are thought to be involved in vasculogenesis and endothelial repair. Low vitamin D concentrations are associated with an increased risk for preeclampsia. Objective We hypothesized that the function of human fetal ECFCs in culture would be suppressed by exposure to preeclampsia-related factors–preeclampsia serum or hypoxic placental conditioned medium– in a fashion reversed by vitamin D. Design, Setting, Patients ECFCs were isolated from cord blood of uncomplicated pregnancies and expanded in culture. Uncomplicated pregnancy villous placenta in explant culture were exposed to either 2% (hypoxic), 8% (normoxic) or 21% (hyperoxic) O2 for 48 h, after which the conditioned media (CM) was collected. Outcome Measures ECFC tubule formation (Matrigel assay) and migration were examined in the presence of either maternal serum from preeclampsia cases or uncomplicated pregnancy controls, or pooled CM, in the presence or absence of 1,25(OH)2 vitamin D3. Results 1,25(OH)2 vitamin D3 reversed the adverse effects of preeclampsia serum or CM from hypoxic placenta on ECFCs capillary-tube formation and migration. Silencing of VDR expression by VDR siRNA, VDR blockade, or VEGF pathway blockade reduced ECFC functional abilities. Effects of VDR or VEGF blockade were partially prevented by vitamin D. Conclusion Vitamin D promotes the capillary-like tubule formation and migration of ECFCs in culture, minimizing the negative effects of exposure to preeclampsia-related factors. Further evaluation of the role of vitamin D in ECFC regulation and preeclampsia is warranted. PMID:24887145

  14. Impact of an endothelial progenitor cell capturing stent on coronary microvascular function: comparison with drug-eluting stents

    PubMed Central

    Choi, Woong Gil; Kim, Soo Hyun; Yoon, Hyung Seok; Lee, Eun Joo

    2015-01-01

    Background/Aims Although drug-eluting stents (DESs) effectively reduce restenosis following percutaneous coronary intervention (PCI), they also delay re-endothelialization and impair microvascular function, resulting in adverse clinical outcomes. Endothelial progenitor cell (EPC) capturing stents, by providing a functional endothelial layer on the stent, have beneficial effects on microvascular function. However, data on coronary microvascular function in patients with EPC stents versus DESs are lacking. Methods Seventy-four patients who previously underwent PCI were enrolled in this study. Microvascular function was evaluated 6 months after PCI based on the index of microvascular resistance (IMR) and the coronary flow reserve (CFR). IMR was calculated as the ratio of the mean distal coronary pressure at maximal hyperemia to the inverse of the hyperemic mean transit time (hTmn). The CFR was calculated by dividing the hTmn by the baseline mean transit time. Results Twenty-one patients (age, 67.2 ± 9.6 years; male:female, 15:6) with an EPC stent and 53 patients (age, 61.5 ± 14.7 years; male:female, 40:13) with second-generation DESs were included in the study. There were no significant differences in the baseline clinical and angiographic characteristics of the two groups. Angiography performed 6 months postoperatively did not show significant differences in their CFR values. However, patients with the EPC stent had a significantly lower IMR than patients with second-generation DESs (median, 25.5 [interquartile range, 12.85 to 28.18] vs. 29.0 [interquartile range, 15.42 to 39.23]; p = 0.043). Conclusions Microvascular dysfunction was significantly improved after 6 months in patients with EPC stents compared to those with DESs. The complete re-endothelialization achieved with the EPC stent may provide clinical benefits over DESs, especially in patients with microvascular dysfunction. PMID:25589834

  15. Immobilization of DNA aptamers via plasma polymerized allylamine film to construct an endothelial progenitor cell-capture surface.

    PubMed

    Qi, Pengkai; Yan, Wei; Yang, Ying; Li, Yalong; Fan, Yi; Chen, Junying; Yang, Zhilu; Tu, Qiufen; Huang, Nan

    2015-02-01

    The endothelial progenitor cells (EPCs) capture stent has drawn increasing attentions and become one of the most promising concepts for the next generation vascular stent. In this regard, it is of great significance to immobilize a molecule with the ability to bind EPC for rapid in vivo endothelialization with high specificity. In this work, a facile two-step method aimed at constructing a coating with specific EPC capturing aptamers is reported. The processes involves as the first-step deposition of plasma polymerized allylamine (PPAam) on a substrate to introduce amine groups, followed by the electrostatic adsorption of a 34 bases single strand DNA sequence to the PPAam surface as a second step (PPAam-DNA). Grazing incidence attenuated total reflection Fourier transform infrared spectroscopy (GATR-FTIR) and X-ray photoelectron spectroscopy (XPS) confirmed the successful immobilization of the aptamers. Quartz crystal microbalance with dissipation (QCM-D) real time monitoring result shows that about 175 ng/cm(2) aptamers were conjugated onto the PPAam surface. The interactions between the modified surfaces and human umbilical vein endothelial cells (ECs), smooth muscle cells (SMCs), and murine induced EPCs derived from mesenchymal stem cells (MSCs) were also investigated. It was demonstrated that PPAam-DNA samples could capture more EPCs, and present a cellular friendly surface for the proliferation of both EPCs and ECs but no effect on the hyperplasia of SMCs. Also, the co-culture results of 3 types of cells confirmed that the aptamer could specifically bond EPCs rather than ECs and SMCs, suggesting the competitive adhesion advantage of EPCs to ECs and SMCs. These data demonstrate that the EPC aptamer has large potential for designing an EPC captured stent and other vascular grafts with targeted in situ endothelialization. PMID:25575347

  16. In vivo endothelization of tubular vascular grafts through in situ recruitment of endothelial and endothelial progenitor cells by RGD-fused mussel adhesive proteins.

    PubMed

    Kang, Tae-Yun; Lee, Jung Ho; Kim, Bum Jin; Kang, Jo-A; Hong, Jung Min; Kim, Byoung Soo; Cha, Hyung Joon; Rhie, Jong-Won; Cho, Dong-Woo

    2015-01-01

    The use of tissue mimics in vivo, including patterned vascular networks, is expected to facilitate the regeneration of functional tissues and organs with large volumes. Maintaining patency of channels in contact with blood is an important issue in the development of a functional vascular network. Endothelium is the only known completely non-thrombogenic material; however, results from treatments to induce endothelialization are inconclusive. The present study was designed to evaluate the clinical applicability of in situ recruitment of endothelial cells/endothelial progenitor cells (EC/EPC) and pre-endothelization using a recombinant mussel adhesive protein fused with arginine-glycine-aspartic acid peptide (MAP-RGD) coating in a model of vascular graft implantation. Microporous polycaprolactone (PCL) scaffolds were fabricated with salt leaching methods and their surfaces were modified with collagen and MAP-RGD. We then evaluated their anti-thrombogenicity with an in vitro hemocompatibility assessment and a 4-week implantation in the rabbit carotid artery. We observed that MAP-RGD coating reduced the possibility of early in vivo graft failure and enhanced re-endothelization by in situ recruitment of EC/EPC (patency rate: 2/3), while endothelization prior to implantation aggravated the formation of thrombosis and/or IH (patency rate: 0/3). The results demonstrated that in situ recruitment of EC/EPC by MAP-RGD could be a promising strategy for vascular applications. In addition, it rules out several issues associated with pre-endothelization, such as cell source, purity, functional modulation and contamination. Further evaluation of long term performance and angiogenesis from the luminal surface may lead to the clinical use of MAP-RGD for tubular vascular grafts and regeneration of large-volume tissues with functional vascular networks. PMID:25599716

  17. Tissue Engineering Special Feature: A macroporous hydrogel for the coculture of neural progenitor and endothelial cells to form functional vascular networks in vivo

    NASA Astrophysics Data System (ADS)

    Ford, Millicent C.; Bertram, James P.; Royce Hynes, Sara; Michaud, Michael; Li, Qi; Young, Michael; Segal, Steven S.; Madri, Joseph A.; Lavik, Erin B.

    2006-02-01

    A microvascular network is critical for the survival and function of most tissues. We have investigated the potential of neural progenitor cells to augment the formation and stabilization of microvascular networks in a previously uncharacterized three-dimensional macroporous hydrogel and the ability of this engineered system to develop a functional microcirculation in vivo. The hydrogel is synthesized by cross-linking polyethylene glycol with polylysine around a salt-leached polylactic-co-glycolic acid scaffold that is degraded in a sodium hydroxide solution. An open macroporous network is formed that supports the efficient formation of tubular structures by brain endothelial cells. After subcutaneous implantation of hydrogel cocultures in mice, blood flow in new microvessels was apparent at 2 weeks with perfused networks established on the surface of implants at 6 weeks. Compared to endothelial cells cultured alone, cocultures of endothelial cells and neural progenitor cells had a significantly greater density of tubular structures positive for platelet endothelial cell adhesion molecule-1 at the 6-week time point. In implant cross sections, the presence of red blood cells in vessel lumens confirmed a functional microcirculation. These findings indicate that neural progenitor cells promote the formation of endothelial cell tubes in coculture and the development of a functional microcirculation in vivo. We demonstrate a previously undescribed strategy for creating stable microvascular networks to support engineered tissues of desired parenchymal cell origin. microvasculature | neural stem cells | polymer | scaffold

  18. Endothelial Progenitor Cell Migration-Enhancing Factors in the Secretome of Placental-Derived Mesenchymal Stem Cells

    PubMed Central

    Kamprom, Witchayaporn; Kheolamai, Pakpoom; U-Pratya, Yaowalak; Supokawej, Aungkura; Wattanapanitch, Methichit; Laowtammathron, Chuti; Roytrakul, Sittiruk; Issaragrisil, Surapol

    2016-01-01

    Therapeutic potentials of mesenchymal stem cells (MSCs) depend largely on their ability to secrete cytokines or factors that modulate immune response, enhance cell survival, and induce neovascularization in the target tissues. We studied the secretome profile of gestational tissue-derived MSCs and their effects on functions of endothelial progenitor cells (EPCs), another angiogenic cell type that plays an important role during the neovascularization. MSCs derived from placental tissues (PL-MSCs) significantly enhanced EPC migration while BM-MSCs, which are the standard source of MSCs for various clinical applications, did not. By using protein fractionation and mass spectrometry analysis, we identified several novel candidates for EPC migration enhancing factor in PL-MSCs secretome that could be used to enhance neovascularization in the injured/ischemic tissues. We recommend that the strategy developed in our study could be used to systematically identify therapeutically useful molecules in the secretomes of other MSC sources for the clinical applications. PMID:26880942

  19. MicroRNAs as potential novel therapeutic targets and tools for regulating paracrine function of endothelial progenitor cells

    PubMed Central

    Xu, Shengjie; Jin, Chongying; Shen, Xiaohua; Ding, Fang; Zhu, Junhui; Fu, Guosheng

    2012-01-01

    Summary Endothelial progenitor cells (EPCs) play a protective role in the cardiovascular system by enhancing the maintenance of endothelium homeostasis and the process of new vessel formation. Recent studies show that EPCs may induce vascular regeneration and neovascularization mainly through paracrine signaling, that is, through the secretion of growth factors and pro-angiogenic cytokines [1]. However, multiple factors might function synergistically and therefore make it difficult to manipulate EPC paracrine effects. MicroRNAs, a family of small, non-coding RNAs, are characterized by post-transcriptionally regulating multiple functionally related genes, which renders them potentially powerful therapeutic targets or tools. In this paper we propose the hypothesis that microRNAs can be utilized as a novel therapeutic strategy for regulating EPC paracrine secretion. PMID:22739741

  20. Stimulatory Influences of Far Infrared Therapy on the Transcriptome and Genetic Networks of Endothelial Progenitor Cells Receiving High Glucose Treatment

    PubMed Central

    Lin, Tzu-Chiao; Lin, Chin-Sheng; Tsai, Tsung-Neng; Cheng, Shu-Meng; Lin, Wei-Shiang; Cheng, Cheng-Chung; Wu, Chun-Hsien; Hsu, Chih-Hsueng

    2015-01-01

    Background Endothelial progenitor cells (EPCs) play a fundamental role in vascular repair and angiogenesis- related diseases. It is well-known that the process of angiogenesis is faulty in patients with diabetes. Long-term exposure of peripheral blood EPCs to high glucose (HG-EPCs) has been shown to impair cell proliferation and other functional competencies. Far infrared (FIR) therapy can promote ischemia-induced angiogenesis in diabetic mice and restore high glucose-suppressed endothelial progenitor cell functions both in vitro and in vivo. However, the detail mechanisms and global transcriptome alternations are still unclear. Methods In this study, we investigated the influences of FIR upon HG-EPC gene expressions. EPCs were obtained from the peripheral blood and treated with high glucose. These cells were then subjected to FIR irradiation and functional assays. Results Those genes responsible for fibroblast growth factors, Mitogen-activated protein kinases (MAPK), Janus kinase/signal transducer and activator of transcription and prostaglandin signaling pathways were significantly induced in HG-EPCs after FIR treatment. On the other hand, mouse double minute 2 homolog, genes involved in glycogen metabolic process, and genes involved in cardiac fibrosis were down-regulated. We also observed complex genetic networks functioning in FIR-treated HG-EPCs, in which several genes, such as GATA binding protein 3, hairy and enhancer of split-1, Sprouty Homolog 2, MAPK and Sirtuin 1, acted as hubs to maintain the stability and connectivity of the whole genetic network. Conclusions Deciphering FIR-affected genes will not only provide us with new knowledge regarding angiogenesis, but also help to develop new biomarkers for evaluating the effects of FIR therapy. Our findings may also be adapted to develop new methods to increase EPC activities for treating diabetes-related ischemia and metabolic syndrome-associated cardiovascular disorders. PMID:27122901

  1. Overexpression of hypoxia-inducible factor-1 alpha improves vasculogenesis-related functions of endothelial progenitor cells.

    PubMed

    Kütscher, Christian; Lampert, Florian M; Kunze, Mirjam; Markfeld-Erol, Filiz; Stark, G Björn; Finkenzeller, Günter

    2016-05-01

    Postnatal vasculogenesis is mediated by mobilization of endothelial progenitor cells (EPCs) from bone marrow and homing to ischemic tissues. This feature emphasizes this cell type for cell-based therapies aiming at the improvement of neovascularization in tissue engineering applications and regenerative medicine. In animal models, it was demonstrated that implantation of EPCs from cord blood (cbEPCs) led to the formation of a complex functional neovasculature, whereas EPCs isolated from adult peripheral blood (pbEPCs) showed a limited vasculogenic potential, which may be attributed to age-related dysfunction. Recently, it was demonstrated that activation of hypoxia-inducible factor-1α (Hif-1α) improves cell functions of progenitor cells of mesenchymal and endothelial origin. Thus, we hypothesized that overexpression of Hif-1α may improve the vasculogenesis-related phenotype of pbEPCs. In the present study, we overexpressed Hif-1α in pbEPCs and cbEPCs by using recombinant adenoviruses and investigated effects on stem cell- and vasculogenesis-related cell parameters. Overexpression of Hif-1α enhanced proliferation, invasion, cell survival and in vitro capillary sprout formation of both EPC populations. Migration was increased in cbEPCs upon Hif-1α overexpression, but not in pbEPCs. Cellular senescence was decreased in pbEPCs, while remained in cbEPCs, which showed, as expected, intrinsically a dramatically lower senescent phenotype in relation to pbEPCs. Similarly, the colony-formation capacity was much higher in cbEPCs in comparison to pbEPCs and was further increased by Hif-1α overexpression, whereas Hif-1α transduction exerted no significant influence on colony formation of pbEPCs. In summary, our experiments illustrated multifarious effects of Hif-1α overexpression on stem cell and vasculogenic parameters. Therefore, Hif-1α overexpression may represent a therapeutic option to improve cellular functions of adult as well as postnatal EPCs. PMID:26827661

  2. LTβR controls thymic portal endothelial cells for haematopoietic progenitor cell homing and T-cell regeneration

    PubMed Central

    Shi, Yaoyao; Wu, Weiwei; Chai, Qian; Li, Qingqing; Hou, Yu; Xia, Huan; Ren, Boyang; Xu, Hairong; Guo, Xiaohuan; Jin, Caiwei; Lv, Mengjie; Wang, Zhongnan; Fu, Yang-Xin; Zhu, Mingzhao

    2016-01-01

    Continuous thymic homing of haematopoietic progenitor cells (HPCs) via the blood is critical for normal T-cell development. However, the nature and the differentiation programme of specialized thymic endothelial cells (ECs) controlling this process remain poorly understood. Here using conditional gene-deficient mice, we find that lymphotoxin beta receptor (LTβR) directly controls thymic ECs to guide HPC homing. Interestingly, T-cell deficiency or conditional ablation of T-cell-engaged LTβR signalling results in a defect in thymic HPC homing, suggesting the feedback regulation of thymic progenitor homing by thymic products. Furthermore, we identify and characterize a special thymic portal EC population with features that guide HPC homing. LTβR is essential for the differentiation and homeostasis of these thymic portal ECs. Finally, we show that LTβR is required for T-cell regeneration on irradiation-induced thymic injury. Together, these results uncover a cellular and molecular pathway that governs thymic EC differentiation for HPC homing. PMID:27493002

  3. New Method for Sorting Endothelial and Neural Progenitors from Human Induced Pluripotent Stem Cells by Sedimentation Field Flow Fractionation.

    PubMed

    Faye, Pierre-Antoine; Vedrenne, Nicolas; De la Cruz-Morcillo, Miguel A; Barrot, Claire-Cécile; Richard, Laurence; Bourthoumieu, Sylvie; Sturtz, Franck; Funalot, Benoît; Lia, Anne-Sophie; Battu, Serge

    2016-07-01

    Human induced pluripotent stem cells (hiPSc) are a very useful solution to create and observe the behavior of specific and usually inaccessible cells, such as human motor neurons. Obtained from a patient biopsy by reprograming dermal fibroblasts (DF), hiPSc present the same properties as embryonic stem cells and can generate any cell type after several weeks of differentiation. Today, there are numerus protocols which aim to control hiPSC differentiation. The principal challenge is to obtain a sufficiently enriched specific cell population to study disease pathophysiology and to provide a good model for further investigation and drug screening. The differentiation process is very costly and time-consuming, because many specific factors and different culture media must be used. In this study, we used Sedimentation Field Flow Fractionation (SdFFF) to prepare enriched populations derived from hiPSc after only 10 days of culture in a classical medium. Based on phenotypic and proteomic characterization, "hyperlayer" elution resulted in a fraction expressing markers of endothelial progenitors while another fraction expressed markers of neural progenitors. The isolation of subpopulations representing various differentiation lineages is of major interest for the production of specialized, cell-enriched fractions and in the preparation of increasingly complex models for the development of new therapeutic tools. PMID:27263863

  4. Circulating endothelial progenitor cells: a new approach to anti-aging medicine?

    PubMed Central

    2009-01-01

    Endothelial dysfunction is associated with major causes of morbidity and mortality, as well as numerous age-related conditions. The possibility of preserving or even rejuvenating endothelial function offers a potent means of preventing/treating some of the most fearful aspects of aging such as loss of mental, cardiovascular, and sexual function. Endothelial precursor cells (EPC) provide a continual source of replenishment for damaged or senescent blood vessels. In this review we discuss the biological relevance of circulating EPC in a variety of pathologies in order to build the case that these cells act as an endogenous mechanism of regeneration. Factors controlling EPC mobilization, migration, and function, as well as therapeutic interventions based on mobilization of EPC will be reviewed. We conclude by discussing several clinically-relevant approaches to EPC mobilization and provide preliminary data on a food supplement, Stem-Kine, which enhanced EPC mobilization in human subjects. PMID:20003528

  5. Intradialytic aerobic cycling exercise alleviates inflammation and improves endothelial progenitor cell count and bone density in hemodialysis patients.

    PubMed

    Liao, Min-Tser; Liu, Wen-Chih; Lin, Fu-Huang; Huang, Ching-Feng; Chen, Shao-Yuan; Liu, Chuan-Chieh; Lin, Shih-Hua; Lu, Kuo-Cheng; Wu, Chia-Chao

    2016-07-01

    Inflammation, endothelial dysfunction, and mineral bone disease are critical factors contributing to morbidity and mortality in hemodialysis (HD) patients. Physical exercise alleviates inflammation and increases bone density. Here, we investigated the effects of intradialytic aerobic cycling exercise on HD patients. Forty end-stage renal disease patients undergoing HD were randomly assigned to either an exercise or control group. The patients in the exercise group performed a cycling program consisting of a 5-minute warm-up, 20 minutes of cycling at the desired workload, and a 5-minute cool down during 3 HD sessions per week for 3 months. Biochemical markers, inflammatory cytokines, nutritional status, the serum endothelial progenitor cell (EPC) count, bone mineral density, and functional capacity were analyzed. After 3 months of exercise, the patients in the exercise group showed significant improvements in serum albumin levels, the body mass index, inflammatory cytokine levels, and the number of cells positive for CD133, CD34, and kinase insert domain-conjugating receptor. Compared with the exercise group, the patients in the control group showed a loss of bone density at the femoral neck and no increases in EPCs. The patients in the exercise group also had a significantly greater 6-minute walk distance after completing the exercise program. Furthermore, the number of EPCs significantly correlated with the 6-minute walk distance both before and after the 3-month program. Intradialytic aerobic cycling exercise programs can effectively alleviate inflammation and improve nutrition, bone mineral density, and exercise tolerance in HD patients. PMID:27399127

  6. High Calcium Bioglass Enhances Differentiation and Survival of Endothelial Progenitor Cells, Inducing Early Vascularization in Critical Size Bone Defects

    PubMed Central

    Nguyen Ngoc, Christina; Meier, Simon; Nau, Christoph; Schaible, Alexander; Marzi, Ingo; Henrich, Dirk

    2013-01-01

    Early vascularization is a prerequisite for successful bone healing and endothelial progenitor cells (EPC), seeded on appropriate biomaterials, can improve vascularization. The type of biomaterial influences EPC function with bioglass evoking a vascularizing response. In this study the influence of a composite biomaterial based on polylactic acid (PLA) and either 20 or 40% bioglass, BG20 and BG40, respectively, on the differentiation and survival of EPCs in vitro was investigated. Subsequently, the effect of the composite material on early vascularization in a rat calvarial critical size defect model with or without EPCs was evaluated. Human EPCs were cultured with β-TCP, PLA, BG20 or BG40, and seeding efficacy, cell viability, cell morphology and apoptosis were analysed in vitro. BG40 released the most calcium, and improved endothelial differentiation and vitality best. This effect was mimicked by adding an equivalent amount of calcium to the medium and was diminished in the presence of the calcium chelator, EGTA. To analyze the effect of BG40 and EPCs in vivo, a 6-mm diameter critical size calvarial defect was created in rats (n = 12). Controls (n = 6) received BG40 and the treatment group (n = 6) received BG40 seeded with 5×105 rat EPCs. Vascularization after 1 week was significantly improved when EPCs were seeded onto BG40, compared to implanting BG40 alone. This indicates that Ca2+ release improves EPC differentiation and is useful for enhanced early vascularization in critical size bone defects. PMID:24244419

  7. Inhibitor of DNA binding 1 regulates cell cycle progression of endothelial progenitor cells through induction of Wnt2 expression.

    PubMed

    Xia, Xi; Yu, Yang; Zhang, Li; Ma, Yang; Wang, Hong

    2016-09-01

    Endothelial injury is a risk factor for atherosclerosis. Endothelial progenitor cell (EPC) proliferation contributes to vascular injury repair. Overexpression of inhibitor of DNA binding 1 (Id1) significantly promotes EPC proliferation; however, the underlying molecular mechanism remains to be fully elucidated. The present study investigated the role of Id1 in cell cycle regulation of EPCs, which is closely associated with proliferation. Overexpression of Id1 increased the proportion of EPCs in the S/G2M phase and significantly increased cyclin D1 expression levels, while knockdown of Id1 arrested the cell cycle progression of EPCs in the G1 phase and inhibited cyclin D1 expression levels. In addition, it was demonstrated that Id1 upregulated wingless‑type mouse mammary tumor virus integration site family member 2 (Wnt2) expression levels and promoted β‑catenin accumulation and nuclear translocation. Furthermore, Wnt2 knockdown counteracted the effects of Id1 on cell cycle progression of EPCs. In conclusion, the results of the present study indicate that Id1 promoted Wnt2 expression, which accelerated cell cycle progression from G1 to S phase. This suggests that Id1 may promote cell cycle progression of EPCs, and that Wnt2 may be important in Id1 regulation of the cell cycle of EPCs. PMID:27432753

  8. The relationship between the number of preprocedural circulating endothelial progenitor cells and angiographic restenosis following coronary artery stent placement

    PubMed Central

    Klomp, Margo; van Tiel, Claudia M; Klous, Anita M; Beijk, Marcel A M; Klees, Margriet I; Scheunhage, Esther M; Tijssen, Jan G P; de Vries, Carlie J M; de Winter, Robbert J

    2011-01-01

    Objective In animals, endothelial progenitor cells (EPCs) beneficially influence the repair of the coronary vessel wall after damage by stent placement. However, their role in humans is less well understood. In the present study, the authors aimed to evaluate the relationship between the number of preprocedural EPCs defined as CD34+/KDR+/CD133+ cells and angiographic late loss as a measure of the growth of in-stent intimal hyperplasia. Design, setting, patients and interventions The 59 study patients were treated in the authors' clinic with a Genous EPC capturing stent, a bare metal stent (BMS) or a drug-eluting stent, and angiographic follow-up occurred between 6 and 13 months. Results The authors found no relationship between preprocedural EPCs and angiographic late loss, irrespective of stent type. Though statistically not significant, patients with a high number of preprocedural CD34 cells and treated with a Genous stent or BMS showed a numerically higher late loss (in Genous patients: 1.03±0.76 mm vs 0.71±0.50 mm, p=0.15; in BMS patients: 1.06±0.73 mm vs 0.35±0.62 mm, p=0.08). Conclusions Considering these and other varied observations, further studies aimed at identifying the biological mechanism and the individual roles of EPCs and/or CD34 cells in endothelial repair after coronary vessel stenting are needed.

  9. Apelin/APJ signaling promotes hypoxia-induced proliferation of endothelial progenitor cells via phosphoinositide-3 kinase/Akt signaling.

    PubMed

    Zhang, Jingchang; Liu, Qiming; Hu, Xinqun; Fang, Zhenfei; Huang, Feng; Tang, Liang; Zhou, Shenghua

    2015-09-01

    Endothelial progenitor cells (EPCs) can adhere to the endothelium at sites of hypoxia/ischemia and participate in the formation of novel vessels through differentiating into endothelial cells (ECs). Apelin is an endogenous ligand for the G protein‑coupled receptor APJ, and apelin/APJ signaling has a role in cardiovascular function. The present study aimed to investigate the role of apelin/APJ signaling in the regulation of EPC proliferation under hypoxia. The results showed that hypoxia was able to induce EPC proliferation, accompanied with an upregulation of hypoxia‑inducible factor (HIF)‑1α as well as apelin/APJ signaling. Further investigation indicated that siRNA‑mediated knockdown of apelin or APJ expression attenuated the hypoxia‑induced proliferation of EPCs, suggesting that apelin/APJ signaling has an important role in hypoxia‑induced EPC proliferation. Moreover, the phosphoinositide‑3 kinase (PI3K)/Akt signaling pathway was found to be involved in the apelin/APJ‑mediated EPC proliferation under hypoxia. Based on these findings, the present study suggested that hypoxia‑induced upregulation of HIF‑1α promotes the expression of apelin and APJ, which further activate the downstream PI3K/Akt signaling pathway, a key promoter of EPC proliferation. In conclusion, the present study highlighted the role of apelin/APJ in the regulation of EPC proliferation, and apelin/APJ may therefore serve as a potential target for the prevention of hypoxic ischemic injury. PMID:26018184

  10. Endothelial Progenitor Cell Fraction Contained in Bone Marrow-Derived Mesenchymal Stem Cell Populations Impairs Osteogenic Differentiation

    PubMed Central

    Duttenhoefer, Fabian; Lara de Freitas, Rafael; Loibl, Markus; Bittermann, Gido; Geoff Richards, R.; Alini, Mauro; Verrier, Sophie

    2015-01-01

    In bone tissue engineering (TE) endothelial cell-osteoblast cocultures are known to induce synergies of cell differentiation and activity. Bone marrow mononucleated cells (BMCs) are a rich source of mesenchymal stem cells (MSCs) able to develop an osteogenic phenotype. Endothelial progenitor cells (EPCs) are also present within BMC. In this study we investigate the effect of EPCs present in the BMC population on MSCs osteogenic differentiation. Human BMCs were isolated and separated into two populations. The MSC population was selected through plastic adhesion capacity. EPCs (CD34+ and CD133+) were removed from the BMC population and the resulting population was named depleted MSCs. Both populations were cultured over 28 days in osteogenic medium (Dex+) or medium containing platelet lysate (PL). MSC population grew faster than depleted MSCs in both media, and PL containing medium accelerated the proliferation for both populations. Cell differentiation was much higher in Dex+ medium in both cases. Real-time RT-PCR revealed upregulation of osteogenic marker genes in depleted MSCs. Higher values of ALP activity and matrix mineralization analyses confirmed these results. Our study advocates that absence of EPCs in the MSC population enables higher osteogenic gene expression and matrix mineralization and therefore may lead to advanced bone neoformation necessary for TE constructs. PMID:26491682

  11. The matrix protein CCN1 (CYR61) promotes proliferation, migration and tube formation of endothelial progenitor cells

    SciTech Connect

    Yu Yang; Gao Yu; Wang, Hong; Huang Lan Qin Jun; Guo Ruiwei; Song Mingbao; Yu Shiyong; Chen Jianfei; Cui Bin; Gao Pan

    2008-10-15

    Neovascularization and re-endothelialization relies on circulating endothelial progenitor cells (EPCs), but their recruitment and angiogenic roles are subjected to regulation by the vascular microenvironment, which remains largely unknown. The present study was designed to investigate the effects of mature ECs and matrix protein CCN1 on the properties of EPCs. In a coculture system, effects of ECs on proliferation, migration and participation in tube-like formation of EPCs were evaluated, and functional assays were employed to identify the exact role of CCN1 in EPCs vitality and function. We demonstrated that ECs, as an indispensable part of the cellular milieu, significantly promoted the proliferation, migration and tube formation activities of EPCs, and more importantly, CCN1 was potentially involved in such effects of ECs. Expression of CCN1 in EPCs was significantly increased by serum, VEGF, ECs-cocultivation and ECs conditioned medium. Moreover, Ad-CCN1-mediated overexpression of CCN1 directly enhanced migration and tube formation of EPCs, whereas silencing of endogenous CCN1 in EPCs inhibits cell functions. Furthermore, CCN1 induced the expressions of chemokines and growth factors, such as MCP-1 and VEGF, suggesting a complex interaction between those proangiogenic factors. Our data suggest that matrix protein CCN1 may play an important role in microenvironment-mediated biological properties of EPCs.

  12. Silencing stromal interaction molecule 1 by RNA interference inhibits the proliferation and migration of endothelial progenitor cells

    SciTech Connect

    Kuang, Chun-yan; Yu, Yang; Guo, Rui-wei; Qian, De-hui; Wang, Kui; Den, Meng-yang; Shi, Yan-kun; Huang, Lan

    2010-07-23

    Research highlights: {yields} STIM1 and TRPC1 are expressed in EPCs. {yields} Knockdown of STIM1 inhibits the proliferation, migration and SOCE of EPCs. {yields} TRPC1-SOC cooperates with STIM1 to mediate the SOCE of EPCs. -- Abstract: Knockdown of stromal interaction molecule 1 (STIM1) significantly suppresses neointima hyperplasia after vascular injury. Endothelial progenitor cells (EPCs) are the major source of cells that respond to endothelium repair and contribute to re-endothelialization by reducing neointima formation after vascular injury. We hypothesized that the effect of STIM1 on neointima hyperplasia inhibition is mediated through its effect on the biological properties of EPCs. In this study, we investigated the effects of STIM1 on the proliferation and migration of EPCs and examined the effect of STIM1 knockdown using cultured rat bone marrow-derived EPCs. STIM1 was expressed in EPCs, and knockdown of STIM1 by adenoviral delivery of small interfering RNA (siRNA) significantly suppressed the proliferation and migration of EPCs. Furthermore, STIM1 knockdown decreased store-operated channel entry 48 h after transfection. Replenishment with recombinant human STIM1 reversed the effects of STIM1 knockdown. Our data suggest that the store-operated transient receptor potential canonical 1 channel is involved in regulating the biological properties of EPCs through STIM1. STIM1 is a potent regulator of cell proliferation and migration in rat EPCs and may play an important role in the biological properties of EPCs.

  13. Advanced glycation end products impair function of late endothelial progenitor cells through effects on protein kinase Akt and cyclooxygenase-2

    SciTech Connect

    Chen Qin; Dong Li; Wang Lian; Kang Lina; Xu Biao

    2009-04-03

    Endothelial progenitor cells (EPCs) exhibit impaired function in the context of diabetes, and advanced glycation end products (AGEs), which accumulate in diabetes, may contribute to this. In the present study, we investigated the mechanism by which AGEs impair late EPC function. EPCs from human umbilical cord blood were isolated, and incubated with AGE-modified albumin (AGE-albumin) at different concentrations found physiologically in plasma. Apoptosis, migration, and tube formation assays were used to evaluate EPC function including capacity for vasculogenesis, and expression of the receptor for AGEs (RAGE), Akt, endothelial nitric oxide synthase (eNOS), and cycloxygenase-2 (COX-2) were determined. Anti-RAGE antibody was used to block RAGE function. AGE-albumin concentration-dependently enhanced apoptosis and depressed migration and tube formation, but did not affect proliferation, of late EPCs. High AGE-albumin increased RAGE mRNA and protein expression, and decreased Akt and COX-2 protein expression, whilst having no effect on eNOS mRNA or protein in these cells. These effects were inhibited by co-incubation with anti-RAGE antibody. These results suggest that RAGE mediates the AGE-induced impairment of late EPC function, through down-regulation of Akt and COX-2 in these cells.

  14. Suppression of oxidative stress in endothelial progenitor cells promotes angiogenesis and improves cardiac function following myocardial infarction in diabetic mice

    PubMed Central

    JIN, PENG; LI, TAO; LI, XUEQI; SHEN, XINGHUA; ZHAO, YANRU

    2016-01-01

    Myocardial infarction is a major contributor to morbidity and mortality in diabetes, which is characterized by inadequate angiogenesis and consequent poor blood reperfusion in the diabetic ischemic heart. The aim of the present study was to investigate the effect that oxidative stress in endothelial progenitor cells (EPCs) has on cardiac angiogenesis in diabetic mice. EPCs derived from diabetic mice revealed reductions in superoxide dismutase (SOD) expression levels and activity compared with those from normal mice. An endothelial tube formation assay showed that angiogenesis was markedly delayed for diabetic EPCs, compared with normal controls. EPCs subjected to various pretreatments were tested as a cell therapy in a diabetic mouse model of myocardial infarction. Induction of oxidative stress in normal EPCs by H2O2 or small interfering RNA-mediated knockdown of SOD reduced their angiogenic activity in the ischemic myocardium of the diabetic mice. Conversely, cell therapy using EPCs from diabetic mice following SOD gene overexpression or treatment with the antioxidant Tempol normalized their ability to promote angiogenesis. These results indicate that decreased expression levels of SOD in EPCs contribute to impaired angiogenesis. In addition, normalization of diabetic EPCs by ex vivo SOD gene therapy accelerates the ability of the EPCs to promote angiogenesis and improve cardiac function when used as a cell therapy following myocardial infarction in diabetic mice. PMID:27284297

  15. Ultrasound Microbubble-Mediated Delivery of Integrin-Linked Kinase Gene Improves Endothelial Progenitor Cells Dysfunction in Pre-Eclampsia

    PubMed Central

    Cui, Kai; Yan, Ting; Luo, Qingqing; Zheng, Yanfang; Liu, Xiaoxia; Huang, Xiaoyu

    2014-01-01

    Pre-eclampsia (PE) is a specific vascular complication in pregnancy whose precise mechanism is still unclear. We hypothesized that endothelial progenitor cells (EPCs), the precursor of endothelial cells, might be impaired in patients with PE and hold a great promise for the treatment of PE. In the present study, we analyzed the EPCs number and expression of integrin-linked kinase (ILK) in PE patients. We confirmed that both EPCs number and ILK expression were diminished in PE patients. Next, we transfected EPCs with ILK gene using ultrasonic microbubble technique (UMT) for the first time, as UMT is a novel type of gene transfer technology showing promising applications in stem cells apart from EPCs. To further investigate the transfection efficiency of UMT, RT-PCR analysis and western blot were used to examine the messenger RNA (mRNA) and protein level of ILK. After transfection of the ILK gene, EPCs function was tested to illustrate the role of ILK in cell proliferation, apoptosis, migration, and secretion. The results of the in vitro study suggested that UMT, a novel gene delivery system, could be considered a potent physical method for EPCs transfection. Moreover, the growth and angiogenetic properties of EPCs are enhanced by introducing ILK. This study may afford a new trend for EPCs transfection and gene therapy in PE. PMID:24564279

  16. PPARα Regulates Mobilization and Homing of Endothelial Progenitor Cells Through the HIF-1α/SDF-1 Pathway

    PubMed Central

    Wang, Zhongxiao; Moran, Elizabeth; Ding, Lexi; Cheng, Rui; Xu, Xun; Ma, Jian-xing

    2014-01-01

    Purpose. The mechanism for the antiangiogenic activity of peroxisome proliferator–activated receptor alpha (PPARα) remains incompletely understood. Endothelial progenitor cells (EPC) are known to participate in neovascularization (NV). The purpose of this study was to investigate whether PPARα regulates EPC during retinal NV. Methods. Retinal NV was induced by oxygen-induced retinopathy (OIR). Mice with OIR were injected intraperitoneally with the PPARα agonist fenofibric acid (FA) or with adenovirus expressing PPARα (Ad-PPARα). Flow cytometry was used to quantify circulating and retinal EPC. Serum stromal cell-derived factor 1 (SDF-1) levels were measured by ELISA. Hypoxia was induced in primary human retinal capillary endothelial cells (HRCEC) and mouse brain endothelial cells (MBEC) by CoCl2. Levels of SDF-1 and hypoxia-inducible factor 1 alpha (HIF-1α) were measured by Western blotting. Results. Fenofibric acid and overexpression of PPARα attenuated the increase of circulating and retinal EPC, correlating with suppressed retinal NV in OIR mice at P17. The PPARα knockout enhanced the OIR-induced increase of circulating and retinal EPC. Fenofibric acid decreased retinal HIF-1α and SDF-1 levels as well as serum SDF-1 levels in the OIR model. In HRCEC, PPARα inhibited HIF-1α nuclear translocation and SDF-1 overexpression induced by hypoxia. Further, MBEC from PPARα−/− mice showed more prominent activation of HIF-1α and overexpression of SDF-1 induced by hypoxia, compared with the wild-type (WT) MBEC. PPARα failed to block SDF-1 overexpression induced by a constitutively active mutant of HIF-1α, suggesting that regulation of SDF-1 by PPARα was through blockade of HIF-1α activation. Conclusions. Peroxisome proliferator-activated receptor alpha suppresses ischemia-induced EPC mobilization and homing through inhibition of the HIF-1α/SDF-1 pathway. This represents a novel molecular mechanism for PPARα's antiangiogenic effects. PMID:24845641

  17. 20-HETE Regulates the Angiogenic Functions of Human Endothelial Progenitor Cells and Contributes to Angiogenesis In Vivo

    PubMed Central

    Chen, Li; Ackerman, Rachel; Saleh, Mohamed; Gotlinger, Katherine H.; Kessler, Michael; Mendelowitz, Lawrence G.; Falck, John R.; Arbab, Ali S.; Scicli, A. Guillermo; Schwartzman, Michal L.

    2014-01-01

    Circulating endothelial progenitor cells (EPC) contribute to postnatal neovascularization. We identified the cytochrome P450 4A/F–20-hydroxyeicosatetraenoic acid (CYP4A/F–20-HETE) system as a novel regulator of EPC functions associated with angiogenesis in vitro. Here, we explored cellular mechanisms by which 20-HETE regulates EPC angiogenic functions and assessed its contribution to EPC-mediated angiogenesis in vivo. Results showed that both hypoxia and vascular endothelial growth factor (VEGF) induce CYP4A11 gene and protein expression (the predominant 20-HETE synthases in human EPC), and this is accompanied by an increase in 20-HETE production by ∼1.4- and 1.8-fold, respectively, compared with the control levels. Additional studies demonstrated that 20-HETE and VEGF have a synergistic effect on EPC proliferation, whereas 20-HETE antagonist 20-HEDGE or VEGF-neutralizing antibody negated 20-HETE- or VEGF-induced proliferation, respectively. These findings are consistent with the presence of a positive feedback regulation on EPC proliferation between the 20-HETE and the VEGF pathways. Furthermore, we found that 20-HETE induced EPC adhesion to fibronectin and endothelial cell monolayer by 40 ± 5.6 and 67 ± 10%, respectively, which was accompanied by a rapid induction of very late antigen-4 and chemokine receptor type 4 mRNA and protein expression. Basal and 20-HETE-stimulated increases in adhesion were negated by the inhibition of the CYP4A–20-HETE system. Lastly, EPC increased angiogenesis in vivo by 3.6 ± 0.2-fold using the Matrigel plug angiogenesis assay, and these increases were markedly reduced by the local inhibition of 20-HETE system. These results strengthened the notion that 20-HETE regulates the angiogenic functions of EPC in vitro and EPC-mediated angiogenesis in vivo. PMID:24403517

  18. 20-HETE regulates the angiogenic functions of human endothelial progenitor cells and contributes to angiogenesis in vivo.

    PubMed

    Chen, Li; Ackerman, Rachel; Saleh, Mohamed; Gotlinger, Katherine H; Kessler, Michael; Mendelowitz, Lawrence G; Falck, John R; Arbab, Ali S; Scicli, A Guillermo; Schwartzman, Michal L; Yang, Jing; Guo, Austin M

    2014-03-01

    Circulating endothelial progenitor cells (EPC) contribute to postnatal neovascularization. We identified the cytochrome P450 4A/F-20-hydroxyeicosatetraenoic acid (CYP4A/F-20-HETE) system as a novel regulator of EPC functions associated with angiogenesis in vitro. Here, we explored cellular mechanisms by which 20-HETE regulates EPC angiogenic functions and assessed its contribution to EPC-mediated angiogenesis in vivo. Results showed that both hypoxia and vascular endothelial growth factor (VEGF) induce CYP4A11 gene and protein expression (the predominant 20-HETE synthases in human EPC), and this is accompanied by an increase in 20-HETE production by ~1.4- and 1.8-fold, respectively, compared with the control levels. Additional studies demonstrated that 20-HETE and VEGF have a synergistic effect on EPC proliferation, whereas 20-HETE antagonist 20-HEDGE or VEGF-neutralizing antibody negated 20-HETE- or VEGF-induced proliferation, respectively. These findings are consistent with the presence of a positive feedback regulation on EPC proliferation between the 20-HETE and the VEGF pathways. Furthermore, we found that 20-HETE induced EPC adhesion to fibronectin and endothelial cell monolayer by 40 ± 5.6 and 67 ± 10%, respectively, which was accompanied by a rapid induction of very late antigen-4 and chemokine receptor type 4 mRNA and protein expression. Basal and 20-HETE-stimulated increases in adhesion were negated by the inhibition of the CYP4A-20-HETE system. Lastly, EPC increased angiogenesis in vivo by 3.6 ± 0.2-fold using the Matrigel plug angiogenesis assay, and these increases were markedly reduced by the local inhibition of 20-HETE system. These results strengthened the notion that 20-HETE regulates the angiogenic functions of EPC in vitro and EPC-mediated angiogenesis in vivo. PMID:24403517

  19. IQ Domain GTPase-Activating Protein 1 is Involved in Shear Stress-Induced Progenitor-Derived Endothelial Cell Alignment

    PubMed Central

    Rami, Lila; Auguste, Patrick; Thebaud, Noélie B.; Bareille, Reine; Daculsi, Richard; Ripoche, Jean; Bordenave, Laurence

    2013-01-01

    Shear stress is one of mechanical constraints which are exerted by blood flow on endothelial cells (ECs). To adapt to shear stress, ECs align in the direction of flow through adherens junction (AJ) remodeling. However, mechanisms regulating ECs alignment under shear stress are poorly understood. The scaffold protein IQ domain GTPase activating protein 1 (IQGAP1) is a scaffold protein which couples cell signaling to the actin and microtubule cytoskeletons and is involved in cell migration and adhesion. IQGAP1 also plays a role in AJ organization in epithelial cells. In this study, we investigated the potential IQGAP1 involvement in the endothelial cells alignment under shear stress. Progenitor-derived endothelial cells (PDECs), transfected (or not) with IQGAP1 small interfering RNA, were exposed to a laminar shear stress (1.2 N/m2) and AJ proteins (VE-cadherin and β-catenin) and IQGAP1 were labeled by immunofluorescence. We show that IQGAP1 is essential for ECs alignment under shear stress. We studied the role of IQGAP1 in AJs remodeling of PDECs exposed to shear stress by studying cell localization and IQGAP1 interactions with VE-cadherin and β-catenin by immunofluorescence and Proximity Ligation Assays. In static conditions, IQGAP1 interacts with VE-cadherin but not with β-catenin at the cell membrane. Under shear stress, IQGAP1 lost its interaction from VE-cadherin to β-catenin. This “switch” was concomitant with the loss of β-catenin/VE-cadherin interaction at the cell membrane. This work shows that IQGAP1 is essential to ECs alignment under shear stress and that AJ remodeling represents one of the mechanisms involved. These results provide a new approach to understand ECs alignment under to shear stress. PMID:24278215

  20. Sympathetic predominance is associated with impaired endothelial progenitor cells and tunneling nanotubes in controlled-hypertensive patients.

    PubMed

    de Cavanagh, Elena M V; González, Sergio A; Inserra, Felipe; Forcada, Pedro; Castellaro, Carlos; Chiabaut-Svane, Jorge; Obregón, Sebastián; Casarini, María Jesús; Kempny, Pablo; Kotliar, Carol

    2014-07-15

    Early endothelial progenitor cells (early EPC) and late EPC are involved in endothelial repair and can rescue damaged endothelial cells by transferring organelles through tunneling nanotubes (TNT). In rodents, EPC mobilization from the bone marrow depends on sympathetic nervous system activity. Indirect evidence suggests a relation between autonomic derangements and human EPC mobilization. We aimed at testing whether hypertension-related autonomic imbalances are associated with EPC impairment. Thirty controlled-essential hypertensive patients [systolic blood pressure/diastolic blood pressure = 130(120-137)/85(61-88) mmHg; 81.8% male] and 20 healthy normotensive subjects [114(107-119)/75(64-79) mmHg; 80% male] were studied. Mononuclear cells were cultured on fibronectin- and collagen-coated dishes for early EPC and late EPC, respectively. Low (LF)- and high (HF)-frequency components of short-term heart rate variability were analyzed during a 5-min rest, an expiration/inspiration maneuver, and a Stroop color-word test. Modulations of cardiac sympathetic and parasympathetic activities were evaluated by LF/HF (%) and HF power (ms(2)), respectively. In controlled-hypertensive patients, the numbers of early EPC, early EPC that emitted TNT, late EPC, and late EPC that emitted TNT were 41, 77, 50, and 88% lower than in normotensive subjects (P < 0.008), respectively. In controlled-hypertensive patients, late EPC number was positively associated with cardiac parasympathetic reserve during the expiration/inspiration maneuver (rho = 0.45, P = 0.031) and early EPC with brachial flow-mediated dilation (rho = 0.655; P = 0.049); also, late TNT number was inversely related to cardiac sympathetic response during the stress test (rho = -0.426, P = 0.045). EPC exposure to epinephrine or norepinephrine showed negative dose-response relationships on cell adhesion to fibronectin and collagen; both catecholamines stimulated early EPC growth, but epinephrine inhibited late EPC growth. In

  1. Pattern of endothelial progenitor cells and apoptotic endothelial cell-derived microparticles in chronic heart failure patients with preserved and reduced left ventricular ejection fraction

    PubMed Central

    Berezin, Alexander E.; Kremzer, Alexander A.; Martovitskaya, Yulia V.; Berezina, Tatyana A.; Gromenko, Elena A.

    2016-01-01

    Background Chronic heart failure (HF) remains a leading cause of cardiovascular (CV) mortality and morbidity worldwide. The aim of the study was to investigate whether the pattern of angiogenic endothelial progenitor cells (EPCs) and apoptotic endothelial cell-derived microparticles (EMPs) would be able to differentiate HF with reduced (HFrEF) and preserved (HFpEF) ejection fraction. Methods One hundred sixty four chronic HF subjects met inclusion criteria. Patients with global left ventricular ejection fraction ≥ 50% were categorized as the HFpEF group (n = 79) and those with ≤ 45% as the HFrEF group (n = 85). Therefore, to compare the circulating levels of biological markers 35 control subjects without HF were included in the study. All control individuals were age- and sex-matched chronic HF patients. The serum level of biomarkers was measured at baseline. The flow cytometric technique was used for predictably distinguishing circulating cell subsets depending on expression of CD45, CD34, CD14, Tie-2, and CD309 antigens and determining endothelial cell-derived microparticles. CD31+/annexin V+ was defined as apoptotic endothelial cell-derived MPs, MPs labeled for CD105+ or CD62E+ were determined as MPs produced due to activation of endothelial cells. Results In multivariate logistic regression model T2DM (R2 = 0.26; P = 0.001), obesity (R2 = 0.22; P = 0.001), previous MI (R2 = 0.17; P = 0.012), galectin-3 (R2 = 0.67; P = 0.012), CD31+/annexin V+ EMPs (R2 = 0.11; P = 0.001), NT-proBNP (R2 = 0.11; P = 0.046), CD14+ CD309+ cells (R2 = 0.058; P = 0.001), and CD14+ СD309+ Tie-2+ cells (R2 = 0.044; P = 0.028) were found as independent predictors of HFpEF. Using multivariate Cox-regression analysis adjusted etiology (previous myocardial infarction), cardiovascular risk factors (obesity, type 2 diabetes mellitus) we found that NT-proBNP (OR 1.08; 95% CI = 1.03–1.12; P = 0.001) and CD31+/annexin V+ EMPs to CD14+ CD309

  2. The Secretome of Hydrogel-Coembedded Endothelial Progenitor Cells and Mesenchymal Stem Cells Instructs Macrophage Polarization in Endotoxemia

    PubMed Central

    Zullo, Joseph A.; Nadel, Ellen P.; Rabadi, May M.; Baskind, Matthew J.; Rajdev, Maharshi A.; Demaree, Cameron M.; Vasko, Radovan; Chugh, Savneek S.; Lamba, Rajat; Goligorsky, Michael S.

    2015-01-01

    We previously reported the delivery of endothelial progenitor cells (EPCs) embedded in hyaluronic acid-based (HA)-hydrogels protects renal function during acute kidney injury (AKI) and promotes angiogenesis. We attempted to further ameliorate renal dysfunction by coembedding EPCs with renal mesenchymal stem cells (MSCs), while examining their paracrine influence on cytokine/chemokine release and proinflammatory macrophages. A live/dead assay determined whether EPC-MSC coculturing improved viability during lipopolysaccharide (LPS) treatment, and HA-hydrogel-embedded delivery of cells to LPS-induced AKI mice was assessed for effects on mean arterial pressure (MAP), renal blood flow (RBF), circulating cytokines/chemokines, serum creatinine, proteinuria, and angiogenesis (femoral ligation). Cytokine/chemokine release from embedded stem cells was examined, including effects on macrophage polarization and release of proinflammatory molecules. EPC-MSC coculturing improved stem cell viability during LPS exposure, an effect augmented by MSC hypoxic preconditioning. The delivery of coembedded EPCs with hypoxic preconditioned MSCs to AKI mice demonstrated additive improvement (compared with EPC delivery alone) in medullary RBF and proteinuria, with comparable effects on serum creatinine, MAP, and angiogenesis. Exposure of proinflammatory M1 macrophages to EPC-MSC conditioned medium changed their polarization to anti-inflammatory M2. Incubation of coembedded EPCs-MSCs with macrophages altered their release of cytokines/chemokines, including enhanced release of anti-inflammatory interleukin (IL)-4 and IL-10. EPC-MSC delivery to endotoxemic mice elevated the levels of circulating M2 macrophages and reduced the circulating cytokines/chemokines. In conclusion, coembedding EPCs-MSCs improved their resistance to stress, impelled macrophage polarization from M1 to M2 while altering their cytokine/chemokines release, reduced circulating cytokines/chemokines, and improved renal and

  3. Sustained Release of Engineered Stromal Cell–Derived Factor 1-α From Injectable Hydrogels Effectively Recruits Endothelial Progenitor Cells and Preserves Ventricular Function After Myocardial Infarction

    PubMed Central

    MacArthur, John W.; Purcell, Brendan P.; Shudo, Yasuhiro; Cohen, Jeffrey E.; Fairman, Alex; Trubelja, Alen; Patel, Jay; Hsiao, Philip; Yang, Elaine; Lloyd, Kelsey; Hiesinger, William; Atluri, Pavan; Burdick, Jason A.; Woo, Y. Joseph

    2014-01-01

    Background Exogenously delivered chemokines have enabled neovasculogenic myocardial repair in models of ischemic cardiomyopathy; however, these molecules have short half-lives in vivo. In this study, we hypothesized that the sustained delivery of a synthetic analog of stromal cell–derived factor 1-α (engineered stromal cell–derived factor analog [ESA]) induces continuous homing of endothelial progenitor cells and improves left ventricular function in a rat model of myocardial infarction. Methods and Results Our previously designed ESA peptide was synthesized by the addition of a fluorophore tag for tracking. Hyaluronic acid was chemically modified with hydroxyethyl methacrylate to form hydrolytically degradable hydrogels through free-radical–initiated crosslinking. ESA was encapsulated in hyaluronic acid hydrogels during gel formation, and then ESA release, along with gel degradation, was monitored for more than 4 weeks in vitro. Chemotactic properties of the eluted ESA were assessed at multiple time points using rat endothelial progenitor cells in a transwell migration assay. Finally, adult male Wistar rats (n=33) underwent permanent ligation of the left anterior descending (LAD) coronary artery, and 100 μL of saline, hydrogel alone, or hydrogel+25 μg ESA was injected into the borderzone. ESA fluorescence was monitored in animals for more than 4 weeks, after which vasculogenic, geometric, and functional parameters were assessed to determine the therapeutic benefit of each treatment group. ESA release was sustained for 4 weeks in vitro, remained active, and enhanced endothelial progenitor cell chemotaxis. In addition, ESA was detected in the rat heart >3 weeks when delivered within the hydrogels and significantly improved vascularity, ventricular geometry, ejection fraction, cardiac output, and contractility compared with controls. Conclusions We have developed a hydrogel delivery system that sustains the release of a bioactive endothelial progenitor cell

  4. Severe Type 2 Diabetes Induces Reversible Modifications of Endothelial Progenitor Cells Which are Ameliorate by Glycemic Control

    PubMed Central

    De Pascale, Maria Rosaria; Bruzzese, Giuseppe; Crimi, Ettore; Grimaldi, Vincenzo; Liguori, Antonio; Brongo, Sergio; Barbieri, Michelangela; Picascia, Antonietta; Schiano, Concetta; Sommese, Linda; Ferrara, Nicola; Paolisso, Giuseppe; Napoli, Claudio

    2016-01-01

    Background Circulating endothelial progenitors cells (EPCs) play a critical role in neovascularization and endothelial repair. There is a growing evidence that hyperglycemia related to Diabetes Mellitus (DM) decreases EPC number and function so promoting vascular complications. Aim of the Study This study investigated whether an intensive glycemic control regimen in Type 2 DM can increase the number of EPCs and restores their function. Methods Sixty-two patients with Type 2 DM were studied. Patients were tested at baseline and after 3 months of an intensive regimen of glycemic control. The Type 2 DM group was compared to control group of subjects without diabetes. Patients with Type 2 DM (mean age 58.2±5.4 years, 25.6% women, disease duration of 15.4±6.3 years) had a baseline HgA1c of 8.7±0.5% and lower EPC levels (CD34+/KDR+) in comparison to healthy controls (p<0.01). Results The intensive glycemic control regimen (HgA1c decreased to 6.2±0.3%) was coupled with a significant increase of EPC levels (mean of 18%, p<0.04 vs. baseline) and number of EPCs CFUs (p<0.05 vs. baseline). Conclusion This study confirms that number and bioactivity of EPCs are reduced in patients with Type 2 DM and, most importantly, that the intensive glycemic control in Type 2 DM promotes EPC improvement both in their number and in bioactivity. PMID:27426095

  5. TNF-α increases endothelial progenitor cell adhesion to the endothelium by increasing bond expression and affinity

    PubMed Central

    Prisco, Anthony R.; Prisco, Michael R.; Carlson, Brian E.

    2014-01-01

    Endothelial progenitor cells (EPCs) are a rare population of cells that participate in angiogenesis. To effectively use EPCs for regenerative therapy, the mechanisms by which they participate in tissue repair must be elucidated. This study focused on the process by which activated EPCs bind to a target tissue. It has been demonstrated that EPCs can bind to endothelial cells (ECs) through the tumore necrosis factor-α (TNF-α)-regulated vascular cell adhesion molecule 1/very-late antigen 4 (VLA4) interaction. VLA4 can bind in a high or low affinity state, a process that is difficult to experimentally isolate from bond expression upregulation. To separate these processes, a new parallel plate flow chamber was built, a detachment assay was developed, and a mathematical model was created that was designed to analyze the detachment assay results. The mathematical model was developed to predict the relative expression of EPC/EC bonds made for a given bond affinity distribution. EPCs treated with TNF-α/vehicle were allowed to bind to TNF-α/vehicle-treated ECs in vitro. Bound cells were subjected to laminar flow, and the cellular adherence was quantified as a function of shear stress. Experimental data were fit to the mathematical model using changes in bond expression or affinity as the only free parameter. It was found that TNF-α treatment of ECs increased adhesion through bond upregulation, whereas TNF-α treatment of EPCs increased adhesion by increasing bond affinity. These data suggest that injured tissue could potentially increase recruitment of EPCs for tissue regeneration via the secretion of TNF-α. PMID:25539711

  6. Prominin-1/CD133 expression as potential tissue-resident vascular endothelial progenitor cells in the pulmonary circulation.

    PubMed

    Sekine, Ayumi; Nishiwaki, Tetsu; Nishimura, Rintaro; Kawasaki, Takeshi; Urushibara, Takashi; Suda, Rika; Suzuki, Toshio; Takayanagi, Shin; Terada, Jiro; Sakao, Seiichiro; Tada, Yuji; Iwama, Atsushi; Tatsumi, Koichiro

    2016-06-01

    Pulmonary vascular endothelial cells could contribute to maintain homeostasis in adult lung vasculature. "Tissue-resident" endothelial progenitor cells (EPCs) play pivotal roles in postnatal vasculogenesis, vascular repair, and tissue regeneration; however, their local pulmonary counterparts remain to be defined. To determine whether prominin-1/CD133 expression can be a marker of tissue-resident vascular EPCs in the pulmonary circulation, we examined the origin and characteristics of prominin-1/CD133-positive (Prom1(+)) PVECs considering cell cycle status, viability, histological distribution, and association with pulmonary vascular remodeling. Prom1(+) PVECs exhibited high steady-state transit through the cell cycle compared with Prom1(-) PVECs and exhibited homeostatic cell division as assessed using the label dilution method and mice expressing green fluorescent protein. In addition, Prom1(+) PVECs showed more marked expression of putative EPC markers and drug resistance genes as well as highly increased activation of aldehyde dehydrogenase compared with Prom1(-) PVECs. Bone marrow reconstitution demonstrated that tissue-resident cells were the source of >98% of Prom1(+) PVECs. Immunofluorescence analyses revealed that Prom1(+) PVECs preferentially resided in the arterial vasculature, including the resistant vessels of the lung. The number of Prom1(+) PVECs was higher in developing postnatal lungs. Sorted Prom1(+) PVECs gave rise to colonies and formed fine vascular networks compared with Prom1(-) PVECs. Moreover, Prom1(+) PVECs increased in the monocrotaline and the Su-5416 + hypoxia experimental models of pulmonary vascular remodeling. Our findings indicated that Prom1(+) PVECs exhibited the phenotype of tissue-resident EPCs. The unique biological characteristics of Prom1(+) PVECs predominantly contribute to neovasculogenesis and maintenance of homeostasis in pulmonary vascular tissues. PMID:27059286

  7. Quercitrin treatment protects endothelial progenitor cells from oxidative damage via inducing autophagy through extracellular signal-regulated kinase.

    PubMed

    Zhi, Kangkang; Li, Maoquan; Bai, Jun; Wu, Yongfa; Zhou, Sili; Zhang, Xiaoping; Qu, Lefeng

    2016-07-01

    Atherosclerosis is a disease resulting from impaired endothelial function, often caused by oxidant injury or inflammation. Endothelial progenitor cells (EPCs) play a critical role in repairing damaged endothelium and protecting against atherosclerosis. Quercitrin, a plant-derived flavonoid compound, displays antioxidant and anti-inflammatory activities. In this study, we showed that quercitrin treatment reduced the apoptosis of EPCs caused by oxidized low-density lipoprotein (ox-LDL) in a dose-dependent manner. Quercitrin improved tube formation, migration and adhesion of ox-LDL-treated EPCs. To determine the effect of quercitrin in vivo, EPCs treated with or without ox-LDL and quercitrin were locally injected into the ischemic hind limb muscle of nude mice. Those injected with EPCs treated with ox-LDL and quercitrin showed significantly increased local accumulation of EPCs, blood flow recovery and capillary density compared with the control and ox-LDL only groups. Furthermore, we showed that quercitrin enhanced autophagy and upregulated mitogen-activated protein kinase and ERK phosphorylation in a dose-dependent manner in vitro. Autophagy inhibitors, chloroquine and 3-methyladenine, abrogated quercitrin-enhanced autophagy caused by ox-LDL as evidenced by decreased numbers of branch points, migratory cells and adherent cells, and increased numbers of apoptotic cells. The ERK inhibitor PD98059 abrogated quercitrin-enhanced autophagy, as identified by decreased autophagosome formation and downregulated ERK phosphorylation. The inhibition of ERK did not affect the expression of Rac1, but enhanced phosphorylation of Akt. Quercitrin treatment also increased the expression of E-cadherin, and PD98059 abrogated the upregulation of E-cadherin induced by quercitrin. Our findings suggested that autophagy is a protective mechanism in EPCs exposed to oxidative damage. Quercitrin can promote autophagy through the activation of ERK and the ERK signaling pathway is therefore

  8. GroEL1, a Heat Shock protein 60 of Chlamydia pneumoniae, Impairs Neovascularization by Decreasing Endothelial Progenitor Cell Function

    PubMed Central

    Lin, Yi-Wen; Huang, Chun-Yao; Chen, Yung-Hsiang; Shih, Chun-Ming; Tsao, Nai-Wen; Lin, Cheng-Yen; Chang, Nen-Chung; Tsai, Chien-Sung; Tsai, Hsiao-Ya; Tsai, Jui-Chi; Huang, Po-Hsun; Li, Chi-Yuan; Lin, Feng-Yen

    2013-01-01

    The number and function of endothelial progenitor cells (EPCs) are sensitive to hyperglycemia, hypertension, and smoking in humans, which are also associated with the development of atherosclerosis. GroEL1 from Chlamydia pneumoniae has been found in atherosclerotic lesions and is related to atherosclerotic pathogenesis. However, the actual effects of GroEL1 on EPC function are unclear. In this study, we investigate the EPC function in GroEL1-administered hind limb-ischemic C57BL/B6 and C57BL/10ScNJ (a toll-like receptor 4 (TLR4) mutation) mice and human EPCs. In mice, laser Doppler imaging, flow cytometry, and immunohistochemistry were used to evaluate the degree of neo-vasculogenesis, circulating level of EPCs, and expression of CD34, vWF, and endothelial nitric oxide synthase (eNOS) in vessels. Blood flow in the ischemic limb was significantly impaired in C57BL/B6 but not C57BL/10ScNJ mice treated with GroEL1. Circulating EPCs were also decreased after GroEL1 administration in C57BL/B6 mice. Additionally, GroEL1 inhibited the expression of CD34 and eNOS in C57BL/B6 ischemic muscle. In vitro, GroEL1 impaired the capacity of differentiation, mobilization, tube formation, and migration of EPCs. GroEL1 increased senescence, which was mediated by caspases, p38 MAPK, and ERK1/2 signaling in EPCs. Furthermore, GroEL1 decreased integrin and E-selectin expression and induced inflammatory responses in EPCs. In conclusion, these findings suggest that TLR4 and impaired NO-related mechanisms could contribute to the reduced number and functional activity of EPCs in the presence of GroEL1 from C. pneumoniae. PMID:24376840

  9. Advanced glycation endproducts induce apoptosis of endothelial progenitor cells by activating receptor RAGE and NADPH oxidase/JNK signaling axis

    PubMed Central

    Chen, Jianfei; Jing, Jun; Yu, Shiyong; Song, Minbao; Tan, Hu; Cui, Bin; Huang, Lan

    2016-01-01

    Elevated levels of advanced glycation endproducts (AGEs) is an important risk factor for atherosclerosis. Dysfunction of endothelial progenitor cells (EPCs), which is essential for re-endothelialization and neovascularization, is a hallmark of atherosclerosis. However, it remains unclear whether and how AGEs acts on EPCs to promote pathogenesis of atherosclerosis. In this study, EPCs were exposed to different concentrations of AGEs. The expression of NADPH and Rac1 was measured to investigate the involvement of NADPH oxidase pathway. ROS was examined to indicate the level of oxidative stress in EPCs. Total JNK and p-JNK were determined by Western blotting. Cell apoptosis was evaluated by both TUNEL staining and flow cytometry. Cell proliferation was measured by 3H thymidine uptake. The results showed that treatment of EPCs with AGEs increased the levels of ROS in EPCs. Mechanistically, AGEs increased the activity of NADPH oxidase and the expression of Rac1, a major component of NADPH. Importantly, treatment of EPCs with AGEs activated the JNK signaling pathway, which was closely associated with cell apoptosis and inhibition of proliferation. Our results suggest that the RAGE activation by AGEs in EPCs upregulates intracellular ROS levels, which contributes to increased activity of NADPH oxidase and expression of Rac1, thus promoting cellular apoptosis and inhibiting proliferation. Mechanistically, AGEs binding to the receptor RAGE in EPCs is associated with hyperactivity of JNK signaling pathway, which is downstream of ROS. Our findings suggest that dysregulation of the AGEs/RAGE axis in EPCs may promote atherosclerosis and identify the NADPH/ROS/JNK signaling axis as a potential target for therapeutic intervention. PMID:27347324

  10. Oxidized High-Density Lipoprotein Impairs Endothelial Progenitor Cells' Function by Activation of CD36-MAPK-TSP-1 Pathways

    PubMed Central

    Wu, Jianxiang; He, Zhiqing; Gao, Xiang; Wu, Feng; Ding, Ru; Ren, Yusheng; Jiang, Qijun; Fan, Min

    2015-01-01

    Abstract Aims: High-density lipoprotein (HDL) levels inversely correlate with cardiovascular events due to the protective effects on vascular wall and stem cells, which are susceptible to oxidative modifications and then lead to potential pro-atherosclerotic effects. We proposed that oxidized HDL (ox-HDL) might lead to endothelial progenitor cells (EPCs) dysfunction and investigated underlying mechanisms. Results: ox-HDL was shown to increase apoptosis and intracellular reactive oxygen species levels, but to reduce migration, angiogenesis, and cholesterol efflux of EPCs in a dose-dependent manner. p38 mitogen-activated protein kinase (MAPK) and NF-κB were activated after ox-HDL stimulation, which also upregulated thrombospondin-1 (TSP-1) expression without affecting vascular endothelial growth factor. Effects caused by ox-HDL could be significantly attenuated by pretreatment with short hairpin RNA-mediated CD36 knockdown or probucol. Data of in vivo experiments and the inverse correlation of ox-HDL and circulating EPC numbers among patients with coronary artery diseases (CAD) or CAD and type 2 diabetes also supported it. Meanwhile, HDL separated from such patients could significantly increase cultured EPC's caspase 3 activity, further supporting our proposal. Innovation: This is the most complete study to date of how ox-HDL would impair EPCs function, which was involved with activation of CD36-p38 MAPK-TSP-1 pathways and proved by not only the inverse relationship between ox-HDL and circulating EPCs in clinic but also pro-apoptotic effects of HDL separated from patients' serum. Conclusion: Activation of CD36-p38 MAPK-TSP-1 pathways contributes to the pathological effects of ox-HDL on EPCs' dysfunction, which might be one of the potential etiological factors responsible for the disturbed neovascularization in chronic ischemic disease. Antioxid. Redox Signal. 22, 308–324. PMID:25313537

  11. Exposure to Inhaled Nickel Nanoparticles Causes a Reduction in Number and Function of Bone Marrow Endothelial Progenitor Cells

    PubMed Central

    Liberda, Eric N; Cuevas, Azita K; Gillespie, Patricia A; Grunig, Gabriele; Qu, Qingshan; Chen, Lung Chi

    2016-01-01

    Introduction Particulate matter (PM), specifically nickel (Ni) found on or in PM, has been associated with an increased risk of mortality in human population studies and significant increases in vascular inflammation, generation of reactive oxygen species, altered vasomotor tone, and potentiated atherosclerosis in murine exposures. Recently, murine inhalation of Ni nanoparticles have been shown to cause pulmonary inflammation which affects cardiovascular tissue and potentiates atherosclerosis. These adverse cardiovascular outcomes may be due to the effects of Ni on endothelial progenitor cells (EPCs), endogenous semi-pluripotent stem cells that aid in endothelial repair. Thus, we hypothesize that Ni nanoparticle exposures decrease cell count and cause impairments in function which may ultimately have significant effects on various cardiovascular diseases such as atherosclerosis. Methods Experiments involving inhaled Ni nanoparticle exposures(2 days/5 hrs/day at ~1000 μg/m3, 3 days/5 hrs/day at ~1000 μg/m3, and 5days/5 hrs/day at ~100 μg/m3), were performed in order to quantify bone marrow resident EPCs using flow cytometry in C57BL/6 mice. Plasma levels of SDF-1α and VEGF were assessed by ELISA and in vitro functional assessments of cultured EPCs were conducted. Results and Conclusions Significant EPC count differences between exposure and control groups for Ni nanoparticle exposures were observed. Differences in EPC tube formation and chemotaxis were also observed for the Ni nanoparticle exposed group. Plasma VEGF and SDF-1α differences were not statistically significant. In conclusion, this study shows that inhalation of Ni nanoparticles results in functionally impaired EPCs and reduced number in the bone marrow, which may lead to enhanced progression of atherosclerosis. PMID:20936915

  12. Enhanced Expression of Stim, Orai, and TRPC Transcripts and Proteins in Endothelial Progenitor Cells Isolated from Patients with Primary Myelofibrosis

    PubMed Central

    Dragoni, Silvia; Laforenza, Umberto; Bonetti, Elisa; Reforgiato, Marta; Poletto, Valentina; Lodola, Francesco; Bottino, Cinzia; Guido, Daniele; Rappa, Alessandra; Pareek, Sumedha; Tomasello, Mario; Guarrera, Maria Rosa; Cinelli, Maria Pia; Aronica, Adele; Guerra, Germano; Barosi, Giovanni; Tanzi, Franco

    2014-01-01

    Background An increase in the frequency of circulating endothelial colony forming cells (ECFCs), the only subset of endothelial progenitor cells (EPCs) truly belonging to the endothelial phenotype, occurs in patients affected by primary myelofibrosis (PMF). Herein, they might contribute to the enhanced neovascularisation of fibrotic bone marrow and spleen. Store-operated Ca2+ entry (SOCE) activated by the depletion of the inositol-1,4,5-trisphosphate (InsP3)-sensitive Ca2+ store drives proliferation in ECFCs isolated from both healthy donors (N-ECFCs) and subjects suffering from renal cellular carcinoma (RCC-ECFCs). SOCE is up-regulated in RCC-ECFCs due to the over-expression of its underlying molecular components, namely Stim1, Orai1, and TRPC1. Methodology/Principal Findings We utilized Ca2+ imaging, real-time polymerase chain reaction, western blot analysis and functional assays to evaluate molecular structure and the functional role of SOCE in ECFCs derived from PMF patients (PMF-ECFCs). SOCE, induced by either pharmacological (i.e. cyclopiazonic acid or CPA) or physiological (i.e. ATP) stimulation, was significantly higher in PMF-ECFCs. ATP-induced SOCE was inhibited upon blockade of the phospholipase C/InsP3 signalling pathway with U73111 and 2-APB. The higher amplitude of SOCE was associated to the over-expression of the transcripts encoding for Stim2, Orai2–3, and TRPC1. Conversely, immunoblotting revealed that Stim2 levels remained constant as compared to N-ECFCs, while Stim1, Orai1, Orai3, TRPC1 and TRPC4 proteins were over-expressed in PMF-ECFCs. ATP-induced SOCE was inhibited by BTP-2 and low micromolar La3+ and Gd3+, while CPA-elicited SOCE was insensitive to Gd3+. Finally, BTP-2 and La3+ weakly blocked PMF-ECFC proliferation, while Gd3+ was ineffective. Conclusions Two distinct signalling pathways mediate SOCE in PMF-ECFCs; one is activated by passive store depletion and is Gd3+-resistant, while the other one is regulated by the InsP3-sensitive Ca2

  13. Construction of a multifunctional coating consisting of phospholipids and endothelial progenitor cell-specific peptides on titanium substrates

    NASA Astrophysics Data System (ADS)

    Chen, Huiqing; Li, Xiaojing; Zhao, Yuancong; Li, Jingan; Chen, Jiang; Yang, Ping; Maitz, Manfred F.; Huang, Nan

    2015-08-01

    A phospholipid/peptide polymer (PMMDP) with phosphorylcholine groups, endothelial progenitor cell (EPC)-specific peptides and catechol groups was anchored onto a titanium (Ti) surface to fabricate a biomimetic multifunctional surface. The PMMDP coating was characterized by X-ray photoelectron spectroscopy (XPS), water contact angle measurements and atomic force microscopy (AFM), respectively. The amount of PMMDP coating on the Ti surface was quantified by using the quartz crystal microbalance with dissipation (QCM-D). Interactions between blood components and the coated and bare Ti substrates were evaluated by platelet adhesion and activation assays and fibrinogen denaturation test using platelet rich plasma (PRP). The results revealed that the PMMDP-modified surface inhibited fibrinogen denaturation and reduced platelet adhesion and activation. EPC cell culture on the PMMDP-modified surface showed increased adhesion and proliferation of EPCs when compared to the cells cultured on untreated Ti surface. The inhibition of fibrinogen denaturation and platelet adhesion and support of EPCs attachment and proliferation indicated that this coating might be beneficial for future applications in blood-contacting implants, such as vascular stents.

  14. Melanoma cell therapy: Endothelial progenitor cells as shuttle of the MMP12 uPAR-degrading enzyme

    PubMed Central

    Laurenzana, Anna; Biagioni, Alessio; D'Alessio, Silvia; Bianchini, Francesca; Chillà, Anastasia; Margheri, Francesca; Luciani, Cristina; Mazzanti, Benedetta; Pimpinelli, Nicola; Torre, Eugenio; Danese, Silvio; Calorini, Lido; Rosso, Mario Del; Fibbi, Gabriella

    2014-01-01

    The receptor for the urokinase-type plasminogen activator (uPAR) accounts for many features of cancer progression, and is therefore considered a target for anti-tumoral therapy. Only full length uPAR mediates tumor progression. Matrix-metallo-proteinase-12 (MMP12)-dependent uPAR cleavage results into the loss of invasion properties and angiogenesis. MMP12 can be employed in the field of “targeted therapies” as a biological drug to be delivered directly in patient's tumor mass. Endothelial Progenitor Cells (EPCs) are selectively recruited within the tumor and could be used as cellular vehicles for delivering anti-cancer molecules. The aim of our study is to inhibit cancer progression by engeneering ECFCs, a subset of EPC, with a lentivirus encoding the anti-tumor uPAR-degrading enzyme MMP12. Ex vivo manipulated ECFCs lost the capacity to perform capillary morphogenesis and acquired the anti-tumor and anti-angiogenetic activity. In vivo MMP12-engineered ECFCs cleaved uPAR within the tumor mass and strongly inhibited tumor growth, tumor angiogenesis and development of lung metastasis. The possibility to exploit tumor homing and activity of autologous MMP12-engineered ECFCs represents a novel way to combat melanoma by a “personalized therapy”, without rejection risk. The i.v. injection of radiolabelled MMP12-ECFCs can thus provide a new theranostic approach to control melanoma progression and metastasis. PMID:25003596

  15. Melanoma cell therapy: Endothelial progenitor cells as shuttle of the MMP12 uPAR-degrading enzyme.

    PubMed

    Laurenzana, Anna; Biagioni, Alessio; D'Alessio, Silvia; Bianchini, Francesca; Chillà, Anastasia; Margheri, Francesca; Luciani, Cristina; Mazzanti, Benedetta; Pimpinelli, Nicola; Torre, Eugenio; Danese, Silvio; Calorini, Lido; Del Rosso, Mario; Fibbi, Gabriella

    2014-06-15

    The receptor for the urokinase-type plasminogen activator (uPAR) accounts for many features of cancer progression, and is therefore considered a target for anti-tumoral therapy. Only full length uPAR mediates tumor progression. Matrix-metallo-proteinase-12 (MMP12)-dependent uPAR cleavage results into the loss of invasion properties and angiogenesis. MMP12 can be employed in the field of "targeted therapies" as a biological drug to be delivered directly in patient's tumor mass. Endothelial Progenitor Cells (EPCs) are selectively recruited within the tumor and could be used as cellular vehicles for delivering anti-cancer molecules. The aim of our study is to inhibit cancer progression by engeneering ECFCs, a subset of EPC, with a lentivirus encoding the anti-tumor uPAR-degrading enzyme MMP12. Ex vivo manipulated ECFCs lost the capacity to perform capillary morphogenesis and acquired the anti-tumor and anti-angiogenetic activity. In vivo MMP12-engineered ECFCs cleaved uPAR within the tumor mass and strongly inhibited tumor growth, tumor angiogenesis and development of lung metastasis. The possibility to exploit tumor homing and activity of autologous MMP12-engineered ECFCs represents a novel way to combat melanoma by a "personalized therapy", without rejection risk. The i.v. injection of radiolabelled MMP12-ECFCs can thus provide a new theranostic approach to control melanoma progression and metastasis. PMID:25003596

  16. Evaluation of the circulating CD34+, CD309+, and endothelial progenitor cells in patients with first attack of optic neuritis

    PubMed Central

    Etemadifar, Masoud; Dehghani, Leila; Ganji, Hamid; Soleimani, Reza; Talebi, Maedeh; Eskandari, Nahid; Samani, Fazel Sahraneshin; Meamar, Rokhsareh

    2015-01-01

    Background: Endothelial progenitor cells (EPCs) are present in circulation and contribute to vasculogenesis in adults. The aim of the present study was to determine the number of circulating EPCs in patients with optic neuritis (ON). Materials and Methods: Fifty patients with ON were diagnosed by expert neurologist and optometrist at the Feiz Hospital, Isfahan, Iran (2012–2013). Blood samples were collected from ON patients in the first attack. The number of EPCs was measured by flow cytometry through the assessment of CD34+ and CD309+ in patients and healthy individuals. Results: With using flow cytometry, CD34+ and CD309+ cells detected in peripheral blood cells of patients (n = 50) with ON, and healthy individuals (n = 30). Patients with ON had (mean = 66.71 ± 17.82) CD34+ and CD309+ cells compared with healthy controls (mean = 28.72 ± 22.46). In addition, there was no significant difference in CD309+ cells in both groups. Conclusion: This study showed elevated CD34+ and CD309+ cells in the early stage of the disease. Regarded to EPC increment in neural repair, it expected the EPC level be increased in these patients, but no detectable differences were observed among both markers in healthy and patient with first attack. PMID:26380236

  17. The biphasic effects of oxidized-low density lipoprotein on the vasculogenic function of endothelial progenitor cells.

    PubMed

    Lin, Feng-Yen; Tsao, Nai-Wen; Shih, Chun-Ming; Lin, Yi-Wen; Yeh, Jong-Shiua; Chen, Jaw-Wen; Nakagami, Hironori; Morishita, Ryuichi; Sawamura, Tatsuya; Huang, Chun-Yao

    2015-01-01

    Late-outgrowth endothelial progenitor cells (EPCs) are stress-resistant and responsible for reparative functions in the cardiovascular system. Oxidized-LDL (oxLDL) plays a critical role in cardiovascular disease pathogenesis. However, it is largely unknown what the impacts of oxLDL are on late-outgrowth EPCs. This study aimed to investigate the concentration-related effects of oxLDL on EPC functions and related angiogenesis, in vitro and in vivo. In this study, early and late-outgrowth EPCs were generated from circulating human mononuclear cells. oxLDL may regulate EPC vasculogenic function via the lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1). Lower concentrations (5 μg/mL) of oxLDL can potentiate EPC tube formation in vitro and in vivo by activating eNOS mechanisms, which are mediated by p38 MAPK- and SAPK/JNK-related pathways. Higher concentrations of oxLDL (10-50 μg/mL) impaired EPC function via the activation of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase pathways and consequent inhibition of eNOS activity, which could be reversed by anti-oxidants (diphenylene iodonium and apocynin) and gp91phox siRNA. In conclusion, oxLDL has concentration-dependent biphasic effects on human late-outgrowth EPC tube formation in vitro and in vivo. PMID:26017136

  18. Exogenous hTERT gene transfected endothelial progenitor cells from bone marrow promoted angiogenesis in ischemic myocardium of rats

    PubMed Central

    Li, Shang-Hai; Wang, Dan-Dan; Xu, Yun-Jun; Ma, Guo-Dong; Li, Xing-Yue; Liang, Wei-Jun

    2015-01-01

    Objective: To explore the biological behavior and the revascularizative ability of endothelial progenitor cells (EPCs) transfected with human telomerase reverse transcriptase (hTERT) gene. Methods: EPCs were isolated from mononuclear cells in bone marrow by using the method of density gradient centrifugation, then cultured with differential velocity adherent method, EPCs were transfected by recombinant plasmid carrying GFP report gene EGFP-hTERT. The EPCs secretion and proliferation ability were detected before and after transfection. The expression of EPCs mRNA were detected by RT-PCR before and after transfection. The new capillaries of infarct area were observed. Results: After transgenesis, the proliferation of EPCs were increased, and the secretion of NO, LDH, iNOS by EPCs were significantly increased compared to the non-transgenesis group. After transplanted the transfected EPCs into the ischemic myocardial of rats, revascularization were increased obviously. Conclusion: EPCs maintained the original biological characteristics after transfecting exogenous hTER gene, the proliferation and survival rate were up-regulated significantly, and the revascularization ability of EPCs were significantly strengthen. PMID:26550433

  19. Wnt3a is critical for endothelial progenitor cell-mediated neural stem cell proliferation and differentiation.

    PubMed

    Du, Yibin; Zhang, Shuo; Yu, Tao; Du, Gongwen; Zhang, Hui; Yin, Zongsheng

    2016-09-01

    The present study aimed to determine whether co-culture with bone marrow‑derived endothelial progenitor cells (EPCs) affects the proliferation and differentiation of spinal cord-derived neural stem cells (NSCs), and to investigate the underlying mechanism. The proliferation and differentiation of the NSCs were evaluated by an MTT cell proliferation and cytotoxicity assay, and immunofluorescence, respectively. The number of neurospheres and the number of β‑tubulin III‑positive cells were detected by microscopy. The wingless‑type MMTV integration site family, member 3a (Wnt3a)/β-catenin signaling pathway was analyzed by western blot analysis and reverse transcription‑quantitative polymerase chain reaction to elucidate the possible mechanisms of EPC‑mediated NSC proliferation and differentiation. The results revealed that co‑culture with EPCs significantly induced NSC proliferation and differentiation. In addition, co‑culture with EPCs markedly induced the expression levels of Wnt3a and β‑catenin and inhibited the phosphorylation of glycogen synthase kinase 3β (GSK‑3β). By contrast, Wnt3a knockdown using a short hairpin RNA plasmid in the EPCs reduced EPC‑mediated NSC proliferation and differentiation, accompanied by inhibition of the EPC‑mediated expression of β‑catenin, and its phosphorylation and activation of GSK‑3β. Taken together, the findings of the present study demonstrated that Wnt3a was critical for EPC‑mediated NSC proliferation and differentiation. PMID:27484039

  20. Erythropoietin-enhanced endothelial progenitor cell recruitment in peripheral blood and renal vessels during experimental acute kidney injury in rats.

    PubMed

    Cakiroglu, Figen; Enders-Comberg, Sora Maria; Pagel, Horst; Rohwedel, Jürgen; Lehnert, Hendrik; Kramer, Jan

    2016-03-01

    Beneficial effects of erythropoietin (EPO) have been reported in acute kidney injury (AKI) when administered prior to induction of AKI. We studied the effects of EPO administration on renal function shortly after ischemic AKI. For this purpose, rats were subjected to renal ischemia for 30 min and EPO was administered at a concentration of 500 U/kg either i.v. as a single shot directly after ischemia or with an additional i.p. dose until 3 days after surgery. The results were compared with AKI rats without EPO application and a sham-operated group. Renal function was assessed by measurement of serum biochemical markers, histological grading, and using an isolated perfused kidney (IPK) model. Furthermore, we performed flow cytometry to analyze the concentration of endothelial progenitor cells (EPCs) in the peripheral blood and renal vessels. Following EPO application, there was only a statistically non-significant tendency of serum creatinine and urea to improve, particularly after daily EPO application. Renal vascular resistance and the renal perfusion rate were not significantly altered. In the histological analysis, acute tubular necrosis was only marginally ameliorated following EPO administration. In summary, we could not demonstrate a significant improvement in renal function when EPO was applied after AKI. Interestingly, however, EPO treatment resulted in a highly significant increase in CD133- and CD34-positive EPC both in the peripheral blood and renal vessels. PMID:26616141

  1. Multilayer Membranes of Glycosaminoglycans and Collagen I Biomaterials Modulate the Function and Microvesicle Release of Endothelial Progenitor Cells

    PubMed Central

    Dai, Bingyan; Pan, Qunwen; Li, Zhanghua; Zhao, Mingyan; Liao, Xiaorong; Wu, Keng; Ma, Xiaotang

    2016-01-01

    Multilayer composite membrane of biomaterials can increase the function of adipose stem cells or osteoprogenitor cells. Recent evidence indicates endothelial progenitor cells (EPCs) and EPCs released microvesicles (MVs) play important roles in angiogenesis and vascular repair. Here, we investigated the effects of biomaterial multilayer membranes of hyaluronic acid (HA) or chondroitin sulfate (CS) and Collagen I (Col I) on the functions and MVs release of EPCs. Layer-by-layer (LBL) technology was applied to construct the multilayer composite membranes. Four types of the membranes constructed by adsorbing either HA or CS and Col I alternatively with different top layers were studied. The results showed that all four types of multilayer composite membranes could promote EPCs proliferation and migration and inhibit cell senility, apoptosis, and the expression of activated caspase-3. Interestingly, these biomaterials increased the release and the miR-126 level of EPCs-MVs. Moreover, the CS-Col I membrane with CS on the top layer showed the most effects on promoting EPCs proliferation, EPCs-MV release, and miR-126 level in EPCs-MVs. In conclusion, HA/CS and Collagen I composed multilayer composite membranes can promote EPCs functions and release of miR-126 riched EPCs-MVs, which provides a novel strategy for tissue repair treatment. PMID:27190523

  2. The Biphasic Effects of Oxidized-Low Density Lipoprotein on the Vasculogenic Function of Endothelial Progenitor Cells

    PubMed Central

    Lin, Feng-Yen; Tsao, Nai-Wen; Shih, Chun-Ming; Lin, Yi-Wen; Yeh, Jong-Shiua; Chen, Jaw-Wen; Nakagami, Hironori; Morishita, Ryuichi; Sawamura, Tatsuya; Huang, Chun-Yao

    2015-01-01

    Late-outgrowth endothelial progenitor cells (EPCs) are stress-resistant and responsible for reparative functions in the cardiovascular system. Oxidized-LDL (oxLDL) plays a critical role in cardiovascular disease pathogenesis. However, it is largely unknown what the impacts of oxLDL are on late-outgrowth EPCs. This study aimed to investigate the concentration-related effects of oxLDL on EPC functions and related angiogenesis, in vitro and in vivo. In this study, early and late-outgrowth EPCs were generated from circulating human mononuclear cells. oxLDL may regulate EPC vasculogenic function via the lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1). Lower concentrations (5 μg/mL) of oxLDL can potentiate EPC tube formation in vitro and in vivo by activating eNOS mechanisms, which are mediated by p38 MAPK- and SAPK/JNK-related pathways. Higher concentrations of oxLDL (10-50 μg/mL) impaired EPC function via the activation of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase pathways and consequent inhibition of eNOS activity, which could be reversed by anti-oxidants (diphenylene iodonium and apocynin) and gp91phox siRNA. In conclusion, oxLDL has concentration-dependent biphasic effects on human late-outgrowth EPC tube formation in vitro and in vivo. PMID:26017136

  3. Endothelial Cells Mediate Islet-Specific Maturation of Human Embryonic Stem Cell-Derived Pancreatic Progenitor Cells

    PubMed Central

    Jaramillo, Maria; Mathew, Shibin; Mamiya, Hikaru; Goh, Saik Kia

    2015-01-01

    It is well recognized that in vitro differentiation of embryonic stem cells (ESC) can be best achieved by closely recapitulating the in vivo developmental niche. Thus, implementation of directed differentiation strategies has yielded encouraging results in the area of pancreatic islet differentiation. These strategies have concentrated on direct addition of chemical signals, however, other aspect of the developmental niche are yet to be explored. During development, pancreatic progenitor (PP) cells grow as an epithelial sheet, which aggregates with endothelial cells (ECs) during the final stages of maturation. Several findings suggest that the interactions with EC play a role in pancreatic development. In this study, we recapitulated this phenomenon in an in vitro environment by maturing the human ESC (hESC)-derived PP cells in close contact with ECs. We find that co-culture with different ECs (but not fibroblast) alone results in pancreatic islet-specific differentiation of hESC-derived PP cells even in the absence of additional chemical induction. The differentiated cells responded to exogenous glucose levels by enhanced C-peptide synthesis. The co-culture system aligned well with endocrine development as determined by comprehensive analysis of involved signaling pathways. By recapitulating cell–cell interaction aspects of the developmental niche we achieved a differentiation model that aligns closely with islet organogenesis. PMID:24943736

  4. The GroEL protein of Porphyromonas gingivalis accelerates tumor growth by enhancing endothelial progenitor cell function and neovascularization.

    PubMed

    Lin, F-Y; Huang, C-Y; Lu, H-Y; Shih, C-M; Tsao, N-W; Shyue, S-K; Lin, C-Y; Chang, Y-J; Tsai, C-S; Lin, Y-W; Lin, S-J

    2015-06-01

    Porphyromonas gingivalis is a bacterial species that causes destruction of periodontal tissues. Additionally, previous evidence indicates that GroEL from P. gingivalis may possess biological activities involved in systemic inflammation, especially inflammation involved in the progression of periodontal diseases. The literature has established a relationship between periodontal disease and cancer. However, it is unclear whether P. gingivalis GroEL enhances tumor growth. Here, we investigated the effects of P. gingivalis GroEL on neovasculogenesis in C26 carcinoma cell-carrying BALB/c mice and chick eggs in vivo as well as its effect on human endothelial progenitor cells (EPC) in vitro. We found that GroEL treatment accelerated tumor growth (tumor volume and weight) and increased the mortality rate in C26 cell-carrying BALB/c mice. GroEL promoted neovasculogenesis in chicken embryonic allantois and increased the circulating EPC level in BALB/c mice. Furthermore, GroEL effectively stimulated EPC migration and tube formation and increased E-selectin expression, which is mediated by eNOS production and p38 mitogen-activated protein kinase activation. Additionally, GroEL may enhance resistance against paclitaxel-induced cell cytotoxicity and senescence in EPC. In conclusion, P. gingivalis GroEL may act as a potent virulence factor, contributing to the neovasculogenesis of tumor cells and resulting in accelerated tumor growth. PMID:25220060

  5. Tumor Necrosis Factor α Regulates Endothelial Progenitor Cell Migration via CADM1 and NF-kB.

    PubMed

    Prisco, Anthony R; Hoffmann, Brian R; Kaczorowski, Catherine C; McDermott-Roe, Chris; Stodola, Timothy J; Exner, Eric C; Greene, Andrew S

    2016-07-01

    Shortly after the discovery of endothelial progenitor cells (EPCs) in 1997, many clinical trials were conducted using EPCs as a cellular based therapy with the goal of restoring damaged organ function by inducing growth of new blood vessels (angiogenesis). Results were disappointing, largely because the cellular and molecular mechanisms of EPC-induced angiogenesis were not clearly understood. Following injection, EPCs must migrate to the target tissue and engraft prior to induction of angiogenesis. In this study EPC migration was investigated in response to tumor necrosis factor α (TNFα), a pro-inflammatory cytokine, to test the hypothesis that organ damage observed in ischemic diseases induces an inflammatory signal that is important for EPC homing. In this study, EPC migration and incorporation were modeled in vitro using a coculture assay where TNFα treated EPCs were tracked while migrating toward vessel-like structures. It was found that TNFα treatment of EPCs increased migration and incorporation into vessel-like structures. Using a combination of genomic and proteomic approaches, NF-kB mediated upregulation of CADM1 was identified as a mechanism of TNFα induced migration. Inhibition of NF-kB or CADM1 significantly decreased migration of EPCs in vitro suggesting a role for TNFα signaling in EPC homing during tissue repair. Stem Cells 2016;34:1922-1933. PMID:26867147

  6. Wnt3a is critical for endothelial progenitor cell-mediated neural stem cell proliferation and differentiation

    PubMed Central

    Du, Yibin; Zhang, Shuo; Yu, Tao; Du, Gongwen; Zhang, Hui; Yin, Zongsheng

    2016-01-01

    The present study aimed to determine whether co-culture with bone marrow-derived endothelial progenitor cells (EPCs) affects the proliferation and differentiation of spinal cord-derived neural stem cells (NSCs), and to investigate the underlying mechanism. The proliferation and differentiation of the NSCs were evaluated by an MTT cell proliferation and cytotoxicity assay, and immunofluorescence, respectively. The number of neurospheres and the number of β-tubulin III-positive cells were detected by microscopy. The wingless-type MMTV integration site family, member 3a (Wnt3a)/β-catenin signaling pathway was analyzed by western blot analysis and reverse transcription-quantitative polymerase chain reaction to elucidate the possible mechanisms of EPC-mediated NSC proliferation and differentiation. The results revealed that co-culture with EPCs significantly induced NSC proliferation and differentiation. In addition, co-culture with EPCs markedly induced the expression levels of Wnt3a and β-catenin and inhibited the phosphorylation of glycogen synthase kinase 3β (GSK-3β). By contrast, Wnt3a knockdown using a short hairpin RNA plasmid in the EPCs reduced EPC-mediated NSC proliferation and differentiation, accompanied by inhibition of the EPC-mediated expression of β-catenin, and its phosphorylation and activation of GSK-3β. Taken together, the findings of the present study demonstrated that Wnt3a was critical for EPC-mediated NSC proliferation and differentiation. PMID:27484039

  7. Platelet lysate gel and endothelial progenitors stimulate microvascular network formation in vitro: tissue engineering implications

    PubMed Central

    Fortunato, Tiago M.; Beltrami, Cristina; Emanueli, Costanza; De Bank, Paul A.; Pula, Giordano

    2016-01-01

    Revascularisation is a key step for tissue regeneration and complete organ engineering. We describe the generation of human platelet lysate gel (hPLG), an extracellular matrix preparation from human platelets able to support the proliferation of endothelial colony forming cells (ECFCs) in 2D cultures and the formation of a complete microvascular network in vitro in 3D cultures. Existing extracellular matrix preparations require addition of high concentrations of recombinant growth factors and allow only limited formation of capillary-like structures. Additional advantages of our approach over existing extracellular matrices are the absence of any animal product in the composition hPLG and the possibility of obtaining hPLG from patients to generate homologous scaffolds for re-implantation. This discovery has the potential to accelerate the development of regenerative medicine applications based on implantation of microvascular networks expanded ex vivo or the generation of fully vascularised organs. PMID:27141997

  8. Platelet lysate gel and endothelial progenitors stimulate microvascular network formation in vitro: tissue engineering implications.

    PubMed

    Fortunato, Tiago M; Beltrami, Cristina; Emanueli, Costanza; De Bank, Paul A; Pula, Giordano

    2016-01-01

    Revascularisation is a key step for tissue regeneration and complete organ engineering. We describe the generation of human platelet lysate gel (hPLG), an extracellular matrix preparation from human platelets able to support the proliferation of endothelial colony forming cells (ECFCs) in 2D cultures and the formation of a complete microvascular network in vitro in 3D cultures. Existing extracellular matrix preparations require addition of high concentrations of recombinant growth factors and allow only limited formation of capillary-like structures. Additional advantages of our approach over existing extracellular matrices are the absence of any animal product in the composition hPLG and the possibility of obtaining hPLG from patients to generate homologous scaffolds for re-implantation. This discovery has the potential to accelerate the development of regenerative medicine applications based on implantation of microvascular networks expanded ex vivo or the generation of fully vascularised organs. PMID:27141997

  9. Rapid chemotherapy-induced acute endothelial progenitor cell mobilization: implications for antiangiogenic drugs as chemosensitizing agents

    PubMed Central

    Shaked, Yuval; Henke, Erik; Roodhart, Jeanine; Mancuso, Patrizia; Langenberg, Marlies; Colleoni, Marco; Daenen, Laura G.; Man, Shan; Xu, Ping; Emmenegger, Urban; Tang, Terence; Zhu, Zhenping; Witte, Larry; Strieter, Robert M.; Bertolini, Francesco; Voest, Emile; Benezra, Robert; Kerbel, Robert S.

    2008-01-01

    SUMMARY Several hypotheses have been proposed to explain how antiangiogenic drugs enhance the treatment efficacy of cytotoxic chemotherapy including impairing the ability of chemotherapy-responsive tumors to regrow after therapy. With respect to the latter, we show that certain chemotherapy drugs, e.g. paclitaxel, can rapidly induce pro-angiogenic bone marrow derived circulating endothelial cell (CEP) mobilization, and subsequent tumor homing, whereas others, e.g. gemcitabine, did not. Acute CEP mobilization was mediated, at least in part, by systemic induction of SDF-1α and could be prevented by various procedures such as treatment with anti-VEGFR2 blocking antibodies or by paclitaxel treatment in CEP-deficient Id-mutant mice, both of which resulted in enhanced anti-tumor effects mediated by paclitaxel, but not gemcitabine. PMID:18772115

  10. Grain and Bean Lysates Improve Function of Endothelial Progenitor Cells from Human Peripheral Blood: Involvement of the Endogenous Antioxidant Defenses

    PubMed Central

    Lucchesi, Daniela; Russo, Rossella; Gabriele, Morena; Longo, Vincenzo; Del Prato, Stefano; Penno, Giuseppe; Pucci, Laura

    2014-01-01

    Increased oxidative stress contributes to the functional impairment of endothelial progenitor cells (EPCs), the pivotal players in the servicing of the endothelial cell lining. Several evidences suggest that decreasing oxidative stress by natural compounds with antioxidant properties may improve EPCs bioactivity. Here, we investigated the effects of Lisosan G (LG), a Triticum Sativum grain powder, and Lady Joy (LJ), a bean lysate, on function of EPCs exposed to oxidative stress. Peripheral blood mononuclear cells were isolated and plated on fibronectin-coated culture dishes; adherent cells, identified as early EPCs, were pre-treated with different concentrations of LG and LJ and incubated with hydrogen peroxide (H2O2). Viability, senescence, adhesion, ROS production and antioxidant enzymes gene expression were evaluated. Lysate-mediated Nrf-2 (nuclear factor (erythroid-derived 2)-like 2)/ARE (antioxidant response element) activation, a modulator of oxidative stress, was assessed by immunocytochemistry. Lady Joy 0.35–0.7 mg/ml increases EPCs viability; pre-treatment with either LG 0.7 mg/ml and LJ 0.35–0.7 mg/ml protect EPCs viability against H2O2-induced injury. LG 0.7 and LJ 0.35–0.7 mg/ml improve EPCs adhesion; pre-treatment with either LG 0.35 and 0.7 mg/ml or LJ 0.35, 0.7 and 1.4 mg/ml preserve adhesiveness of EPCs exposed to H2O2. Senescence is attenuated in EPCs incubated with lysates 0.35 mg/ml. After exposure to H2O2, LG pre-treated cells show a lower senescence than untreated EPCs. Lysates significantly decrease H2O2-induced ROS generation. Both lysates increase glutathione peroxidase-1 and superoxide dismutase-2 (SOD-2) expression; upon H2O2 exposure, pre-treatment with LJ allows higher SOD-2 expression. Heme oxigenase-1 increases in EPCs pre-treated with LG even upon H2O2 exposure. Finally, incubation with LG 0.7 mg/ml results in Nrf-2 translocation into the nucleus both at baseline and after the oxidative challenge. Our data suggest a protective

  11. Changes in circulating endothelial progenitor cells predict responses of multiple myeloma patients to treatment with bortezomib and dexamethasone

    PubMed Central

    Wang, L.; Du, F.; Zhang, H.M.; Zhang, W.J.; Wang, H.X.

    2015-01-01

    Four cycles of chemotherapy are required to assess responses of multiple myeloma (MM) patients. We investigated whether circulating endothelial progenitor cells (cEPCs) could be a biomarker for predicting patient response in the first cycle of chemotherapy with bortezomib and dexamethasone, so patients might avoid ineffective and costly treatments and reduce exposure to unwanted side effects. We measured cEPCs and stromal cell-derived factor-1α (SDF-1α) in 46 MM patients in the first cycle of treatment with bortezomib and dexamethasone, and investigated clinical relevance based on patient response after four 21-day cycles. The mononuclear cell fraction was analyzed for cEPC by FACS analysis, and SDF-1α was analyzed by ELISA. The study population was divided into 3 groups according to the response to chemotherapy: good responders (n=16), common responders (n=12), and non-responders (n=18). There were no significant differences among these groups at baseline day 1 (P>0.05). cEPC levels decreased slightly at day 21 (8.2±3.3 cEPCs/μL) vs day 1 (8.4±2.9 cEPCs/μL) in good responders (P>0.05). In contrast, cEPC levels increased significantly in the other two groups (P<0.05). SDF-1α changes were closely related to changes in cEPCs. These findings indicate that change in cEPCs at day 21 in the first cycle might be considered a noninvasive biomarker for predicting a later response, and extent of change could help decide whether to continue this costly chemotherapy. cEPCs and the SDF-1α/CXCR4 axis are potential therapeutic targets for improved response and outcomes in MM patients. PMID:26108099

  12. Preconditioned endothelial progenitor cells reduce formation of melanoma metastases through SPARC-driven cell-cell interactions and endocytosis.

    PubMed

    Defresne, Florence; Bouzin, Caroline; Grandjean, Marie; Dieu, Marc; Raes, Martine; Hatzopoulos, Antonis K; Kupatt, Christian; Feron, Olivier

    2011-07-15

    Tumor progression is associated with the release of signaling substances from the primary tumor into the bloodstream. Tumor-derived cytokines are known to promote the mobilization and the recruitment of cells from the bone marrow, including endothelial progenitor cells (EPC). Here, we examined whether such paracrine influence could also influence the capacity of EPC to interfere with circulating metastatic cells. We therefore consecutively injected EPC prestimulated by tumor-conditioned medium (EPC-CM) and luciferase-expressing B16 melanoma cells to mice. A net decrease in metastases spreading (vs. nonstimulated EPC) led us to carry out a 2-dimensional difference gel electrophoresis (2D-DIGE) proteomic study to identify possible mediators of EPC-driven protection. Among 33 proteins exhibiting significant changes in expression, secreted protein, acidic and rich in cysteine (SPARC) presented the highest induction after EPC exposure to CM. We then showed that contrary to control EPC, SPARC-silenced EPC were not able to reduce the extent of metastases when injected with B16 melanoma cells. Using adhesion tests and the hanging drop assay, we further documented that cell-cell interactions between EPC-CM and melanoma cells were promoted in a SPARC-dependent manner. This interaction led to the engulfment of melanoma cells by EPC-CM, a process prevented by SPARC silencing and mimicked by recombinant SPARC. Finally, we showed that contrary to melanoma cells, the prometastatic human breast cancer cell line MDA-MB231-D3H2 reduced SPARC expression in human EPC and stimulated metastases spreading. Our findings unravel the influence of tumor cells on EPC phenotypes through a SPARC-driven accentuation of macrophagic capacity associated with limitations to metastatic spread. PMID:21616936

  13. Interleukin-1β induces fibroblast growth factor 2 expression and subsequently promotes endothelial progenitor cell angiogenesis in chondrocytes.

    PubMed

    Chien, Szu-Yu; Huang, Chun-Yin; Tsai, Chun-Hao; Wang, Shih-Wei; Lin, Yu-Min; Tang, Chih-Hsin

    2016-05-01

    Arthritis is a process of chronic inflammation that results in joint damage. IL (interleukin)-1β is an inflammatory cytokine that acts as a key mediator of cartilage degradation, and is abundantly expressed in arthritis. Neovascularization is one of the pathological characteristics of arthritis. However, the role of IL-1β in the angiogenesis of chondrocytes remains unknown. In the present study, we demonstrate that stimulating chondrocytes (ATDC5) with IL-1β increased the expression of FGF (fibroblast growth factor)-2, a potent angiogenic inducer, and then promoted EPC (endothelial progenitor cell) tube formation and migration. In addition, FGF-2-neutralizing antibody abolished ATDC5-conditional medium-mediated angiogenesis in vitro, as well as its angiogenic effects in the CAM (chick chorioallantoic membrane) assay and Matrigel plug nude mice model in vivo. IHC (immunohistochemistry) staining from a CIA (collagen-induced arthritis) mouse model also demonstrates that arthritis increased the expression of IL-1β and FGF-2, as well as EPC homing in articular cartilage. Moreover, IL-1β-induced FGF-2 expression via IL-1RI (type-1 IL-1 receptor), ROS (reactive oxygen species) generation, AMPK (AMP-activated protein kinase), p38 and NF-κB (nuclear factor κB) pathway has been demonstrated. On the basis of these findings, we conclude that IL-1β promotes FGF-2 expression in chondrocytes through the ROS/AMPK/p38/NF-κB signalling pathway and subsequently increases EPC angiogenesis. Therefore IL-1β serves as a link between inflammation and angiogenesis during arthritis. PMID:26811540

  14. Period 2 is essential to maintain early endothelial progenitor cell function in vitro and angiogenesis after myocardial infarction in mice.

    PubMed

    Sun, Yuan-Yuan; Bai, Wen-Wu; Wang, Bo; Lu, Xiao-Ting; Xing, Yi-Fan; Cheng, Wen; Liu, Xiao-Qiong; Zhao, Yu-Xia

    2014-05-01

    Cellular therapeutic neovascularization has been successfully performed in clinical trials for patients with ischaemia diseases. Despite the vast knowledge of cardiovascular disease and circadian biology, the role of the circadian clock in regulating angiogenesis in myocardial infarction (MI) remains poorly understood. In this study, we aimed to investigate the role and underlying mechanisms of Period 2 (Per2) in endothelial progenitor cell (EPC) function. Flow cytometry revealed lower circulating EPC proportion in per2(-/-) than in wild-type (WT) mice. PER2 was abundantly expressed in early EPCs in mice. In vitro, EPCs from per2(-/-) mice showed impaired proliferation, migration, tube formation and adhesion. Western blot analysis demonstrated inhibited PI3k/Akt/FoxO signalling and reduced C-X-C chemokine receptor type 4 (CXCR4) protein level in EPCs of per2(-/-) mice. The impaired proliferation was blocked by activated PI3K/Akt/FoxO signalling. Direct interaction of CXCR4 and PER2 was detected in WT EPCs. To further study the effect of per2 on in vivo EPC survival and angiogenesis, we injected saline or DiI-labelled WT or per2(-/-) EPC intramyocardially into mice with induced MI. Per2(-/-) reduced the retention of transplanted EPCs in the myocardium, which was associated with significantly reduced DiI expression in the myocardium of MI mice. Decreased angiogenesis in the myocardium of per2(-/-) EPC-treated mice coincided with decreased LV function and increased infarct size in the myocardium. Per2 may be a key factor in maintaining EPC function in vitro and in therapeutic angiogenesis in vivo. PMID:24621388

  15. Insulin sensitizers prevent fine particulate matter-induced vascular insulin resistance and changes in endothelial progenitor cell homeostasis.

    PubMed

    Haberzettl, Petra; McCracken, James P; Bhatnagar, Aruni; Conklin, Daniel J

    2016-06-01

    Exposure to fine particular matter (PM2.5) increases the risk of developing cardiovascular disease and Type 2 diabetes. Because blood vessels are sensitive targets of air pollutant exposure, we examined the effects of concentrated ambient PM2.5 (CAP) on vascular insulin sensitivity and circulating levels of endothelial progenitor cells (EPCs), which reflect cardiovascular health. We found that CAP exposure for 9 days decreased insulin-stimulated Akt phosphorylation in the aorta of mice maintained on control diet. This change was accompanied by the induction of IL-1β and increases in the abundance of cleaved IL-18 and p10 subunit of Casp-1, consistent with the activation of the inflammasome pathway. CAP exposure also suppressed circulating levels of EPCs (Flk-1(+)/Sca-1(+) cells), while enhancing the bone marrow abundance of these cells. Although similar changes in vascular insulin signaling and EPC levels were observed in mice fed high-fat diet, CAP exposure did not exacerbate diet-induced changes in vascular insulin resistance or EPC homeostasis. Treatment with an insulin sensitizer, metformin or rosiglitazone, prevented CAP-induced vascular insulin resistance and NF-κB and inflammasome activation and restored peripheral blood and bone marrow EPC levels. These findings suggest that PM2.5 exposure induces diet-independent vascular insulin resistance and inflammation and prevents EPC mobilization, and that this EPC mobilization defect could be mediated by vascular insulin resistance. Impaired vascular insulin sensitivity may be an important mechanism underlying PM2.5-induced vascular injury, and pharmacological sensitization to insulin action could potentially prevent deficits in vascular repair and mitigate vascular inflammation due to exposure to elevated levels of ambient air pollution. PMID:27016579

  16. Development of a Xeno-Free Autologous Culture System for Endothelial Progenitor Cells Derived from Human Umbilical Cord Blood

    PubMed Central

    Park, Soon-Jung; Kim, Hojin; Bae, Daekyeong

    2013-01-01

    Despite promising preclinical outcomes in animal models, a number of challenges remain for human clinical use. In particular, expanding a large number of endothelial progenitor cells (EPCs) in vitro in the absence of animal-derived products is the most critical hurdle remaining to be overcome to ensure the safety and efficiency of human therapy. To develop in vitro culture conditions for EPCs derived from human cord blood (hCB-EPCs), we isolated extracts (UCE) and collagen (UC-collagen) from umbilical cord tissue to replace their animal-derived counterparts. UC-collagen and UCE efficiently supported the attachment and proliferation of hCB-EPCs in a manner comparable to that of animal-derived collagen in the conventional culture system. Our developed autologous culture system maintained the typical characteristics of hCB-EPCs, as represented by the expression of EPC-associated surface markers. In addition, the therapeutic potential of hCB-EPCs was confirmed when the transplantation of hCB-EPCs cultured in this autologous culture system promoted limb salvage in a mouse model of hindlimb ischemia and was shown to contribute to attenuating muscle degeneration and fibrosis. We suggest that the umbilical cord represents a source for autologous biomaterials for the in vitro culture of hCB-EPCs. The main characteristics and therapeutic potential of hCB-EPCs were not compromised in developed autologous culture system. The absence of animal-derived products in our newly developed in vitro culture removes concerns associated with secondary contamination. Thus, we hope that this culture system accelerates the realization of therapeutic applications of autologous hCB-EPCs for human vascular diseases. PMID:24086472

  17. Interleukin-1β induces fibroblast growth factor 2 expression and subsequently promotes endothelial progenitor cell angiogenesis in chondrocytes

    PubMed Central

    Chien, Szu-Yu; Huang, Chun-Yin; Tsai, Chun-Hao; Wang, Shih-Wei

    2016-01-01

    Arthritis is a process of chronic inflammation that results in joint damage. IL (interleukin)-1β is an inflammatory cytokine that acts as a key mediator of cartilage degradation, and is abundantly expressed in arthritis. Neovascularization is one of the pathological characteristics of arthritis. However, the role of IL-1β in the angiogenesis of chondrocytes remains unknown. In the present study, we demonstrate that stimulating chondrocytes (ATDC5) with IL-1β increased the expression of FGF (fibroblast growth factor)-2, a potent angiogenic inducer, and then promoted EPC (endothelial progenitor cell) tube formation and migration. In addition, FGF-2-neutralizing antibody abolished ATDC5-conditional medium-mediated angiogenesis in vitro, as well as its angiogenic effects in the CAM (chick chorioallantoic membrane) assay and Matrigel plug nude mice model in vivo. IHC (immunohistochemistry) staining from a CIA (collagen-induced arthritis) mouse model also demonstrates that arthritis increased the expression of IL-1β and FGF-2, as well as EPC homing in articular cartilage. Moreover, IL-1β-induced FGF-2 expression via IL-1RI (type-1 IL-1 receptor), ROS (reactive oxygen species) generation, AMPK (AMP-activated protein kinase), p38 and NF-κB (nuclear factor κB) pathway has been demonstrated. On the basis of these findings, we conclude that IL-1β promotes FGF-2 expression in chondrocytes through the ROS/AMPK/p38/NF-κB signalling pathway and subsequently increases EPC angiogenesis. Therefore IL-1β serves as a link between inflammation and angiogenesis during arthritis. PMID:26811540

  18. Cytoprotective effect of dieckol on human endothelial progenitor cells (hEPCs) from oxidative stress-induced apoptosis.

    PubMed

    Lee, S H; Kim, J Y; Yoo, S Y; Kwon, S M

    2013-07-01

    Although endothelial progenitor cells (EPCs) have been used to promote revascularization after peripheral or myocardial ischemia, excess amounts of reactive oxygen species (ROS) are often involved in senescence and apoptosis of EPCs, thereby causing defective neovascularization and reduced or failed recovery. Here, we examined the cytoprotective effect of Ecklonia cava-derived antioxidant dieckol (DK) on oxidative stress-induced apoptosis in EPCs to improve EPC bioactivity for vessel repair. Although H2O2 (10 (- 3) M) increased the intracellular ROS level in EPCs, DK (10ug/ml) pretreatment suppressed the H2O2-induced ROS increase and drastically reduced the ratios of apoptotic cells. H2O2-induced ROS increased the phosphorylation of p38 MAPK and JNK; this was inhibited by DK pretreatment. H2O2 treatment increased the phosphorylation of NF-κB, which was blocked by pretreatment with SB 203580, a p38 MAPK inhibitor, or SP 600125, a JNK inhibitor. H2O2 decreased the cellular levels of Bcl-2 and c-IAPs, cellular inhibitors of apoptosis proteins, but increased caspase-3 activation. However, all these effects were inhibited by pretreatment with DK. Injection of DK-mixed EPCs (DK + EPCs) into myocardial ischemic sites in vivo induced cellular proliferation and survival of cells at the ischemic sites and, thereby, enhanced the secretion of angiogenic cytokines at the ischemic sites. These results show that DK + EPC exhibit markedly enhanced anti-apoptotic and antioxidative capabilities, unlike that shown by EPCs alone; thus, they contribute to improved repair of ischemic myocardial injury through cell survival and angiogenic cytokine production. PMID:23607503

  19. The Ape-1/Ref-1 redox antagonist E3330 inhibits the growth of tumor endothelium and endothelial progenitor cells: therapeutic implications in tumor angiogenesis.

    PubMed

    Zou, Gang-Ming; Karikari, Collins; Kabe, Yasuaki; Handa, Hiroshi; Anders, Robert A; Maitra, Anirban

    2009-04-01

    The apurinic/apyrimidinic endonuclease 1/redox factor-1 (Ape-1/Ref-1) is a multi-functional protein, involved in DNA repair and the activation of redox-sensitive transcription factors. The Ape-1/Ref-1 redox domain acts as a cytoprotective element in normal endothelial cells, mitigating the deleterious effects of apoptotic stimuli through induction of survival signals. We explored the role of the Ape-1/Ref-1 redox domain in the maintenance of tumor-associated endothelium, and of endothelial progenitor cells (EPCs), which contribute to tumor angiogenesis. We demonstrate that E3330, a small molecule inhibitor of the Ape-1/Ref-1 redox domain, blocks the in vitro growth of pancreatic cancer-associated endothelial cells (PCECs) and EPCs, which is recapitulated by stable expression of a dominant-negative redox domain mutant. Further, E3330 blocks the differentiation of bone marrow-derived mesenchymal stem cells (BMSCs) into CD31(+) endothelial progeny. Exposure of PCECs to E3330 results in a reduction of H-ras expression and intracellular nitric oxide (NO) levels, as well as decreased DNA-binding activity of the hypoxia-inducible transcription factor, HIF-1alpha. E3330 also reduces secreted and intracellular vascular endothelial growth factor expression by pancreatic cancer cells, while concomitantly downregulating the cognate receptor Flk-1/KDR on PCECs. Inhibition of the Ape-1/Ref-1 redox domain with E3330 or comparable angiogenesis inhibitors might be a potent therapeutic strategy in solid tumors. PMID:19097035

  20. Increased expression of microRNA-221 inhibits PAK1 in endothelial progenitor cells and impairs its function via c-Raf/MEK/ERK pathway

    SciTech Connect

    Zhang, Xiaoping; Mao, Haian; Chen, Jin-yuan; Wen, Shengjun; Li, Dan; Ye, Meng; Lv, Zhongwei

    2013-02-15

    Highlights: ► MicroRNA-221 is upregulated in the endothelial progenitor cells of atherosclerosis patients. ► PAK1 is a direct target of microRNA-221. ► MicroRNA-221 inhibits EPCs proliferation through c-Raf/MEK/ERK pathway. -- Abstract: Coronary artery disease (CAD) is associated with high mortality and occurs via endothelial injury. Endothelial progenitor cells (EPCs) restore the integrity of the endothelium and protect it from atherosclerosis. In this study, we compared the expression of microRNAs (miRNAs) in EPCs in atherosclerosis patients and normal controls. We found that miR-221 expression was significantly up-regulated in patients compared with controls. We predicted and identified p21/Cdc42/Rac1-activated kinase 1 (PAK1) as a novel target of miR-221 in EPCs. We also demonstrated that miR-221 targeted a putative binding site in the 3′UTR of PAK1, and absence of this site was inversely associated with miR-221 expression in EPCs. We confirmed this relationship using a luciferase reporter assay. Furthermore, overexpression of miR-221 in EPCs significantly decreased EPC proliferation, in accordance with the inhibitory effects induced by decreased PAK1. Overall, these findings demonstrate that miR-221 affects the MEK/ERK pathway by targeting PAK1 to inhibit the proliferation of EPCs.

  1. Vascular endothelial growth factors A and C are induced in the SVZ following neonatal hypoxia-ischemia and exert different effects on neonatal glial progenitors

    PubMed Central

    Bain, Jennifer M.; Moore, Lisamarie; Ren, Zhihua; Simonishvili, Sophia; Levison, Steven W.

    2012-01-01

    Episodes of neonatal hypoxia-ischemia (H-I) are strongly associated with cerebral palsy and a wide spectrum of other neurological deficits in children. Two key processes required to repair damaged organs are to amplify the number of precursors capable of regenerating damaged cells and to direct their differentiation towards the cell types that need to be replaced. Since hypoxia induces vascular endothelial growth factor (VEGF) production, it is logical to predict that VEGFs are key mediators of tissue repair after H-I injury. The goal of this study was to test the hypothesis that certain VEGF isoforms increase during recovery from neonatal H-I and that they would differentially affect the proliferation and differentiation of subventricular zone (SVZ) progenitors. During the acute recovery period from H-I both VEGF-A and VEGF-C were transiently induced in the SVZ, which correlated with an increase in SVZ blood vessel diameter. These growth factors were produced by glial progenitors, astrocytes and to a lesser extent, microglia. VEGF-A promoted the production of astrocytes from SVZ glial progenitors while VEGF-C stimulated the proliferation of both early and late oligodendrocyte progenitors, which was abolished by blocking the VEGFR-3. Altogether, these results provide new insights into the signals that coordinate the reactive responses of the progenitors in the SVZ to neonatal H-I. Our studies further suggest that therapeutics that extend VEGF-C production and/or agonists that stimulate the VEGFR-3 will promote oligodendrocyte progenitor cell development to enhance myelination after perinatal brain injury. PMID:23565129

  2. Endothelial Cell-Selective Adhesion Molecule Expression in Hematopoietic Stem/Progenitor Cells Is Essential for Erythropoiesis Recovery after Bone Marrow Injury

    PubMed Central

    Sudo, Takao; Yokota, Takafumi; Okuzaki, Daisuke; Ueda, Tomoaki; Ichii, Michiko; Ishibashi, Tomohiko; Isono, Tomomi; Habuchi, Yoko; Oritani, Kenji; Kanakura, Yuzuru

    2016-01-01

    Numerous red blood cells are generated every second from proliferative progenitor cells under a homeostatic state. Increased erythropoietic activity is required after myelo-suppression as a result of chemo-radio therapies. Our previous study revealed that the endothelial cell-selective adhesion molecule (ESAM), an authentic hematopoietic stem cell marker, plays essential roles in stress-induced hematopoiesis. To determine the physiological importance of ESAM in erythroid recovery, ESAM-knockout (KO) mice were treated with the anti-cancer drug, 5-fluorouracil (5-FU). ESAM-KO mice experienced severe and prolonged anemia after 5-FU treatment compared to wild-type (WT) mice. Eight days after the 5-FU injection, compared to WT mice, ESAM-KO mice showed reduced numbers of erythroid progenitors in bone marrow (BM) and spleen, and reticulocytes in peripheral blood. Megakaryocyte-erythrocyte progenitors (MEPs) from the BM of 5-FU-treated ESAM-KO mice showed reduced burst forming unit-erythrocyte (BFU-E) capacities than those from WT mice. BM transplantation revealed that hematopoietic stem/progenitor cells from ESAM-KO donors were more sensitive to 5-FU treatment than that from WT donors in the WT host mice. However, hematopoietic cells from WT donors transplanted into ESAM-KO host mice could normally reconstitute the erythroid lineage after a BM injury. These results suggested that ESAM expression in hematopoietic cells, but not environmental cells, is critical for hematopoietic recovery. We also found that 5-FU treatment induces the up-regulation of ESAM in primitive erythroid progenitors and macrophages that do not express ESAM under homeostatic conditions. The phenotypic change seen in macrophages might be functionally involved in the interaction between erythroid progenitors and their niche components during stress-induced acute erythropoiesis. Microarray analyses of primitive erythroid progenitors from 5-FU-treated WT and ESAM-KO mice revealed that various signaling

  3. Effects of lycopene on number and function of human peripheral blood endothelial progenitor cells cultivated with high glucose

    PubMed Central

    Zeng, Yao-Chi; Mu, Gui-Ping; Huang, Shu-Fen; Zeng, Xue-Hui; Cheng, Hong

    2014-01-01

    BACKGROUND/OBJECTIVES The objectives of this study were to investigate the effects of lycopene on the migration, adhesion, tube formation capacity, and p38 mitogen-activated protein kinase (p38 MAPK) activity of endothelial progenitor cells (EPCs) cultivated with high glucose (HG) and as well as explore the mechanism behind the protective effects of lycopene on peripheral blood EPCs. MATERIALS/METHODS Mononuclear cells were isolated from human peripheral blood by Ficoll density gradient centrifugation. EPCs were identified after induction of cellular differentiation. Third generation EPCs were incubated with HG (33 mmol/L) or 10, 30, and 50 µg/mL of lycopene plus HG. MTT assay and flow cytometry were performed to assess proliferation and apoptosis of EPCs. EPC migration was assessed by MTT assay with a modified boyden chamber. Adhesion assay was performed by replating EPCs on fibronectin-coated dishes, after which adherent cells were counted. In vitro vasculogenesis activity was assayed by Madrigal network formation assay. Western blotting was performed to analyze protein expression of both phosphorylated and non-phosphorylated p38 MAPK. RESULTS The proliferation, migration, adhesion, and in vitro vasculogenesis capacity of EPCs treated with 10, 30, and 50 µg/mL of lycopene plus HG were all significantly higher comapred to the HG group (P < 0.05). Rates of apoptosis were also significantly lower than that of the HG group. Moreover, lycopene blocked phosphorylation of p38 MAPK in EPCs (P < 0.05). To confirm the causal relationship between MAPK inhibition and the protective effects of lycopene against HG-induced cellular injury, we treated cells with SB203580, a phosphorylation inhibitor. The inhibitor significantly inhibited HG-induced EPC injury. CONCLUSIONS Lycopene promotes proliferation, migration, adhesion, and in vitro vasculogenesis capacity as well as reduces apoptosis of EPCs. Further, the underlying molecular mechanism of the protective effects of

  4. Association between Microalbuminuria Predicting In-Stent Restenosis after Myocardial Infarction and Cellular Senescence of Endothelial Progenitor Cells

    PubMed Central

    Ota, Hisanobu; Takehara, Naofumi; Aonuma, Tatsuya; Kabara, Maki; Matsuki, Motoki; Yamauchi, Atsushi; Takeuchi, Toshiharu; Kawabe, Jun-ichi; Hasebe, Naoyuki

    2015-01-01

    Objective Relationship between microalbuminuria and worse outcome of coronary artery disease patients is discussed, but its underlying pathophysiological mechanism remains unclear. We investigated the role of microalbuminuria to the function of endothelial progenitor cells (EPCs), that might affect to outcome of acute myocardial infarction (AMI) patients. Methods Forty-five AMI patients were divided into two groups according to their urinary albumin excretion: normal (n = 24) and microalbuminuria (>30 mg/day, n = 21). At day-2 and day-7 after AMI onset, circulating-EPCs (CD34+Flk1+) were quantified by flow cytometry. The number of lectin-acLDL-positive cultured-EPCs immobilized on fibronectin was determined. To assess the cellular senescence of cultured-EPCs, the expression level of sirtuin-1 mRNA and the number of SA-β-gal positive cell were evaluated. Angiographic late in-stent loss after percutaneous coronary intervention (PCI) was evaluated at a six-month follow-up. Results No significant differences in coronary risk and the extent of myocardial damage were observed between the two groups. Late in-stent loss at the six-month follow-up was significantly higher in the microalbuminuria group (normal : microalbuminuria = 0.76±0.34 : 1.18±0.57 mm, p=0.021). The number of circulating-EPCs was significantly increased in microalbuminuria group at day-7, however, improved adhesion of EPCs was observed in normal group but not in microalbuminuria group from baseline to day-7 (+3.1±8.3 : -1.3±4.4 %: p<0.05). On the other hand, in microalbuminuria group at day-7, the level of sirtuin-1 mRNA expression of cultured-EPCs was significantly decreased (7.1±8.9 : 2.5±3.7 fold, p<0.05), which was based on the negative correlation between the level of sirtuin-1 mRNA expression and the extent of microalbuminuria. The ratio of SA-β-gal-positive cells in microalbuminuria group was increased compared to that of normal group. Conclusions Microalbuminuria in AMI patients is

  5. Impact of obesity control on circulating level of endothelial progenitor cells and angiogenesis in response to ischemic stimulation

    PubMed Central

    2012-01-01

    Background and aim We tested the hypothesis that obesity reduced circulating number of endothelial progenitor cells (EPCs), angiogenic ability, and blood flow in ischemic tissue that could be reversed after obesity control. Methods 8-week-old C57BL/6J mice (n = 27) were equally divided into group 1 (fed with 22-week control diet), group 2 (22-week high fat diet), and group 3 (14-week high fat diet, followed by 8-week control diet). Critical limb ischemia (CLI) was induced at week 20 in groups 2 and 3. The animals were sacrificed at the end of 22 weeks. Results Heart weight, body weight, abdominal fat weight, serum total cholesterol level, and fasting blood sugar were highest in group 2 (all p < 0.001). The numbers of circulating EPCs (C-kit/CD31+, Sca-1/KDR + and CXCR4/CD34+) were lower in groups 1 and 2 than in group 3 at 18 h after CLI induction (p < 0.03). The numbers of differentiated EPCs (C-kit/CD31+, CXCR4/CD34+ and CD133+) from adipose tissue after 14-day cultivation were also lowest in group 2 (p < 0.001). Protein expressions of VCAM-1, oxidative index, Smad3, and TGF-β were higher, whereas the Smad1/5 and BMP-2, mitochondrial cytochrome-C SDF-1α and CXCR4 were lower in group 2 than in groups 1 and 3 (all p < 0.02). Immunofluorescent staining of CD31+ and vWF + cells, the number of small vessel (<15 μm), and blood flow through Laser Doppler scanning of ischemic area were lower in group 2 compared to groups 1 and 3 on day 14 after CLI induction (all p < 0.001). Conclusion Obesity suppressed abilities of angiogenesis and recovery from CLI that were reversed by obesity control. PMID:22568992

  6. Synergy of endothelial and neural progenitor cells from adipose-derived stem cells to preserve neurovascular structures in rat hypoxic-ischemic brain injury

    PubMed Central

    Hsueh, Yuan-Yu; Chang, Ya-Ju; Huang, Chia-Wei; Handayani, Fitri; Chiang, Yi-Lun; Fan, Shih-Chen; Ho, Chien-Jung; Kuo, Yu-Min; Yang, Shang-Hsun; Chen, Yuh-Ling; Lin, Sheng-Che; Huang, Chao-Ching; Wu, Chia-Ching

    2015-01-01

    Perinatal cerebral hypoxic-ischemic (HI) injury damages the architecture of neurovascular units (NVUs) and results in neurological disorders. Here, we differentiated adipose-derived stem cells (ASCs) toward the progenitor of endothelial progenitor cells (EPCs) and neural precursor cells (NPCs) via microenvironmental induction and investigated the protective effect by transplanting ASCs, EPCs, NPCs, or a combination of EPCs and NPCs (E+N) into neonatal HI injured rat pups. The E+N combination produced significant reduction in brain damage and cell apoptosis and the most comprehensive restoration in NVUs regarding neuron number, normal astrocytes, and vessel density. Improvements in cognitive and motor functions were also achieved in injured rats with E+N therapy. Synergistic interactions to facilitate transmigration under in vitro hypoxic microenvironment were discovered with involvement of the neuropilin-1 (NRP1) signal in EPCs and the C-X-C chemokine receptor 4 (CXCR4) and fibroblast growth factor receptor 1 (FGFR1) signals in NPCs. Therefore, ASCs exhibit great potential for cell sources in endothelial and neural lineages to prevent brain from HI damage. PMID:26447335

  7. Synergy of endothelial and neural progenitor cells from adipose-derived stem cells to preserve neurovascular structures in rat hypoxic-ischemic brain injury.

    PubMed

    Hsueh, Yuan-Yu; Chang, Ya-Ju; Huang, Chia-Wei; Handayani, Fitri; Chiang, Yi-Lun; Fan, Shih-Chen; Ho, Chien-Jung; Kuo, Yu-Min; Yang, Shang-Hsun; Chen, Yuh-Ling; Lin, Sheng-Che; Huang, Chao-Ching; Wu, Chia-Ching

    2015-01-01

    Perinatal cerebral hypoxic-ischemic (HI) injury damages the architecture of neurovascular units (NVUs) and results in neurological disorders. Here, we differentiated adipose-derived stem cells (ASCs) toward the progenitor of endothelial progenitor cells (EPCs) and neural precursor cells (NPCs) via microenvironmental induction and investigated the protective effect by transplanting ASCs, EPCs, NPCs, or a combination of EPCs and NPCs (E+N) into neonatal HI injured rat pups. The E+N combination produced significant reduction in brain damage and cell apoptosis and the most comprehensive restoration in NVUs regarding neuron number, normal astrocytes, and vessel density. Improvements in cognitive and motor functions were also achieved in injured rats with E+N therapy. Synergistic interactions to facilitate transmigration under in vitro hypoxic microenvironment were discovered with involvement of the neuropilin-1 (NRP1) signal in EPCs and the C-X-C chemokine receptor 4 (CXCR4) and fibroblast growth factor receptor 1 (FGFR1) signals in NPCs. Therefore, ASCs exhibit great potential for cell sources in endothelial and neural lineages to prevent brain from HI damage. PMID:26447335

  8. Statins, HMG-CoA Reductase Inhibitors, Improve Neovascularization by Increasing the Expression Density of CXCR4 in Endothelial Progenitor Cells

    PubMed Central

    Chiang, Kuang-Hsing; Cheng, Wan-Li; Shih, Chun-Ming; Lin, Yi-Wen; Tsao, Nai-Wen; Kao, Yung-Ta; Lin, Chih-Ting; Wu, Shinn-Chih; Huang, Chun-Yao; Lin, Feng-Yen

    2015-01-01

    Statins, inhibitors of 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase, are used to reduce cholesterol biosynthesis in the liver. Accordingly, statins regulate nitric oxide (NO) and glutamate metabolism, inflammation, angiogenesis, immunity and endothelial progenitor cells (EPCs) functions. The function of EPCs are regulated by stromal cell-derived factor 1 (SDF-1), vascular endothelial growth factor (VEGF), and transforming growth factor β (TGF-β), etc. Even though the pharmacologic mechanisms by which statins affect the neovasculogenesis of circulating EPCs, it is still unknown whether statins affect the EPCs function through the regulation of CXCR4, a SDF-1 receptor expression. Therefore, we desired to explore the effects of statins on CXCR4 expression in EPC-mediated neovascularization by in vitro and in vivo analyses. In animal studies, we analyzed the effects of atorvastatin or rosuvastatin treatments in recovery of capillary density and blood flow, the expression of vWF and CXCR4 at ischemia sites in hindlimb ischemia ICR mice. Additionally, we analyzed whether the atorvastatin or rosuvastatin treatments increased the mobilization, homing, and CXCR4 expression of EPCs in hindlimb ischemia ICR mice that underwent bone marrow transplantation. The results indicated that statins treatment led to significantly more CXCR4-positive endothelial progenitor cells incorporated into ischemic sites and in the blood compared with control mice. In vivo, we isolated human EPCs and analyzed the effect of statins treatment on the vasculogenic ability of EPCs and the expression of CXCR4. Compared with the control groups, the neovascularization ability of EPCs was significantly improved in the atorvastatin or rosuvastatin group; this improvement was dependent on CXCR4 up-regulation. The efficacy of statins on improving EPC neovascularization was related to the SDF-1α/CXCR4 axis and might be regulated by the NO. In conclusion, atorvastatin and rosuvastatin improved

  9. Irradiated human endothelial progenitor cells induce bystander killing in human non-small cell lung and pancreatic cancer cells.

    PubMed

    Turchan, William T; Shapiro, Ronald H; Sevigny, Garrett V; Chin-Sinex, Helen; Pruden, Benjamin; Mendonca, Marc S

    2016-08-01

    Purpose To investigate whether irradiated human endothelial progenitor cells (hEPC) could induce bystander killing in the A549 non-small cell lung cancer (NSCLC) cells and help explain the improved radiation-induced tumor cures observed in A549 tumor xenografts co-injected with hEPC. Materials and methods We investigated whether co-injection of CBM3 hEPC with A549 NSCLC cells would alter tumor xenograft growth rate or tumor cure after a single dose of 0 or 5 Gy of X-rays. We then utilized dual chamber Transwell dishes, to test whether medium from irradiated CBM3 and CBM4 hEPC would induce bystander cell killing in A549 cells, and as an additional control, in human pancreatic cancer MIA PaCa-2 cells. The CBM3 and CBM4 hEPC were plated into the upper Transwell chamber and the A549 or MIA PaCa-2 cells were plated in the lower Transwell chamber. The top inserts with the CBM3 or CBM4 hEPC cells were subsequently removed, irradiated, and then placed back into the Transwell dish for 3 h to allow for diffusion of any potential bystander factors from the irradiated hEPC in the upper chamber through the permeable membrane to the unirradiated cancer cells in the lower chamber. After the 3 h incubation, the cancer cells were re-plated for clonogenic survival. Results We found that co-injection of CBM3 hEPC with A549 NSCLC cells significantly increased the tumor growth rate compared to A549 cells alone, but paradoxically also increased A549 tumor cure after a single dose of 5 Gy of X-rays (p < 0.05). We hypothesized that irradiated hEPC may be inducing bystander killing in the A549 NSCLC cells in tumor xenografts, thus improving tumor cure. Bystander studies clearly showed that exposure to the medium from irradiated CBM3 and CBM4 hEPC induced significant bystander killing and decreased the surviving fraction of A549 and MIA PaCa-2 cells to 0.46 (46%) ± 0.22 and 0.74 ± 0.07 (74%) respectively (p < 0.005, p < 0.0001). In addition, antibody depletion

  10. Data regarding association between serum osteoprotegerin level, numerous of circulating endothelial-derived and mononuclear-derived progenitor cells in patients with metabolic syndrome.

    PubMed

    Berezin, Alexander E; Kremzer, Alexander A; Berezina, Tatyana A; Martovitskaya, Yulia V; Gronenko, Elena A

    2016-09-01

    Metabolic syndrome (MetS) is defined as cluster of multiple metabolic and cardiovascular (CV) abnormalities included abdominal obesity, high-normal blood pressure, dyslipidaemia, and impaired fasting glucose tolerance that exhibits has a growing prevalence worldwide. We investigated whether an elevated level of osteoprotegerin (OPG) predicts imbalance between different phenotypes of circulating endothelial (EPCs) and mononuclear (MPCs) progenitor cells in MetS patients. We have analyzed data regarding dysmetabolic disorder subjects without known CV disease), as well as with known type two diabetes mellitus. All patients have given their informed written consent for participation in the study. This article contains data on the independent predictors of depletion in numerous of circulating EPCs and MPCs in MetS patients. The data are supplemental to our original research article describing detailed associations of elevated OPG level in MetS patients with numerous of EPCs and MPCs beyond traditional CV risk factors. PMID:27508223

  11. Self-Renewal and High Proliferative Colony Forming Capacity of Late-Outgrowth Endothelial Progenitors Is Regulated by Cyclin-Dependent Kinase Inhibitors Driven by Notch Signaling.

    PubMed

    Patel, Jatin; Wong, Ho Yi; Wang, Weili; Alexis, Josue; Shafiee, Abbas; Stevenson, Alexander J; Gabrielli, Brian; Fisk, Nicholas M; Khosrotehrani, Kiarash

    2016-04-01

    Since the discovery of endothelial colony forming cells (ECFC), there has been significant interest in their therapeutic potential to treat vascular injuries. ECFC cultures display significant heterogeneity and a hierarchy among cells able to give rise to high proliferative versus low proliferative colonies. Here we aimed to define molecularly this in vitro hierarchy. Based on flow cytometry, CD34 expression levels distinguished two populations. Only CD34 + ECFC had the capacity to reproduce high proliferative potential (HPP) colonies on replating, whereas CD34- ECFCs formed only small clusters. CD34 + ECFCs were the only ones to self-renew in stringent single-cell cultures and gave rise to both CD34 + and CD34- cells. Upon replating, CD34 + ECFCs were always found at the centre of HPP colonies and were more likely in G0/1 phase of cell cycling. Functionally, CD34 + ECFC were superior at restoring perfusion and better engrafted when injected into ischemic hind limbs. Transcriptomic analysis identified cyclin-dependent kinase (CDK) cell cycle inhibiting genes (p16, p21, and p57), the Notch signaling pathway (dll1, dll4, hes1, and hey1), and the endothelial cytokine il33 as highly expressed in CD34 + ECFC. Blocking the Notch pathway using a γ-secretase inhibitor (DAPT) led to reduced expression of cell cycle inhibitors, increased cell proliferation followed by a loss of self-renewal, and HPP colony formation capacity reflecting progenitor exhaustion. Similarly shRNA knockdown of p57 strongly affected self-renewal of ECFC colonies. ECFC hierarchy is defined by Notch signalling driving cell cycle regulators, progenitor quiescence and self-renewal potential. Stem Cells 2016;34:902-912. PMID:26732848

  12. TGFβ inhibition enhances the generation of hematopoietic progenitors from human ES cell-derived hemogenic endothelial cells using a stepwise strategy

    PubMed Central

    Wang, Chengyan; Tang, Xuming; Sun, Xiaomeng; Miao, Zhenchuan; Lv, Yaxin; Yang, Yanlei; Zhang, Huidan; Zhang, Pengbo; Liu, Yang; Du, Liying; Gao, Yang; Yin, Ming; Ding, Mingxiao; Deng, Hongkui

    2012-01-01

    Embryonic hematopoiesis is a complex process. Elucidating the mechanism regulating hematopoietic differentiation from pluripotent stem cells would allow us to establish a strategy to efficiently generate hematopoietic cells. However, the mechanism governing the generation of hematopoietic progenitors from human embryonic stem cells (hESCs) remains unknown. Here, on the basis of the emergence of CD43+ hematopoietic cells from hemogenic endothelial (HE) cells, we demonstrated that VEGF was essential and sufficient, and that bFGF was synergistic with VEGF to specify the HE cells and the subsequent transition into CD43+ hematopoietic cells. Significantly, we identified TGFβ as a novel signal to regulate hematopoietic development, as the TGFβ inhibitor SB 431542 significantly promoted the transition from HE cells into CD43+ hematopoietic progenitor cells (HPCs) during hESC differentiation. By defining these critical signaling factors during hematopoietic differentiation, we can efficiently generate HPCs from hESCs. Our strategy could offer an in vitro model to study early human hematopoietic development. PMID:21862970

  13. Cilostazol Enhances Mobilization of Circulating Endothelial Progenitor Cells and Improves Endothelium-Dependent Function in Patients at High Risk of Cardiovascular Disease.

    PubMed

    Chao, Ting-Hsing; Chen, I-Chih; Lee, Cheng-Han; Chen, Ju-Yi; Tsai, Wei-Chuan; Li, Yi-Heng; Tseng, Shih-Ya; Tsai, Liang-Miin; Tseng, Wei-Kung

    2016-08-01

    This is the first study to investigate the vasculoangiogenic effects of cilostazol on endothelial progenitor cells (EPCs) and flow-mediated dilatation (FMD) in patients at high risk of cardiovascular disease (CVD). This double-blind, placebo-controlled study included 71 patients (37 received 200 mg/d cilostazol and 34 received placebo for 12 weeks). Use of cilostazol, but not placebo, significantly increased circulating EPC (kinase insert domain receptor(+)CD34(+)) counts (percentage changes: 149.0% [67.9%-497.8%] vs 71.9% [-31.8% to 236.5%], P = .024) and improved triglyceride and high-density lipoprotein cholesterol levels (P = .002 and P = .003, respectively). Plasma levels of vascular endothelial growth factor (VEGF)-A165 and FMD significantly increased (72.5% [32.9%-120.4%] vs -5.8% [-46.0% to 57.6%], P = .001; 232.8% ± 83.1% vs -46.9% ± 21.5%, P = .003, respectively) in cilostazol-treated patients. Changes in the plasma triglyceride levels significantly inversely correlated with the changes in the VEGF-A165 levels and FMD. Cilostazol significantly enhanced the mobilization of EPCs and improved endothelium-dependent function by modifying some metabolic and angiogenic markers in patients at high risk of CVD. PMID:27401788

  14. Surface modification of a polyhedral oligomeric silsesquioxane poly(carbonate-urea) urethane (POSS-PCU) nanocomposite polymer as a stent coating for enhanced capture of endothelial progenitor cells

    PubMed Central

    Tan, Aaron; Farhatnia, Yasmin; Goh, Debbie; G, Natasha; de Mel, Achala; Lim, Jing; Teoh, Swee-Hin; Malkovskiy, Andrey V; Chawla, Reema; Rajadas, Jayakumar; Cousins, Brian G; Hamblin, Michael R; Alavijeh, Mohammad S; Seifalian, Alexander M

    2013-01-01

    An unmet need exists for the development of next-generation multifunctional nanocomposite materials for biomedical applications, particularly in the field of cardiovascular regenerative biology. Herein, we describe the preparation and characterization of a novel polyhedral oligomeric silsesquioxane poly(carbonate-urea) urethane (POSS-PCU) nanocomposite polymer with covalently attached anti-CD34 antibodies to enhance capture of circulating endothelial progenitor cells (EPC). This material may be used as a new coating for bare metal stents used after balloon angioplasty to improve re-endothelialization. Biophysical characterization techniques were used to assess POSS-PCU and its subsequent functionalization with anti-CD34 antibodies. Results indicated successful covalent attachment of anti-CD34 antibodies on the surface of POSS-PCU leading to an increased propensity for EPC capture, whilst maintaining in vitro biocompatibility and hemocompatibility. POSS-PCU has already been used in 3 first-in-man studies, as a bypass graft, lacrimal duct and a bioartificial trachea. We therefore postulate that its superior biocompatibility and unique biophysical properties would render it an ideal candidate for coating medical devices, with stents as a prime example. Taken together, anti-CD34 functionalized POSS-PCU could form the basis of a nano-inspired polymer platform for the next generation stent coatings. PMID:24706135

  15. Cilostazol Improves Proangiogenesis Functions in Human Early Endothelial Progenitor Cells through the Stromal Cell-Derived Factor System and Hybrid Therapy Provides a Synergistic Effect In Vivo.

    PubMed

    Tseng, Shih-Ya; Chao, Ting-Hsing; Li, Yi-Heng; Cho, Chung-Lung

    2016-01-01

    This study investigated the effect of cilostazol on proangiogenesis functions in human early endothelial progenitor cells (EPCs) in vitro and the therapeutic implication of hybrid therapy with cilostazol and human early EPCs in vivo. Cilostazol significantly increased colony-forming units and enhanced differentiation of EPCs toward endothelial lineage. Treatments resulted in antiapoptotic effects and stimulated proliferation and migration and in vitro vascular tube formation through activation of stromal cell-derived factor-1 (SDF-1)/C-X-C chemokine receptor type 4 (CXCR4)/phosphatidylinositol-3 kinase (PI3K)/Akt signaling pathway. Blood flow recovery and capillary density in murine ischemic hindlimbs were significantly improved in cilostazol-treated, human early EPCs-treated, and cotreatment groups. The effects were attenuated with SDF-1α inhibition. Plasma SDF-1α levels were significantly higher in 3 active treatment groups after surgery, with greatest effects observed in hybrid therapy. The angiogenic effects of transplanted EPCs pretreated with cilostazol ex vivo were superior to untreated EPCs using in vivo Matrigel assay. Implanted EPCs were incorporated into the capillary, with pretreatment or cotreatment with cilostazol resulting in enhanced effects. Taken together, cilostazol promotes a large number of proangiogenic functions in human early EPCs through activation of SDF-1/CXCR4/PI3K/Akt signaling, and hybrid therapy provides a synergistic effect in vivo. Cotreatment may be beneficial in ischemic disease. PMID:27595100

  16. Cilostazol Improves Proangiogenesis Functions in Human Early Endothelial Progenitor Cells through the Stromal Cell-Derived Factor System and Hybrid Therapy Provides a Synergistic Effect In Vivo

    PubMed Central

    Tseng, Shih-Ya; Li, Yi-Heng; Cho, Chung-Lung

    2016-01-01

    This study investigated the effect of cilostazol on proangiogenesis functions in human early endothelial progenitor cells (EPCs) in vitro and the therapeutic implication of hybrid therapy with cilostazol and human early EPCs in vivo. Cilostazol significantly increased colony-forming units and enhanced differentiation of EPCs toward endothelial lineage. Treatments resulted in antiapoptotic effects and stimulated proliferation and migration and in vitro vascular tube formation through activation of stromal cell-derived factor-1 (SDF-1)/C-X-C chemokine receptor type 4 (CXCR4)/phosphatidylinositol-3 kinase (PI3K)/Akt signaling pathway. Blood flow recovery and capillary density in murine ischemic hindlimbs were significantly improved in cilostazol-treated, human early EPCs-treated, and cotreatment groups. The effects were attenuated with SDF-1α inhibition. Plasma SDF-1α levels were significantly higher in 3 active treatment groups after surgery, with greatest effects observed in hybrid therapy. The angiogenic effects of transplanted EPCs pretreated with cilostazol ex vivo were superior to untreated EPCs using in vivo Matrigel assay. Implanted EPCs were incorporated into the capillary, with pretreatment or cotreatment with cilostazol resulting in enhanced effects. Taken together, cilostazol promotes a large number of proangiogenic functions in human early EPCs through activation of SDF-1/CXCR4/PI3K/Akt signaling, and hybrid therapy provides a synergistic effect in vivo. Cotreatment may be beneficial in ischemic disease. PMID:27595100

  17. Cardiac shockwave therapy improves myocardial function in patients with refractory coronary artery disease by promoting VEGF and IL-8 secretion to mediate the proliferation of endothelial progenitor cells

    PubMed Central

    CAI, HONG-YAN; LI, LIN; GUO, TAO; WANG, YU; MA, TIE-KUN; XIAO, JIAN-MING; ZHAO, LING; FANG, YIN; YANG, PING; ZHAO, HU

    2015-01-01

    Cardiac shockwave therapy (CSWT) is a potential and effective remedy to promote revascularization in the ischemic myocardium of patients with refractory coronary heart disease (CHD). The technique is both safe and non-invasive; however, the underlying molecular mechanism remains unclear. The aim of this study was to evaluate the efficacy of CSWT in treating CHD patients and investigate a potential mechanism. A total of 26 patients with CHD were enrolled in the study, and CSWT was performed over a 3-month period. The efficacy of CSWT was assessed using several clinical parameters. Peripheral blood (PB) was collected prior to and following treatment. The number of circulating endothelial progenitor cells (EPCs) in the PB was counted using a flow cytometer, and the levels of vascular endothelial growth factor (VEGF), interleukin-8 (IL-8), stromal cell-derived factor 1 and matrix metalloproteinase 9 in the PB were analyzed. Mononuclear cells were isolated from the PB and cultured in vitro. The EPCs and EPC-colony forming units (EPC-CFUs) in the PB mononuclear cell culture were counted using an inverted phase contrast microscope. Following CSWT, the tested clinical parameters were significantly improved. The levels of circulating EPCs, VEGF and IL-8 in the PB were significantly increased, as were the EPCs and EPC-CFUs from the PB mononuclear cell culture. We suggest that EPC proliferation, mediated by VEGF and IL-8 secretion, may be among the potential mechanisms associated with CSWT. PMID:26668649

  18. The role of FGF2 in migration and tubulogenesis of endothelial progenitor cells in relation to pro-angiogenic growth factor production.

    PubMed

    Litwin, Monika; Radwańska, Agata; Paprocka, Maria; Kieda, Claudine; Dobosz, Tadeusz; Witkiewicz, Wojciech; Baczyńska, Dagmara

    2015-12-01

    In recent years, special attention has been paid to finding new pro-angiogenic factors which could be used in gene therapy of vascular diseases such as critical limb ischaemia (CLI). Angiogenesis, the formation of new blood vessels, is a complex process dependent on different cytokines, matrix proteins, growth factors and other pro- or anti-angiogenic stimuli. Numerous lines of evidence suggest that key mediators of angiogenesis, vascular endothelial growth factor (VEGF) and hepatocyte growth factor (HGF) together with fibroblast growth factor2 (FGF2) are involved in regulation of the normal and pathological process of angiogenesis. However, less information is available on the complex interactions between these and other angiogenic factors. The aim of this study was to characterise the effect of fibroblast growth factor2 on biological properties of human endothelial progenitor cells with respect to the expression level of other regulatory cytokines. Ectopic expression of FGF2 in EP cells stimulates their pro-angiogenic behaviour, leading to increased proliferation, migration and tube formation abilities. Moreover, we show that the expression profile of VEGF and other pro-angiogenic cytokines, such as HGF, MCP2, and interleukins, is affected differently by FGF2 in EPC. In conclusion, we provide evidence that FGF2 directly affects not only the biological properties of EP cells but also the expression pattern and secretion of numerous chemocytokines. Our results suggest that FGF2 could be applied in therapeutic approaches for CLI and other ischaemic diseases of the vascular system in vivo. PMID:26314253

  19. Surface modification of a polyhedral oligomeric silsesquioxane poly(carbonate-urea) urethane (POSS-PCU) nanocomposite polymer as a stent coating for enhanced capture of endothelial progenitor cells.

    PubMed

    Tan, Aaron; Farhatnia, Yasmin; Goh, Debbie; G, Natasha; de Mel, Achala; Lim, Jing; Teoh, Swee-Hin; Malkovskiy, Andrey V; Chawla, Reema; Rajadas, Jayakumar; Cousins, Brian G; Hamblin, Michael R; Alavijeh, Mohammad S; Seifalian, Alexander M

    2013-12-01

    An unmet need exists for the development of next-generation multifunctional nanocomposite materials for biomedical applications, particularly in the field of cardiovascular regenerative biology. Herein, we describe the preparation and characterization of a novel polyhedral oligomeric silsesquioxane poly(carbonate-urea) urethane (POSS-PCU) nanocomposite polymer with covalently attached anti-CD34 antibodies to enhance capture of circulating endothelial progenitor cells (EPC). This material may be used as a new coating for bare metal stents used after balloon angioplasty to improve re-endothelialization. Biophysical characterization techniques were used to assess POSS-PCU and its subsequent functionalization with anti-CD34 antibodies. Results indicated successful covalent attachment of anti-CD34 antibodies on the surface of POSS-PCU leading to an increased propensity for EPC capture, whilst maintaining in vitro biocompatibility and hemocompatibility. POSS-PCU has already been used in 3 first-in-man studies, as a bypass graft, lacrimal duct and a bioartificial trachea. We therefore postulate that its superior biocompatibility and unique biophysical properties would render it an ideal candidate for coating medical devices, with stents as a prime example. Taken together, anti-CD34 functionalized POSS-PCU could form the basis of a nano-inspired polymer platform for the next generation stent coatings. PMID:24706135

  20. MRI Tracking of FePro Labeled Fresh and Cryopreserved Long Term In Vitro Expanded Human Cord Blood AC133+ Endothelial Progenitor Cells in Rat Glioma

    PubMed Central

    Janic, Branislava; Jafari-Khouzani, Kourosh; Babajani-Feremi, Abbas; Iskander, A. S. M.; Varma, Nadimpalli Ravi S.; Ali, Meser M.; Knight, Robert A.; Arbab, Ali S.

    2012-01-01

    Background Endothelial progenitors cells (EPCs) are important for the development of cell therapies for various diseases. However, the major obstacles in developing such therapies are low quantities of EPCs that can be generated from the patient and the lack of adequate non-invasive imaging approach for in vivo monitoring of transplanted cells. The objective of this project was to determine the ability of cord blood (CB) AC133+ EPCs to differentiate, in vitro and in vivo, toward mature endothelial cells (ECs) after long term in vitro expansion and cryopreservation and to use magnetic resonance imaging (MRI) to assess the in vivo migratory potential of ex vivo expanded and cryopreserved CB AC133+ EPCs in an orthotopic glioma rat model. Materials, Methods and Results The primary CB AC133+ EPC culture contained mainly EPCs and long term in vitro conditions facilitated the maintenance of these cells in a state of commitment toward endothelial lineage. At days 15–20 and 25–30 of the primary culture, the cells were labeled with FePro and cryopreserved for a few weeks. Cryopreserved cells were thawed and in vitro differentiated or IV administered to glioma bearing rats. Different groups of rats also received long-term cultured, magnetically labeled fresh EPCs and both groups of animals underwent MRI 7 days after IV administration of EPCs. Fluorescent microscopy showed that in vitro differentiation of EPCs was not affected by FePro labeling and cryopreservation. MRI analysis demonstrated that in vivo accumulation of previously cryopreserved transplanted cells resulted in significantly higher R2 and R2* values indicating a higher rate of migration and incorporation into tumor neovascularization of previously cryopreserved CB AC133+ EPCs to glioma sites, compared to non-cryopreserved cells. Conclusion Magnetically labeled CB EPCs can be in vitro expanded and cryopreserved for future use as MRI probes for monitoring the migration and incorporation to the sites of

  1. Apolipoprotein A-I mimetic peptide D-4F promotes human endothelial progenitor cell proliferation, migration, adhesion though eNOS/NO pathway.

    PubMed

    Zhang, Zhengang; Qun, Jianhua; Cao, Chunmei; Wang, Jun; Li, Wei; Wu, Yong; Du, Lin; Zhao, Pei; Gong, Kaizheng

    2012-04-01

    Circulating endothelial progenitor cells (EPCs) have a critical role in endothelial maintenance and repair. Apolipoprotein A-I mimetic peptide D-4F has been shown to posses anti-atherogenic properties via sequestration of oxidized phospholipids, induction of remodeling of high density lipoprotein and promotion of cholesterol efflux from macrophage-derived foam cells. In this study, we test the effects of D-4F on EPC biology. EPCs were isolated from the peripheral venous blood of healthy male volunteers and characterized by 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine-labeled acetylated LDL uptake and ulex europaeus agglutinin binding and flow cytometry. Cell proliferation, migration, adhesion, nitric oxide production and endothelial nitric oxide synthase (eNOS) expression in the absence and presence of D-4F or simvastatin (as a positive control), were assayed. We demonstrated that D-4F significantly enhanced EPC proliferation, migration and adhesion in a dose-dependent manner compared with vehicle. However, all of the favorable effects of D-4F on EPCs were dramatically attenuated by preincubation with NOS inhibitor L-NAME. Further, D-4F also increased nitric oxide production in culture supernatant and the levels of eNOS expression and phosphorylation. The stimulatory effects of D-4F (10 μg/ml) on EPC biology were comparable to 0.5 μM simvastatin. These results suggest that eNOS/NO pathway mediates the functional modulation of EPC biology in response to D-4F treatment and support the notion that the beneficial role of D-4F on EPCs may be one of the important components of its anti-atherogenic potential. PMID:21947883

  2. Effects of pitavastatin versus atorvastatin on the peripheral endothelial progenitor cells and vascular endothelial growth factor in high-risk patients: a pilot prospective, double-blind, randomized study

    PubMed Central

    2014-01-01

    Background Circulating endothelial progenitor cells (EPCs) reflect endothelial repair capacity and may be a significant marker for the clinical outcomes of cardiovascular disease. While some high-dose statin treatments may improve endothelial function, it is not known whether different statins may have similar effects on EPCs.This study aimed to investigate the potential class effects of different statin treatment including pitavastatin and atorvastatin on circulating EPCs in clinical setting. Methods A pilot prospective, double-blind, randomized study was conducted to evaluate the ordinary dose of pitavastatin (2 mg daily) or atorvastatin (10 mg daily) treatment for 12 weeks on circulating EPCs in patients with cardiovascular risk such as hypercholesterolemia and type 2 diabetes mellitus (T2DM). Additional in vitro study was conducted to clarify the direct effects of both statins on EPCs from the patients. Results A total of 26 patients (19 with T2DM) completed the study. While the lipid-lowering effects were similar in both treatments, the counts of circulating CD34+KDR+EPCs were significantly increased (from 0.021 ± 0.015 to 0.054 ± 0.044% of gated mononuclear cells, P < 0.05) only by pitavastatin treatment. Besides, plasma asymmetric dimethylarginine level was reduced (from 0.68 ± 0.10 to 0.53 ± 0.12 μmol/L, P < 0.05) by atorvastatin, and plasma vascular endothelial growth factor (VEGF) level was increased (from 74.33 ± 32.26 to 98.65 ± 46.64 pg/mL, P < 0.05) by pitavastatin. In the in vitro study, while both statins increased endothelial nitric oxide synthase (eNOS) expression, only pitavastatin increased the phosphorylation of eNOS in EPCs. Pitavastatin but not atorvastatin ameliorated the adhesion ability of early EPCs and the migration and tube formation capacities of late EPCs. Conclusions While both statins similarly reduced plasma lipids, only pitavastatin increased plasma VEGF level and circulating EPCs in

  3. Human umbilical cord blood-derived f-macrophages retain pluripotentiality after thrombopoietin expansion

    SciTech Connect

    Zhao Yong . E-mail: yongzhao@uic.edu; Mazzone, Theodore

    2005-11-01

    We have previously characterized a new type of stem cell from human peripheral blood, termed fibroblast-like macrophage (f-M{phi}). Here, using umbilical cord blood as a source, we identified cells with similar characteristics including expression of surface markers (CD14, CD34, CD45, CD117, and CD163), phagocytosis, and proliferative capacity. Further, thrombopoietin (TPO) significantly stimulated the proliferation of cord blood-derived f-M{phi} (CB f-M{phi}) at low dosage without inducing a megakaryocytic phenotype. Additional experiments demonstrated that TPO-expanded cord blood-derived f-M{phi} (TCB f-M{phi}) retained their surface markers and differentiation ability. Treatment with vascular endothelial cell growth factor (VEGF) gave rise to endothelial-like cells, expressing Flt-1, Flk-1, von Willebrand Factor (vWF), CD31, acetylated low density lipoprotein internalization, and the ability to form endothelial-like cell chains. In the presence of lipopolyssacharide (LPS) and 25 mM glucose, the TCB f-M{phi} differentiated to express insulin mRNA, C-peptide, and insulin. In vitro functional analysis demonstrated that these insulin-positive cells could release insulin in response to glucose and other secretagogues. These findings demonstrate a potential use of CB f-M{phi} and may lead to develop new therapeutic strategy for treating dominant disease.

  4. Ex vivo reconstitution of arterial endothelium by embryonic stem cell-derived endothelial progenitor cells in baboons.

    PubMed

    Shi, Qiang; Hodara, Vida; Simerly, Calvin R; Schatten, Gerald P; VandeBerg, John L

    2013-02-15

    There is an increasing need for an animal model that can be used to translate basic research into clinical therapy. We documented the differentiation and functional competence of embryonic stem cell (ESC)-derived endothelial cells in baboons. Baboon angioblasts were sequentially differentiated from embryoid body cultures for 9 days in an angioblast differentiation medium with varying concentrations of BMP-4, FLT-3 ligand, stem cell factor, thrombopoietin, basic fibroblast growth factor (FGF), vascular endothelial growth factor (VEGF), and knockout serum replacement. Real-time polymerase chain reaction results showed that ESC-derived angioblasts downregulated NANOG and OCT3/4, upregulated T-brachyury and GATA2, and moderately expressed CD34; they did not express CD144, TEK, or VWF, and varied in levels of CD31 expression. Several populations of putative angioblasts appeared 3 days and 9 days after differentiation, as identified by flow cytometry. Angioblasts at this stage exhibited dual paths of differentiation toward hematopoietic and vascular fates. To examine whether derived angioblasts could reconstitute the endothelium, we built an ex vivo culture system and seeded fluorescently labeled angioblast cultures onto a denuded segment of the femoral artery. We found that the seeded cells were able to grow into the endothelium on the interior surface of denuded artery segments within 5 days after seeding. After 14 days of ex vivo culture, the transplanted cells expressed CD31, an endothelial marker. The control arteries, seeded with vehicle only, did not harbor cells with endothelial markers. We conclude that ESC-derived angioblasts are promising therapeutic agents for repairing damaged vasculature, and that the baboon model will be vital for optimizing therapies for human clinical studies. PMID:22931470

  5. Bovine Posterior Limbus: An Evaluation of an Alternative Source for Corneal Endothelial and Trabecular Meshwork Stem/Progenitor Cells

    PubMed Central

    Yu, Wing Yan; Grierson, Ian; Sheridan, Carl; Lo, Amy Cheuk-Yin

    2015-01-01

    A growing body of evidence has revealed that stem-like cells in the posterior limbus of the eye between the corneal endothelium (CE) and trabecular meshwork (TM) may be able to rejuvenate these tissues in disease. However, these cells have not been clearly defined and we have named them PET cells (progenitor cells of the endothelium and trabeculum). A good and inexpensive animal model for PET cells is lacking, so we investigated bovine eyes as an effective large tissue source. We showed the presence of stem/progenitor cells in the bovine CE, transition zone, and TM in situ. Floating spheres cultured from the CE and TM showed similar stem cell marker expression patterns. Both the CE and TM spheres were bipotent and highly proliferative, but with limited secondary sphere-forming capability. They were highly prone to differentiate back into the cell type of their tissue of origin. It is speculated that the PET cells become more tissue-specific as they migrate away from their niche. Here, we showed that PET cells are present in the posterior limbus of bovine eyes and that they can be successfully cultured and expanded. PET cells represent an attractive target for developing new treatments to regenerate both the CE and TM, thereby reducing the requirement for donor tissue for corneal transplant and invasive treatments for glaucomatous patients. PMID:25323922

  6. The effects of microvesicles on endothelial progenitor cells are compromised in type 2 diabetic patients via downregulation of the miR-126/VEGFR2 pathway.

    PubMed

    Wu, Keng; Yang, Yi; Zhong, Yun; Ammar, Hala Mustafa; Zhang, Peihua; Guo, Runmin; Liu, Hua; Cheng, Chuanfang; Koroscil, Thomas M; Chen, Yanfang; Liu, Shiming; Bihl, Ji C

    2016-05-15

    Our previous study showed that circulating microvesicles (cMVs) of diabetic mice have negative effects on the function of endothelial progenitor cells (EPCs). Whether this is true in diabetic patients deserves further study. In this study, the effects of cMVs and EPC-derived MVs (EPC-MVs) on EPC migration, apoptosis, and reactive oxygen species (ROS) production in healthy controls, well-controlled, and uncontrolled diabetic patients were investigated. The levels of miR-126 and vascular endothelial growth factor receptor 2 (VEGFR2) in cMVs, EPC-MVs, and/or EPCs were analyzed. Moreover, miR-126 inhibitor or mimic was applied to EPCs to modulate the miR-126 level in EPC-MVs. We found the following: 1) the circulating EPC level was reduced but the circulating EPC-MV level increased in uncontrolled diabetic patients; 2) the cMVs and EPC-MVs of healthy controls had beneficial effects on EPCs (migration, apoptosis, ROS), whereas the effects were reversely changed in the cMVs and EPC-MVs of uncontrolled diabetic patients; and 3) the cMVs and EPC-MVs of uncontrolled diabetic patients carried less miR-126 and had downregulated VEGFR2 expression in EPCs. Manipulating the miR-126 level in EPC-MVs with inhibitor or mimic changed their function. The effects of cMVs and EPC-MVs are compromised in diabetes due to the reduction of their carried miR-126, which might provide a therapy target for diabetic vascular complications. PMID:26956185

  7. Mesenchymal stem cells and endothelial progenitor cells accelerate intra-aneurysmal tissue organization after treatment with SDF-1α-coated coils.

    PubMed

    Gao, Yuyuan; Lu, Ziming; Chen, Chengwei; Cui, Xubo; Liu, Yaqi; Zheng, Tao; Jiang, Xiaodan; Zeng, Chi; Quan, Daping; Wang, Qiujing

    2016-04-01

    Recurrences of aneurysms remain the major drawback of detachable coils for the endovascular treatment of intracranial aneurysms. The aim of the present study is to develop new modified coils, coating the surface of platinum coils with silk fibroin (SF) consisting of stromal cell-derived factor-1α (SDF-1α), and evaluate its acceleration of organization of cavities and reduction of lumen size in a rat aneurysm model. The morphological characteristics of SDF-1α-coated coils were examined using scanning electron microscopy (SEM). Fifty experimental aneurysms were created and randomly divided into five groups: three groups were embolized with SDF-1α-coated coils (8 mm) and two of these groups need transplantation of mesenchymal stem cells (MSCs) or endothelial progenitor cells (EPCs); one group was embolized with bare coils (8 mm) and another group severed as control. After coil implantation for 14 or 28 days, the coils were harvested and histological analysis was performed. SEM photographs showed that SF/SDF-1α-coated coils have uniform size and a thin film compared with bare coils. In the group treated with SDF-1α-coated coils, tissue organization was accelerated and the proliferation of α-smooth muscle actin positive cells was promoted in the aneurysmal sac. Compared with unmodified coils, on day 28, tissue organization was significantly greater in the group treated with SDF-1α-coated coils and MSC or EPC transplantation. These results suggest that SDF-1α-coated coils with MSC or EPC transplantation may be beneficial in the aneurysm healing and endothelialization at the orifice of embolized aneurysm. PMID:27125512

  8. In vitro interactions between rat bone marrow-derived endothelial progenitor cells and hepatic stellate cells: interaction between EPCs and HSCs.

    PubMed

    Liu, Feng; Liu, Zhi-da; Wu, Nan; Wang, Jiang-Hua; Zhang, Heng-Hui; Fei, Ran; Cong, Xu; Chen, Hong-song; Wei, Lai

    2013-08-01

    Transplantation of bone marrow (BM)-derived endothelial progenitor cells (EPCs) has been reported to improve liver fibrosis, but there is no direct evidence for the mechanism of improvement. We investigated the mechanism in vitro by coculturing BM-derived EPCs with activated hepatic stellate cells (HSCs) to mimic the hepatic environment. EPCs and HSCs were cultured alone and indirectly cocultured at a 1:1 ratio in a Transwell system. The characteristics of HSCs and EPCs were examined at different time points. An invasion assay showed the time-dependent effect on degradation of the extracellular matrix (ECM) layer in EPCs cultured alone. Real-time PCR and enzyme-linked immunosorbent assay analysis revealed that EPCs served as a source of matrix metalloproteinase-9 (MMP-9), and MMP-9 expression levels significantly increased during the 2 d of coculture. CFSE labeling showed that EPCs inhibited proliferation of HSCs. Annexin-V/PI staining, erminal deoxynucleotidyl transferase X-dUTP nick end labeling analysis, and (cleaved) caspase-3 activity revealed that EPCs promoted HSC apoptosis. However, the proliferation and apoptosis of EPCs were unaffected by cocultured HSCs. Coculturing increased the expression of inducible nitric oxide synthase, vascular endothelial growth factor, and hepatocyte growth factor (HGF) in EPCs, promoted differentiation of EPCs, and reduced the expression of types I and III collagens and transforming growth factor beta 1. Knockdown of HGF expression attenuated EPC-induced activation of HSC apoptosis and profibrotic ability. These findings demonstrated that BM-derived EPCs could degrade ECM, promoting activated HSC apoptosis, suppressing proliferation and profibrotic ability of activated HSCs. HGF secretion by EPCs plays a key role in inducing activated HSC apoptosis and HSC profibrotic ability. PMID:23722413

  9. Co-Transplantation of Endothelial Progenitor Cells and Pancreatic Islets to Induce Long-Lasting Normoglycemia in Streptozotocin-Treated Diabetic Rats

    PubMed Central

    Spiga, Saturnino; Mazzanti, Benedetta; Curcio, Michele; Mulas, Giovanna; Diana, Marco; Marzola, Pasquina; Mosca, Franco; Longoni, Biancamaria

    2014-01-01

    Graft vascularization is a crucial step to obtain stable normoglycemia in pancreatic islet transplantation. Endothelial progenitor cells (EPCs) contribute to neoangiogenesis and to the revascularization process during ischaemic events and play a key role in the response to pancreatic islet injury. In this work we co-transplanted EPCs and islets in the portal vein of chemically-induced diabetic rats to restore islet vascularization and to improve graft survival. Syngenic islets were transplanted, either alone or with EPCs derived from green fluorescent protein (GFP) transgenic rats, into the portal vein of streptozotocin-induced diabetic rats. Blood glucose levels were monitored and intraperitoneal glucose tolerance tests were performed. Real time-PCR was carried out to evaluate the gene expression of angiogenic factors. Diabetic-induced rats showed long-lasting (6 months) normoglycemia upon co-transplantation of syngenic islets and EPCs. After 3–5 days from transplantation, hyperglycaemic levels dropped to normal values and lasted unmodified as long as they were checked. Further, glucose tolerance tests revealed the animals' ability to produce insulin on-demand as indexed by a prompt response in blood glucose clearance. Graft neovascularization was evaluated by immunohistochemistry: for the first time the measure of endothelial thickness revealed a donor-EPC-related neovascularization supporting viable islets up to six months after transplant. Our results highlight the importance of a newly formed viable vascular network together with pancreatic islets to provide de novo adequate supply in order to obtain enduring normoglycemia and prevent diabetes-related long-term health hazards. PMID:24733186

  10. Co-transplantation of endothelial progenitor cells and pancreatic islets to induce long-lasting normoglycemia in streptozotocin-treated diabetic rats.

    PubMed

    Quaranta, Paola; Antonini, Sara; Spiga, Saturnino; Mazzanti, Benedetta; Curcio, Michele; Mulas, Giovanna; Diana, Marco; Marzola, Pasquina; Mosca, Franco; Longoni, Biancamaria

    2014-01-01

    Graft vascularization is a crucial step to obtain stable normoglycemia in pancreatic islet transplantation. Endothelial progenitor cells (EPCs) contribute to neoangiogenesis and to the revascularization process during ischaemic events and play a key role in the response to pancreatic islet injury. In this work we co-transplanted EPCs and islets in the portal vein of chemically-induced diabetic rats to restore islet vascularization and to improve graft survival. Syngenic islets were transplanted, either alone or with EPCs derived from green fluorescent protein (GFP) transgenic rats, into the portal vein of streptozotocin-induced diabetic rats. Blood glucose levels were monitored and intraperitoneal glucose tolerance tests were performed. Real time-PCR was carried out to evaluate the gene expression of angiogenic factors. Diabetic-induced rats showed long-lasting (6 months) normoglycemia upon co-transplantation of syngenic islets and EPCs. After 3-5 days from transplantation, hyperglycaemic levels dropped to normal values and lasted unmodified as long as they were checked. Further, glucose tolerance tests revealed the animals' ability to produce insulin on-demand as indexed by a prompt response in blood glucose clearance. Graft neovascularization was evaluated by immunohistochemistry: for the first time the measure of endothelial thickness revealed a donor-EPC-related neovascularization supporting viable islets up to six months after transplant. Our results highlight the importance of a newly formed viable vascular network together with pancreatic islets to provide de novo adequate supply in order to obtain enduring normoglycemia and prevent diabetes-related long-term health hazards. PMID:24733186

  11. Circulating endothelial progenitor cells and residual in vivo thromboxane biosynthesis in low-dose aspirin-treated polycythemia vera patients.

    PubMed

    Santilli, Francesca; Romano, Mario; Recchiuti, Antonio; Dragani, Alfredo; Falco, Angela; Lessiani, Gianfranco; Fioritoni, Francesca; Lattanzio, Stefano; Mattoscio, Domenico; De Cristofaro, Raimondo; Rocca, Bianca; Davì, Giovanni

    2008-08-15

    Polycythemia vera (PV) is associated with high morbidity and mortality for thrombosis. We hypothesized that in PV altered sensitivity to aspirin might be related to dysfunction of the endothelial repair and/or of the nitric oxide (NO) system. Urinary thromboxane (TX) A(2) metabolite (TXM), endothelial colony-forming cells (ECFCs), plasma asymmetric dimethylarginine (ADMA) and von Willebrand factor (VWF) were measured in 37 PV patients on low-dose aspirin and 12 healthy controls. Patients showed an approximately 2-fold increase in median TXM and plasma ADMA levels (P < .001), while ECFC numbers were reduced by approximately 7-fold (P < .001) as compared with non-aspirinated control. These differences were more pronounced in patients with previous thrombosis. An 8-week course of aspirin did not affect ECFCs in 6 controls. VWF and TXM correlated directly with ADMA, and inversely with ECFCs. By multiple regression analysis, lower ECFC quartiles (beta = -0.39; SE = 0.17; P = .028) and higher VWF levels (beta = 0.338, SE = 0.002, P = .034) were independent predictors of higher TXM quartiles (R(2) = 0.39). Serum TXB(2), measured in 22 patients, was approximately 10-fold higher than aspirin-treated controls. PV patients appear to have an unbalanced ECFC/NO axis, and an apparent altered sensitivity of platelet TXA(2) production, all potentially contributing to aspirin-insensitive TXM formation. Thus, additional antithrombotic strategies may be beneficial in PV. PMID:18541722

  12. Endothelial progenitor cells, defined by the simultaneous surface expression of VEGFR2 and CD133, are not detectable in healthy peripheral and cord blood.

    PubMed

    Lanuti, Paola; Rotta, Gianluca; Almici, Camillo; Avvisati, Giuseppe; Budillon, Alfredo; Doretto, Paolo; Malara, Natalia; Marini, Mirella; Neva, Arabella; Simeone, Pasquale; Di Gennaro, Elena; Leone, Alessandra; Falda, Alessandra; Tozzoli, Renato; Gregorj, Chiara; Di Cerbo, Melania; Trunzo, Valentina; Mollace, Vincenzo; Marchisio, Marco; Miscia, Sebastiano

    2016-03-01

    Circulating endothelial cells (CEC) and their progenitors (EPC) are restricted subpopulations of peripheral blood (PB), cord blood (CB), and bone marrow (BM) cells, involved in the endothelial homeostasis maintenance. Both CEC and EPC are thought to represent potential biomarkers in several clinical conditions involving endothelial turnover/remodeling. Although different flow cytometry methods for CEC and EPC characterization have been published so far, none of them have reached consistent conclusions, therefore consensus guidelines with respect to CEC and EPC identification and quantification need to be established. Here, we have carried out an in depth investigation of CEC and EPC phenotypes in healthy PB, CB and BM samples, by optimizing a reliable polychromatic flow cytometry (PFC) panel. Results showed that the brightness of CD34 expression on healthy PB and CB circulating cells represents a key benchmark for the identification of CEC (CD45neg/CD34bright/CD146pos) respect to the hematopoietic stem cell (HSC) compartment (CD45dim/CD34pos/CD146neg). This approach, combined with a dual-platform counting technique, allowed a sharp CEC enumeration in healthy PB (n = 38), and resulting in consistent CEC counts with previously reported data (median = 11.7 cells/ml). In parallel, by using rigorous PFC conditions, CD34pos/CD45dim/CD133pos/VEGFR2pos EPC were not found in any healthy PB or CB sample, since VEGFR2 expression was never detectable on the surface of CD34pos/CD45dim/CD133pos cells. Notably, the putative EPC phenotype was observed in all analyzed BM samples (n = 12), and the expression of CD146 and VEGFR2, on BM cells, was not restricted to the CD34bright compartment, but also appeared on the HSC surface. Altogether, our findings suggest that the previously reported EPC antigen profile, defined by the simultaneous expression of VEGFR2 and CD133 on the surface of CD45dim/CD34pos cells, should be carefully re-evaluated and further studies should be conducted to

  13. tPA-MMP-9 Axis Plays a Pivotal Role in Mobilization of Endothelial Progenitor Cells from Bone Marrow to Circulation and Ischemic Region for Angiogenesis

    PubMed Central

    Day, Yuan-Ji

    2016-01-01

    We examined the role of tissue plasminogen activator- (tPA-) matrix metalloproteinase- (MMP-) 9 in mobilizing endothelial progenitor cells (EPCs) from bone marrow to circulation and critical limb ischemia (CLI) region. Male C57BL/6J mice having been irradiated were categorized into wild-type mice (WT) receiving WT bone marrow cell (BMC) transfusion (group 1), WT mice receiving MMP-9 knockout (MMP-9−/−) BMC (group 2), MMP-9−/− receiving MMP-9−/− BMC (group 3), and MMP-9−/− receiving WT BMC (group 4), each of which was subdivided into sham control (SC), CLI, SC-tPA, and CLI-tPA. In groups 1 and 4, by post-CLI 18 h and day 14, circulating EPC (C-kit+/CD31+, Sca-1+/KDR+) levels were highest in CLI-tPA subgroup. In groups 2 and 3, EPC levels did not differ among all subgroups. The EPC levels in bone marrow were higher in groups 2 and 3 than those in groups 1 and 4. By day 14, in animals with CLI, expression levels of proangiogenic factors (CXCR4, SDF-1α, and VEGF) showed similar trends as circulating EPC levels. Moreover, the number of infiltrated neutrophils and macrophages in quadriceps was higher in groups 1 and 4 than groups in 2 and 3. In conclusion, tPA-MMP-9 axis plays a crucial role in EPC mobilization and angiogenesis in experimental CLI. PMID:27610138

  14. Effect of Periodic Granulocyte Colony-Stimulating Factor Administration on Endothelial Progenitor Cells and Different Monocyte Subsets in Pediatric Patients with Muscular Dystrophies

    PubMed Central

    Sienkiewicz, Dorota; Grubczak, Kamil; Okurowska-Zawada, Bożena; Paszko-Patej, Grażyna; Miklasz, Paula; Singh, Paulina; Radzikowska, Urszula; Kulak, Wojciech

    2016-01-01

    Muscular dystrophies (MD) are heterogeneous group of diseases characterized by progressive muscle dysfunction. There is a large body of evidence indicating that angiogenesis is impaired in muscles of MD patients. Therefore, induction of dystrophic muscle revascularization should become a novel approach aimed at diminishing the extent of myocyte damage. Recently, we and others demonstrated that administration of granulocyte colony-stimulating factor (G-CSF) resulted in clinical improvement of patients with neuromuscular disorders. To date, however, the exact mechanisms underlying these beneficial effects of G-CSF have not been fully understood. Here we used flow cytometry to quantitate numbers of CD34+ cells, endothelial progenitor cells, and different monocyte subsets in peripheral blood of pediatric MD patients treated with repetitive courses of G-CSF administration. We showed that repetitive cycles of G-CSF administration induced efficient mobilization of above-mentioned cells including cells with proangiogenic potential. These findings contribute to better understanding the beneficial clinical effects of G-CSF in pediatric MD patients. PMID:26770204

  15. Direct Cell-Cell Contact between Mesenchymal Stem Cells and Endothelial Progenitor Cells Induces a Pericyte-Like Phenotype In Vitro

    PubMed Central

    Richards, R. Geoff; Nerlich, Michael; Alini, Mauro

    2014-01-01

    Tissue engineering techniques for the regeneration of large bone defects require sufficient vascularisation of the applied constructs to ensure a sufficient supply of oxygen and nutrients. In our previous work, prevascularised 3D scaffolds have been successfully established by coculture of bone marrow derived stem cells (MSCs) and endothelial progenitor cells (EPCs). We identified stabilising pericytes (PCs) as part of newly formed capillary-like structures. In the present study, we report preliminary data on the interactions between MSCs and EPCs, leading to the differentiation of pericyte-like cells. MSCs and EPCs were seeded in transwell cultures, direct cocultures, and single cultures. Cells were cultured for 10 days in IMDM 10% FCS or IMDM 5% FCS 5% platelet lysate medium. Gene expression of PC markers, CD146, NG2, αSMA, and PDGFR-β, was analysed using RT-PCR at days 0, 3, 7, and 10. The upregulation of CD146, NG2, and αSMA in MSCs in direct coculture with EPCs advocates the MSCs' differentiation towards a pericyte-like phenotype in vitro. These results suggest that pericyte-like cells derive from MSCs and that cell-cell contact with EPCs is an important factor for this differentiation process. These findings emphasise the concept of coculture strategies to promote angiogenesis for cell-based tissue engineered bone grafts. PMID:24563864

  16. tPA-MMP-9 Axis Plays a Pivotal Role in Mobilization of Endothelial Progenitor Cells from Bone Marrow to Circulation and Ischemic Region for Angiogenesis.

    PubMed

    Leu, Steve; Day, Yuan-Ji; Sun, Cheuk-Kwan; Yip, Hon-Kan

    2016-01-01

    We examined the role of tissue plasminogen activator- (tPA-) matrix metalloproteinase- (MMP-) 9 in mobilizing endothelial progenitor cells (EPCs) from bone marrow to circulation and critical limb ischemia (CLI) region. Male C57BL/6J mice having been irradiated were categorized into wild-type mice (WT) receiving WT bone marrow cell (BMC) transfusion (group 1), WT mice receiving MMP-9 knockout (MMP-9(-/-)) BMC (group 2), MMP-9(-/-) receiving MMP-9(-/-) BMC (group 3), and MMP-9(-/-) receiving WT BMC (group 4), each of which was subdivided into sham control (SC), CLI, SC-tPA, and CLI-tPA. In groups 1 and 4, by post-CLI 18 h and day 14, circulating EPC (C-kit+/CD31+, Sca-1+/KDR+) levels were highest in CLI-tPA subgroup. In groups 2 and 3, EPC levels did not differ among all subgroups. The EPC levels in bone marrow were higher in groups 2 and 3 than those in groups 1 and 4. By day 14, in animals with CLI, expression levels of proangiogenic factors (CXCR4, SDF-1α, and VEGF) showed similar trends as circulating EPC levels. Moreover, the number of infiltrated neutrophils and macrophages in quadriceps was higher in groups 1 and 4 than groups in 2 and 3. In conclusion, tPA-MMP-9 axis plays a crucial role in EPC mobilization and angiogenesis in experimental CLI. PMID:27610138

  17. Inhibition of p38 Mitogen-Activated Protein Kinase Enhances the Apoptosis Induced by Oxidized Low-Density Lipoprotein in Endothelial Progenitor Cells.

    PubMed

    Tie, Guodong; Yan, Jinglian; Messina, Julia A; Raffai, Robert L; Messina, Louis M

    2015-01-01

    Oxidized low-density lipoprotein (oxLDL) is an important risk factor in the development of atherosclerosis. oxLDL has been shown to decrease endothelial progenitor cell (EPC) number by inducing apoptosis. p38 mitogen-activated protein kinase (MAPK) was shown to be activated by oxLDL and participated in the regulation of EPC number and function. However, the role of p38 remains unknown. Here, we show that oxLDL-induced p38 phosphorylation in EPCs is time and dose dependent. Treatment with antioxidant N-acetyl cysteine restored oxLDL-induced p38 phosphorylation to basal levels. LOX-1-blocking antibody also significantly decreased oxLDL-induced p38 phosphorylation. Interestingly, TUNEL staining showed that pretreatment with the p38 inhibitor SB203580 further increased oxLDL-induced apoptosis in EPCs. In accordance with these findings, pretreatment with SB203580 further attenuated Akt phosphorylation in EPCs challenged with oxLDL, indicating an interaction between Akt and p38 MAPK pathways. In agreement, inhibition of p38 MAPK further attenuated Akt phosphorylation and increased apoptosis in EPCs isolated from hypercholesterolemic ApoE-/- mice. In conclusion, p38 MAPK serves as an anti-apoptotic pathway by supporting Akt activity when EPCs are challenged with oxLDL. PMID:27031525

  18. Skeletal Myogenic Progenitors Originating from Embryonic Dorsal Aorta Coexpress Endothelial and Myogenic Markers and Contribute to Postnatal Muscle Growth and Regeneration

    PubMed Central

    De Angelis, Luciana; Berghella, Libera; Coletta, Marcello; Lattanzi, Laura; Zanchi, Malvina; Gabriella, M.; Ponzetto, Carola; Cossu, Giulio

    1999-01-01

    Skeletal muscle in vertebrates is derived from somites, epithelial structures of the paraxial mesoderm, yet many unrelated reports describe the occasional appearance of myogenic cells from tissues of nonsomite origin, suggesting either transdifferentiation or the persistence of a multipotent progenitor. Here, we show that clonable skeletal myogenic cells are present in the embryonic dorsal aorta of mouse embryos. This finding is based on a detailed clonal analysis of different tissue anlagen at various developmental stages. In vitro, these myogenic cells show the same morphology as satellite cells derived from adult skeletal muscle, and express a number of myogenic and endothelial markers. Surprisingly, the latter are also expressed by adult satellite cells. Furthermore, it is possible to clone myogenic cells from limbs of mutant c-Met−/− embryos, which lack appendicular muscles, but have a normal vascular system. Upon transplantation, aorta-derived myogenic cells participate in postnatal muscle growth and regeneration, and fuse with resident satellite cells. The potential of the vascular system to generate skeletal muscle cells may explain observations of nonsomite skeletal myogenesis and raises the possibility that a subset of satellite cells may derive from the vascular system. PMID:10562287

  19. Effect of Periodic Granulocyte Colony-Stimulating Factor Administration on Endothelial Progenitor Cells and Different Monocyte Subsets in Pediatric Patients with Muscular Dystrophies.

    PubMed

    Eljaszewicz, Andrzej; Sienkiewicz, Dorota; Grubczak, Kamil; Okurowska-Zawada, Bożena; Paszko-Patej, Grażyna; Miklasz, Paula; Singh, Paulina; Radzikowska, Urszula; Kulak, Wojciech; Moniuszko, Marcin

    2016-01-01

    Muscular dystrophies (MD) are heterogeneous group of diseases characterized by progressive muscle dysfunction. There is a large body of evidence indicating that angiogenesis is impaired in muscles of MD patients. Therefore, induction of dystrophic muscle revascularization should become a novel approach aimed at diminishing the extent of myocyte damage. Recently, we and others demonstrated that administration of granulocyte colony-stimulating factor (G-CSF) resulted in clinical improvement of patients with neuromuscular disorders. To date, however, the exact mechanisms underlying these beneficial effects of G-CSF have not been fully understood. Here we used flow cytometry to quantitate numbers of CD34+ cells, endothelial progenitor cells, and different monocyte subsets in peripheral blood of pediatric MD patients treated with repetitive courses of G-CSF administration. We showed that repetitive cycles of G-CSF administration induced efficient mobilization of above-mentioned cells including cells with proangiogenic potential. These findings contribute to better understanding the beneficial clinical effects of G-CSF in pediatric MD patients. PMID:26770204

  20. Bradykinin inhibits oxidative stress-induced senescence of endothelial progenitor cells through the B2R/AKT/RB and B2R/EGFR/RB signal pathways.

    PubMed

    Fu, Cong; Li, Bing; Sun, Yuning; Ma, Genshan; Yao, Yuyu

    2015-09-22

    Circulating endothelial progenitor cells (EPCs) have multiple protective effects that facilitate repair of damage to tissues and organs. However, while various stressors are known to impair EPC function, the mechanisms of oxidative stress-induced EPC senescence remains unknown. We demonstrated that B2 receptor (B2R) expression on circulating CD34(+) cells was significantly reduced in patients with diabetes mellitus (DM) as compared to healthy controls. Furthermore, CD34(+) cell B2R expression in patients with DM was inversely correlated with plasma myeloperoxidase concentrations. Bradykinin (BK) treatment decreased human EPC (hEPC) senescence and intracellular oxygen radical production, resulting in reduced retinoblastoma 1 (RB) RNA expression in H2O2-induced senescent hEPCs and a reversal of the B2R downregulation that is normally observed in senescent cells. Furthermore, BK treatment of H2O2-exposed cells leads to elevated phosphorylation of RB, AKT, and cyclin D1 compared with H2O2-treatment alone. Antagonists of B2R, PI3K, and EGFR signaling pathways and B2R siRNA blocked BK protective effects. In summary, this study demonstrates that BK significantly inhibits oxidative stress-induced hEPC senescence though B2R-mediated activation of PI3K and EGFR signaling pathways. PMID:26360782

  1. Cell-Surface MMP-9 Protein Is a Novel Functional Marker to Identify and Separate Proangiogenic Cells from Early Endothelial Progenitor Cells Derived from CD133(+) Cells.

    PubMed

    Kanayasu-Toyoda, Toshie; Tanaka, Takeshi; Kikuchi, Yutaka; Uchida, Eriko; Matsuyama, Akifumi; Yamaguchi, Teruhide

    2016-05-01

    To develop cell therapies for ischemic diseases, endothelial progenitor cells (EPCs) have been expected to play a pivotal role in vascular regeneration. It is desirable to use a molecular marker that is related to the function of the cells. Here, a quantitative polymerase chain reaction array revealed that early EPCs derived from CD133(+) cells exhibited significant expression of MMP-9. Some populations of early EPCs expressed MMP-9 on the cell surface and others did not. We also attempted to separate the proangiogenic fraction from early EPCs derived from CD133(+) cells using a functional cell surface marker, and we then analyzed the MMP-9(+) and MMP-9(-) cell fractions. The MMP-9(+) cells not only revealed higher invasion ability but also produced a high amount of IL-8. Moreover, the stimulative effect of MMP-9(+) cells on angiogenesis in vitro and in vivo was prohibited by anti-IL-8 antibody. These data indicate that MMP-9 is one of the useful cell surface markers for the separation of angiogenic cells. Our treatment of early EPCs with hyaluronidase caused not only a downregulation of cell-surface MMP-9 but also a decrease in invasion ability, indicating that membrane-bound MMP-9, which is one of the useful markers for early EPCs, plays an important role in angiogenesis. Stem Cells 2016;34:1251-1262. PMID:26824798

  2. Visfatin attenuates the ox-LDL-induced senescence of endothelial progenitor cells by upregulating SIRT1 expression through the PI3K/Akt/ERK pathway.

    PubMed

    Ming, Guang-Feng; Tang, Yong-Jun; Hu, Kai; Chen, Yao; Huang, Wei-Hua; Xiao, Jian

    2016-08-01

    Endothelial progenitor cells (EPCs) play an important role in aging-associated senescence, thereby potentially contributing to vascular pathologies. Visfatin, identified as a new adipocytokine, is closely associated with the senescence of human cells. However, the effects of visfatin on the oxidized low-density lipoprotein (ox-LDL)-induced senescence of EPCs has not yet been explored, to the best of our knowledge. For this purpose, in the present study, we examined the effects of visfatin in ox-LDL-stimulated EPCs as well as the underlying mechanism responsible for these effects. We found that visfatin attenuated the ox-LDL-induced senescence of EPCs by repressing β-galactosidase expression and recovering telomerase activity. Western blot analysis confirmed that visfatin induced a dose-dependent increase in sirtuin 1 (SIRT1) expression in EPCs and ox-LDL exposure decreased SIRT1 expression. Silencing SIRT1 abolished the inhibition of EPC senescence and the suppression of p53 expression induced by visfatin. Moreover, visfatin attenuated the inhibition of phosphorylation of Akt, phosphoinositide-3-kinase (PI3K) and extracellular signal-regulated kinase (ERK) induced by ox-LDL. Taken together, these findings suggest that the treatment of EPCs with visfatin markedly attenuates the ox-LDL-induced senescence of EPCs by upregulating SIRT1 expression through the PI3K/Akt/ERK pathway. PMID:27277186

  3. NAMPT regulates senescence, proliferation, and migration of endothelial progenitor cells through the SIRT1 AS lncRNA/miR-22/SIRT1 pathway.

    PubMed

    Ming, Guang-Feng; Wu, Kai; Hu, Kai; Chen, Yao; Xiao, Jian

    2016-09-23

    The importance of endothelial progenitor cells (EPCs) in cardiovascular diseases has been demonstrated by numerous studies. Previous studies have shown that Nicotinamide phosphoribosyltransferase (NAMPT) plays a role in EPC development by regulating Sirtuin 1 (SIRT1), but the specific mechanism has not yet been elucidated. After stimulating EPCs with NAMPT, expression of SIRT1 and SIRT1 antisense long non-coding RNA (AS lncRNA) was upregulated. Upon transfection of an SIRT1 AS lncRNA overexpression vector into EPCs, SIRT1 expression was upregulated. Upon transfection of a small interfering RNA (siRNA) that targets SIRT1 AS lncRNA along with NAMPT, SIRT1 AS lncRNA was downregulated and NAMPT-induced SIRT1 expression was reduced. We used software analyses and a dual-luciferase reporter assay to demonstrate that microRNA (miR)-22 regulated SIRT1 and SIRT1 AS lncRNA. Our data suggest that SIRT1 AS lncRNA relieves miR-22-induced SIRT1 downregulation by competitively sponging miR-22. By measuring EPC senescence, proliferation, and migration, we found that NAMPT inhibited EPC senescence through an SIRT1 AS lncRNA/miR-22/SIRT1 pathway and promoted EPC proliferation and migration. These findings provide a new theoretical basis for the prevention and treatment of atherosclerosis (AS) and other cardiovascular diseases. PMID:27569277

  4. The Effect of Simulated Microgravity on Differentiation of Porcine Blood-Derived Vascular Stem Cells.

    PubMed

    Ramaswamy, Vidhya; Dirr, Elliott W; Allen, Josephine B

    2016-02-15

    The negative effects of space flight on cardiovascular health of astronauts have been demonstrated and documented over many years. Endothelial cells (ECs) play an important role in regulating weightlessness-induced cardiovascular dysfunction. This project seeks to study the effect of microgravity on the differentiation of endothelial progenitor cells (EPCs) into ECs and the downstream functions of the differentiated cells. Initial exposure of EPCs to microgravity indicated an inhibition of migratory, proliferative, and antithrombogenic capacity of the differentiated ECs. However, our results indicate a potential recovery of proliferative and antithrombogenic functions on prolonged exposure to microgravity. PMID:26649474

  5. Endothelial Progenitor Cell Dysfunction in Myelodysplastic Syndromes: Possible Contribution of a Defective Vascular Niche to Myelodysplasia12

    PubMed Central

    Teofili, Luciana; Martini, Maurizio; Nuzzolo, Eugenia Rosa; Capodimonti, Sara; Iachininoto, Maria Grazia; Cocomazzi, Alessandra; Fabiani, Emiliano; Voso, Maria Teresa; Larocca, Luigi M.

    2015-01-01

    We set a model to replicate the vascular bone marrow niche by using endothelial colony forming cells (ECFCs), and we used it to explore the vascular niche function in patients with low-risk myelodysplastic syndromes (MDS). Overall, we investigated 56 patients and we observed higher levels of ECFCs in MDS than in healthy controls; moreover, MDS ECFCs were found variably hypermethylated for p15INK4b DAPK1, CDH1, or SOCS1. MDS ECFCs exhibited a marked adhesive capacity to normal mononuclear cells. When normal CD34 + cells were co-cultured with MDS ECFCs, they generated significant lower amounts of CD11b + and CD41 + cells than in co-culture with normal ECFCs. At gene expression profile, several genes involved in cell adhesion were upregulated in MDS ECFCs, while several members of the Wingless and int (Wnt) pathways were underexpressed. Furthermore, at miRNA expression profile, MDS ECFCs hypo-expressed various miRNAs involved in Wnt pathway regulation. The addition of Wnt3A reduced the expression of intercellular cell adhesion molecule-1 on MDS ECFCs and restored the defective expression of markers of differentiation. Overall, our data demonstrate that in low-risk MDS, ECFCs exhibit various primary abnormalities, including putative MDS signatures, and suggest the possible contribution of the vascular niche dysfunction to myelodysplasia. PMID:26025663

  6. Vasculogenic Conditioning of Peripheral Blood Mononuclear Cells Promotes Endothelial Progenitor Cell Expansion and Phenotype Transition of Anti‐Inflammatory Macrophage and T Lymphocyte to Cells With Regenerative Potential

    PubMed Central

    Masuda, Haruchika; Tanaka, Rica; Fujimura, Satoshi; Ishikawa, Masakazu; Akimaru, Hiroshi; Shizuno, Tomoko; Sato, Atsuko; Okada, Yoshinori; Iida, Yumi; Itoh, Jobu; Itoh, Yoshiko; Kamiguchi, Hiroshi; Kawamoto, Atsuhiko; Asahara, Takayuki

    2014-01-01

    Background Cell‐based therapies involving mononuclear cells (MNCs) have been developed for vascular regeneration to treat ischemic diseases; however, quality control of therapeutic MNCs has not been evaluated. We investigated the therapeutic potential of peripheral blood (PB) MNCs, operated by recently developed quality and quantity (QQ) culture of endothelial progenitor cells (EPCs). Methods and Results PBs were collected from healthy volunteers; peripheral blood mononuclear cells (PBMNCs) isolated from these PBs were subjected to QQ culture for 7 days with medium containing stem cell factor, thrombopoietin, Flt‐3 ligand, vascular endothelial growth factor, and interleukin‐6. The resulting cells (QQMNCs) in EPC colony‐forming assay generated significantly more definitive EPC colonies than PBMNCs. In flow cytometry, macrophages and helper T lymphocytes of QQMNCs became phenotypically polarized into angiogenic, anti‐inflammatory, and regenerative subsets: classical M1 to alternative M2; T helper (Th)1 to Th2; angiogenic or regulatory T‐cell expansion. Quantitative real‐time polymerase chain reaction (qRT‐PCR) assay revealed the predominant proangiogenic gene expressions in QQMNCs versus PBMNCs. Using murine ischemic hindlimb models, the efficacy of QQMNC intramuscular transplantation (Tx) was compared to that of PBMNCTx, cultured “early EPC” Tx (eEPCTx), and granulocyte colony‐stimulating factor mobilized CD34+ cell Tx (GmCD34Tx). Laser Doppler imaging revealed the blood perfusion recovery in ischemic hindlimbs after QQMNCTx superior to after PBMNCTx and eEPCTx, but also earlier than after GmCD34Tx. Histological evaluations and qRT‐PCR assays in ischemic hindlimbs demonstrated that QQMNCTx, similarly to GmCD34Tx, enhanced angiovasculogenesis and myogenesis, whereas it preponderantly inhibited inflammation and fibrosis versus PBMNCTx and eEPCTx. Conclusions QQ culture potentiates the ability of PBMNCs to promote regeneration of injured tissue

  7. Sleep-disordered breathing is associated with depletion of circulating endothelial progenitor cells and elevation in pulmonary arterial pressure in patients with decompensated systolic heart failure

    PubMed Central

    Zhang, Han; Feng, Liu; Wan, Qi-Lin; Hong, Yan; Li, Yan-Ming; Cheng, Guan-Chang; Han, Xin-Qiang

    2015-01-01

    Background Sleep-disordered breathing (SDB) is known to occur frequently in and may predict worsening progression of patients with congestive heart failure (CHF). SDB is also known to play an important role in the development of idiopathic pulmonary arterial hypertension (PAH) via inducing endothelial dysfunction and vascular remodeling, a pathological process that can be significantly influenced by factors such as osteoprotegerin (OPG) and endothelial progenitor cells (EPCs). The objective of this study is to determine if CHF with SDB is associated with changes in OPG, EPCs, and PAH. Methods EPCs were isolated, cultured, and quantified from CHF patients with SDB (n = 52), or without SDB (n = 68). OPG and N-terminal pro-brain natriuretic peptide (NT-proBNP) from each group was analyzed and correlated with EPCs and the mean pulmonary artery pressure (mPAP) measured by right heart catheterization. Results A significant decrease in circulating EPCs (29.30 ± 9.01 vs. 45.17 ± 10.51 EPCs/× 200 field; P < 0.05) was found in CHF patients with SDB compared to those without SDB. Both OPG (789.83 ± 89.38 vs. 551.29 ± 42.12 pg/mL; P < 0.05) and NT-proBNP (5946.50 ± 1434.50 vs. 3028.60 ± 811.90 ng/mL; P < 0.05) were also significantly elevated in SDB CHF patients who also had significantly elevated mPAP (50.2 ± 9.5 vs. 36.4 ± 4.1 mm Hg; P < 0.05). EPC numbers correlated inversely with the episodes of apnea and hypopnea per hour (RDI, r = –0.45, P = 0.037) and blood level of OPG (r = –0.53, P = 0.011). Although NT-proBNP was also increased significantly in patients with SDB, it had no correlation with either EPCs or RDI. Conclusions SDB due to hypoxemia from decompensated CHF is associated with (1) OPG elevation, (2) EPC depletion, and (3) mPAP elevation. The inverse relationship of circulating OPG with EPCs suggests a likely mechanism for hypoxemia and OPG in the development of pulmonary vascular dysfunction via depleting EPCs, thus worsening prognosis of CHF. PMID

  8. Black Raspberry Extract Increased Circulating Endothelial Progenitor Cells and Improved Arterial Stiffness in Patients with Metabolic Syndrome: A Randomized Controlled Trial.

    PubMed

    Jeong, Han Saem; Kim, Sohyeon; Hong, Soon Jun; Choi, Seung Cheol; Choi, Ji-Hyun; Kim, Jong-Ho; Park, Chi-Yeon; Cho, Jae Young; Lee, Tae-Bum; Kwon, Ji-Wung; Joo, Hyung Joon; Park, Jae Hyoung; Yu, Cheol Woong; Lim, Do-Sun

    2016-04-01

    Administration of black raspberry (Rubus occidentalis) is known to improve vascular endothelial function in patients at a high risk for cardiovascular (CV) disease. We investigated short-term effects of black raspberry on circulating endothelial progenitor cells (EPCs) and arterial stiffness in patients with metabolic syndrome. Patients with metabolic syndrome (n = 51) were prospectively randomized into the black raspberry group (n = 26, 750 mg/day) and placebo group (n = 25) during the 12-week follow-up. Central blood pressure, augmentation index, and EPCs, such as CD34/KDR(+), CD34/CD117(+), and CD34/CD133(+), were measured at baseline and at 12-week follow-up. Radial augmentation indexes were significantly decreased in the black raspberry group compared to the placebo group (-5% ± 10% vs. 3% ± 14%, P < .05). CD34/CD133(+) cells at 12-week follow-up were significantly higher in the black raspberry group compared to the placebo group (19 ± 109/μL vs. -28 ± 57/μL, P < .05). Decreases from the baseline in interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-α) were significantly greater in the black raspberry group compared to the placebo group (-0.5 ± 1.4 pg/mL vs. -0.1 ± 1.1 pg/mL, P < .05 and -5.4 ± 4.5 pg/mL vs. -0.8 ± 4.0 pg/mL, P < .05, respectively). Increases from the baseline in adiponectin levels (2.9 ± 2.1 μg/mL vs. -0.2 ± 2.5 μg/mL, P < .05) were significant in the black raspberry group. The use of black raspberry significantly lowered the augmentation index and increased circulating EPCs, thereby improving CV risks in patients with metabolic syndrome during the 12-week follow-up. PMID:26891216

  9. Blood-derived topical therapy for ocular surface diseases.

    PubMed

    Soni, Nishant G; Jeng, Bennie H

    2016-01-01

    Human serum-derived and plasma-derived therapies have become increasingly popular in the treatment of ocular surface disorders, with mounting clinical and scientific evidence suggesting good safety and efficacy profiles. These therapies may be considered for various ocular surface conditions, such as dry eye syndrome and persistent epithelial defect, when conservative management does not suffice. The costly and inconvenient process of obtaining the blood-derived products is the barrier to their more widespread use. Some blood-derived therapies, such as umbilical cord serum-derived and platelet-derived plasma preparations, may be more viable options since these therapies can be made readily available to patients. In this review, the existing literature on the safety and efficacy of blood-derived products, such as autologous serum tears, in the treatment of ocular surface diseases is discussed. Issues relevant to the production of autologous serum tears are also described. PMID:26178904

  10. Differential uPAR recruitment in caveolar-lipid rafts by GM1 and GM3 gangliosides regulates endothelial progenitor cells angiogenesis.

    PubMed

    Margheri, Francesca; Papucci, Laura; Schiavone, Nicola; D'Agostino, Riccardo; Trigari, Silvana; Serratì, Simona; Laurenzana, Anna; Biagioni, Alessio; Luciani, Cristina; Chillà, Anastasia; Andreucci, Elena; Del Rosso, Tommaso; Margheri, Giancarlo; Del Rosso, Mario; Fibbi, Gabriella

    2015-01-01

    Gangliosides and the urokinase plasminogen activator receptor (uPAR) tipically partition in specialized membrane microdomains called lipid-rafts. uPAR becomes functionally important in fostering angiogenesis in endothelial progenitor cells (EPCs) upon recruitment in caveolar-lipid rafts. Moreover, cell membrane enrichment with exogenous GM1 ganglioside is pro-angiogenic and opposite to the activity of GM3 ganglioside. On these basis, we first checked the interaction of uPAR with membrane models enriched with GM1 or GM3, relying on the adoption of solid-supported mobile bilayer lipid membranes with raft-like composition formed onto solid hydrophilic surfaces, and evaluated by surface plasmon resonance (SPR) the extent of uPAR recruitment. We estimated the apparent dissociation constants of uPAR-GM1/GM3 complexes. These preliminary observations, indicating that uPAR binds preferentially to GM1-enriched biomimetic membranes, were validated by identifying a pro-angiogenic activity of GM1-enriched EPCs, based on GM1-dependent uPAR recruitment in caveolar rafts. We have observed that addition of GM1 to EPCs culture medium promotes matrigel invasion and capillary morphogenesis, as opposed to the anti-angiogenesis activity of GM3. Moreover, GM1 also stimulates MAPKinases signalling pathways, typically associated with an angiogenesis program. Caveolar-raft isolation and Western blotting of uPAR showed that GM1 promotes caveolar-raft partitioning of uPAR, as opposed to control and GM3-challenged EPCs. By confocal microscopy, we have shown that in EPCs uPAR is present on the surface in at least three compartments, respectively, associated to GM1, GM3 and caveolar rafts. Following GM1 exogenous addition, the GM3 compartment is depleted of uPAR which is recruited within caveolar rafts thereby triggering angiogenesis. PMID:25313007

  11. Differential uPAR recruitment in caveolar-lipid rafts by GM1 and GM3 gangliosides regulates endothelial progenitor cells angiogenesis

    PubMed Central

    Margheri, Francesca; Papucci, Laura; Schiavone, Nicola; D'Agostino, Riccardo; Trigari, Silvana; Serratì, Simona; Laurenzana, Anna; Biagioni, Alessio; Luciani, Cristina; Chillà, Anastasia; Andreucci, Elena; Del Rosso, Tommaso; Margheri, Giancarlo; Del Rosso, Mario; Fibbi, Gabriella

    2015-01-01

    Gangliosides and the urokinase plasminogen activator receptor (uPAR) tipically partition in specialized membrane microdomains called lipid-rafts. uPAR becomes functionally important in fostering angiogenesis in endothelial progenitor cells (EPCs) upon recruitment in caveolar-lipid rafts. Moreover, cell membrane enrichment with exogenous GM1 ganglioside is pro-angiogenic and opposite to the activity of GM3 ganglioside. On these basis, we first checked the interaction of uPAR with membrane models enriched with GM1 or GM3, relying on the adoption of solid-supported mobile bilayer lipid membranes with raft-like composition formed onto solid hydrophilic surfaces, and evaluated by surface plasmon resonance (SPR) the extent of uPAR recruitment. We estimated the apparent dissociation constants of uPAR-GM1/GM3 complexes. These preliminary observations, indicating that uPAR binds preferentially to GM1-enriched biomimetic membranes, were validated by identifying a pro-angiogenic activity of GM1-enriched EPCs, based on GM1-dependent uPAR recruitment in caveolar rafts. We have observed that addition of GM1 to EPCs culture medium promotes matrigel invasion and capillary morphogenesis, as opposed to the anti-angiogenesis activity of GM3. Moreover, GM1 also stimulates MAPKinases signalling pathways, typically associated with an angiogenesis program. Caveolar-raft isolation and Western blotting of uPAR showed that GM1 promotes caveolar-raft partitioning of uPAR, as opposed to control and GM3-challenged EPCs. By confocal microscopy, we have shown that in EPCs uPAR is present on the surface in at least three compartments, respectively, associated to GM1, GM3 and caveolar rafts. Following GM1 exogenous addition, the GM3 compartment is depleted of uPAR which is recruited within caveolar rafts thereby triggering angiogenesis. PMID:25313007

  12. May the remodeling of the Ca²⁺ toolkit in endothelial progenitor cells derived from cancer patients suggest alternative targets for anti-angiogenic treatment?

    PubMed

    Moccia, Francesco; Poletto, Valentina

    2015-09-01

    Endothelial progenitor cells (EPCs) may be recruited from bone marrow to sustain the metastatic switch in a number of solid cancers, including breast cancer (BC) and renal cellular carcinoma (RCC). Preventing EPC mobilization causes tumor shrinkage. Novel anti-angiogenic treatments have been introduced in therapy to inhibit VEGFR-2 signaling; unfortunately, these drugs blocked tumor angiogenesis in pre-clinical murine models, but resulted far less effective in human patients. Understanding the molecular mechanisms driving EPC proliferation and tubulogenesis in cancer patients could outline novel targets for alternative anti-angiogenic treatments. Store-operated Ca²⁺ entry (SOCE) regulates the growth of human EPCs, and it is mediated by the interaction between the endoplasmic reticulum Ca²⁺-sensor, Stim1, and the plasmalemmal Ca²⁺ channels, Orai1 and TRPC1. EPCs do not belong to the neoplastic clone: thus, unlike tumor endothelium and neoplastic cells, they should not remodel their Ca²⁺ toolkit in response to tumor microenvironment. However, our recent work demonstrated that EPCs isolated from naïve RCC patients (RCC-EPCs) undergo a dramatic remodeling of their Ca²⁺ toolkit by displaying a remarkable drop in the endoplasmic reticulum Ca²⁺ content, by down-regulating the expression of inositol-1,4,5-receptors (InsP3Rs), and by up-regulating Stim1, Orai1 and TRPC1. Moreover, EPCs are dramatically less sensitive to VEGF stimulation both in terms of Ca²⁺ signaling and of gene expression when isolated from tumor patients. Conversely, the pharmacological abolition of SOCE suppresses proliferation in these cells. These results question the suitability of VEGFR-2 as a therapeutically relevant target for anti-angiogenic treatments and hint at Orai1 and TRPC1 as more promising alternatives. This article is part of a Special Issue entitled: 13th European Symposium on Calcium. PMID:25447551

  13. Dynamics of bone marrow-derived endothelial progenitor cell/mesenchymal stem cell interaction in co-culture and its implications in angiogenesis

    SciTech Connect

    Aguirre, A.; Planell, J.A.; Engel, E.

    2010-09-17

    Research highlights: {yields} BM-EPCs and MSCs establish complex, self-organizing structures in co-culture. {yields} Co-culture decreases proliferation by cellular self-regulatory mechanisms. {yields} Co-cultured cells present an activated proangiogenic phenotype. {yields} qRT-PCR and cluster analysis identify new target genes playing important roles. -- Abstract: Tissue engineering aims to regenerate tissues and organs by using cell and biomaterial-based approaches. One of the current challenges in the field is to promote proper vascularization in the implant to prevent cell death and promote host integration. Bone marrow endothelial progenitor cells (BM-EPCs) and mesenchymal stem cells (MSCs) are bone marrow resident stem cells widely employed for proangiogenic applications. In vivo, they are likely to interact frequently both in the bone marrow and at sites of injury. In this study, the physical and biochemical interactions between BM-EPCs and MSCs in an in vitro co-culture system were investigated to further clarify their roles in vascularization. BM-EPC/MSC co-cultures established close cell-cell contacts soon after seeding and self-assembled to form elongated structures at 3 days. Besides direct contact, cells also exhibited vesicle transport phenomena. When co-cultured in Matrigel, tube formation was greatly enhanced even in serum-starved, growth factor free medium. Both MSCs and BM-EPCs contributed to these tubes. However, cell proliferation was greatly reduced in co-culture and morphological differences were observed. Gene expression and cluster analysis for wide panel of angiogenesis-related transcripts demonstrated up-regulation of angiogenic markers but down-regulation of many other cytokines. These data suggest that cross-talk occurs in between BM-EPCs and MSCs through paracrine and direct cell contact mechanisms leading to modulation of the angiogenic response.

  14. TNF-TNFR2/p75 Signaling Inhibits Early and Increases Delayed Nontargeted Effects in Bone Marrow-derived Endothelial Progenitor Cells*

    PubMed Central

    Sasi, Sharath P.; Song, Jin; Park, Daniel; Enderling, Heiko; McDonald, J. Tyson; Gee, Hannah; Garrity, Brittany; Shtifman, Alexander; Yan, Xinhua; Walsh, Kenneth; Natarajan, Mohan; Kishore, Raj; Goukassian, David A.

    2014-01-01

    TNF-α, a pro-inflammatory cytokine, is highly expressed after being irradiated (IR) and is implicated in mediating radiobiological bystander responses (RBRs). Little is known about specific TNF receptors in regulating TNF-induced RBR in bone marrow-derived endothelial progenitor cells (BM-EPCs). Full body γ-IR WT BM-EPCs showed a biphasic response: slow decay of p-H2AX foci during the initial 24 h and increase between 24 h and 7 days post-IR, indicating a significant RBR in BM-EPCs in vivo. Individual TNF receptor (TNFR) signaling in RBR was evaluated in BM-EPCs from WT, TNFR1/p55KO, and TNFR2/p75KO mice, in vitro. Compared with WT, early RBR (1–5 h) were inhibited in p55KO and p75KO EPCs, whereas delayed RBR (3–5 days) were amplified in p55KO EPCs, suggesting a possible role for TNFR2/p75 signaling in delayed RBR. Neutralizing TNF in γ-IR conditioned media (CM) of WT and p55KO BM-EPCs largely abolished RBR in both cell types. ELISA protein profiling of WT and p55KO EPC γ-IR-CM over 5 days showed significant increases in several pro-inflammatory cytokines, including TNF-α, IL-1α (Interleukin-1 alpha), RANTES (regulated on activation, normal T cell expressed and secreted), and MCP-1. In vitro treatments with murine recombinant (rm) TNF-α and rmIL-1α, but not rmMCP-1 or rmRANTES, increased the formation of p-H2AX foci in nonirradiated p55KO EPCs. We conclude that TNF-TNFR2 signaling may induce RBR in naïve BM-EPCs and that blocking TNF-TNFR2 signaling may prevent delayed RBR in BM-EPCs, conceivably, in bone marrow milieu in general. PMID:24711449

  15. Spliced stromal cell-derived factor-1α analog stimulates endothelial progenitor cell migration and improves cardiac function in a dose-dependent manner after myocardial infarction

    PubMed Central

    Hiesinger, William; Frederick, John R.; Atluri, Pavan; McCormick, Ryan C.; Marotta, Nicole; Muenzer, Jeffrey R.; Woo, Y. Joseph

    2011-01-01

    Objectives Stromal cell-derived factor (SDF)-1α is a potent endogenous endothelial progenitor cell (EPC) chemokine and key angiogenic precursor. Recombinant SDF-1α has been demonstrated to improve neovasculogenesis and cardiac function after myocardial infarction (MI) but SDF-1α is a bulky protein with a short half-life. Small peptide analogs might provide translational advantages, including ease of synthesis, low manufacturing costs, and the potential to control delivery within tissues using engineered biomaterials. We hypothesized that a minimized peptide analog of SDF-1α, designed by splicing the N-terminus (activation and binding) and C-terminus (extracellular stabilization) with a truncated amino acid linker, would induce EPC migration and preserve ventricular function after MI. Methods EPC migration was first determined in vitro using a Boyden chamber assay. For in vivo analysis, male rats (n=48) underwent left anterior descending coronary artery ligation. At infarction, the rats were randomized into 4 groups and received peri-infarct intramyocardial injections of saline, 3 μg/kg of SDF-1α, 3 μg/kg of spliced SDF analog, or 6 μg/kg spliced SDF analog. After 4 weeks, the rats underwent closed chest pressure volume conductance catheter analysis. Results EPCs showed significantly increased migration when placed in both a recombinant SDF-1α and spliced SDF analog gradient. The rats treated with spliced SDF analog at MI demonstrated a significant dose-dependent improvement in end-diastolic pressure, stroke volume, ejection fraction, cardiac output, and stroke work compared with the control rats. Conclusions A spliced peptide analog of SDF-1α containing both the N- and C- termini of the native protein induced EPC migration, improved ventricular function after acute MI, and provided translational advantages compared with recombinant human SDF-1α. PMID:20951261

  16. Stromal Cell-Derived Factor-1α Activation of Tissue Engineered Endothelial Progenitor Cell Matrix Enhances Ventricular Function after Myocardial Infarction by Inducing Neovasculogenesis

    PubMed Central

    Frederick, John R.; Fitzpatrick, J. Raymond; McCormick, Ryan C.; Harris, David A.; Kim, Ah-Young; Muenzer, Jeffrey R.; Marotta, Nicole; Smith, Maximilian J.; Cohen, Jeffrey E.; Hiesinger, William; Atluri, Pavan; Woo, Y. Joseph

    2014-01-01

    Background Myocardial ischemia causes cardiomyocyte death, adverse ventricular remodeling, and ventricular dysfunction. Endothelial progenitor cells (EPC) have been shown to ameliorate this process, particularly when activated with stromal cell-derived factor-1α (SDF). We hypothesized that implantation of a tissue engineered extracellular matrix scaffold seeded with EPCs primed with SDF could induce neovasculogenesis, prevent adverse remodeling, and preserve ventricular function after myocardial infarction (MI). Methods and Results Lewis rats (n=82) underwent left anterior descending artery ligation to induce MI. EPCs were cultured on a vitronectin/collagen scaffold, and primed with SDF to generate the activated EPC matrix (EPCM). EPCM was sutured to the anterolateral left ventricular (LV) wall including the region of ischemia.. At four weeks, when compared to controls, borderzone myocardial tissue demonstrated increased levels of VEGF in the EPCM group. Vessel density as assessed by immunohistochemical microscopy was significantly increased in the EPCM group (4.1 vs 6.2 vessels/high-powered field, p<0.001), and microvascular perfusion measured by lectin microangiography was enhanced four-fold (0.7 vs. 2.7% vessel volume/section volume, p=0.04). Ventricular geometry and scar fraction assessed by analysis of sectioned hearts exhibited significantly preserved LV internal diameter (9.7mm vs. 8.6mm, p=0.005) and decreased infarct scar expressed as percent of total section area (16% vs. 7%, p=0.002) when compared to all other groups. In addition, EPCM animals showed a significant preservation of function as measured by echocardiography, pressure volume-conductance, and Doppler flow. Conclusions Extracellular matrix seeded with EPCs primed with SDF induces borderzone neovasculogenesis, attenuates adverse ventricular remodeling, and preserves ventricular function after MI. PMID:20837901

  17. SIRT1 Protects Against Oxidative Stress-Induced Endothelial Progenitor Cells Apoptosis by Inhibiting FOXO3a via FOXO3a Ubiquitination and Degradation.

    PubMed

    Wang, Yu-Qiang; Cao, Qing; Wang, Fei; Huang, Li-Ya; Sang, Tian-Tian; Liu, Fang; Chen, Shu-Yan

    2015-09-01

    Cell loss due to apoptosis induced by oxidative stress is a major hurdle for endothelial progenitor cells (EPCs)-based therapy. Sirtuin 1 (SIRT1) plays important roles in many pathophysiological processes by deacetylating various substrates, including forkhead transcription factor (FOXO). However, after deacetylation, the fate of FOXO protein remains to be explored. In the present study, we investigated whether SIRT1 exerted a protective effect on hydrogen peroxide (H(2)O(2))-induced EPCs apoptosis and, if so, what the underlying mechanism might be. EPCs were isolated and obtained from human umbilical cord blood by density gradient centrifugation and identified by morphology, tube formation ability, cell surface markers, and the ability to take up acetylated low-density lipoprotein (Dil-Ac-LDL) and bind ulex europaeus agglutinin 1 (FITC-UEA-1). Immunofluorescence showed that SIRT1 is localized in the nucleus of EPCs in the presence or absence of H(2)O(2). SIRT1 protein level in EPCs was increased by the treatment with H(2)O(2) for 24 h. Incubation of EPCs with H(2)O(2) dose dependently induced EPCs apoptosis. SIRT1 overexpression reduced the rate of EPCs apoptosis induced by H(2)O(2), whereas SIRT1 downregulation and EX527, a specific SIRT1 inhibitor, exerted the opposite effect. SIRT1 overexpression decreased the total FOXO3a protein expression, whereas SIRT1 downregulation and EX527 increased the amount of FOXO3a protein. SIRT1 reduced FOXO3a transcriptional activity according to Bim expression. Co-immunoprecipitation assay showed that SIRT1 could bind to FOXO3a, reduce its acetylation level and increase its ubiquitination level. To sum up, our work demonstrated that SIRT1 had a pivotally protective role in the regulation of EPCs apoptosis induced by H(2)O(2) and that SIRT1 protected against apoptosis by inhibiting FOXO3a via FOXO3a ubiquitination and subsequent degradation. PMID:25640014

  18. Melittin inhibits tumor angiogenesis modulated by endothelial progenitor cells associated with the SDF-1α/CXCR4 signaling pathway in a UMR-106 osteosarcoma xenograft mouse model

    PubMed Central

    QIN, GANG; CHEN, YONGQIANG; LI, HAIDONG; XU, SUYANG; LI, YUMEI; SUN, JIAN; RAO, WU; CHEN, CHAOWEI; DU, MINDONG; HE, KAIYI; YE, YONG

    2016-01-01

    Endothelial progenitor cells (EPCs) are important in tumor angiogenesis. Stromal cell-derived factor-1α (SDF-1α) and its receptor C-X-C chemokine receptor type 4 (CXCR4) are key in stem cell homing. Melittin, a component of bee venom, exerts antitumor activity, however, the underlying mechanisms remain to be elucidated. The present study aimed to assess the effects of melittin on EPCs and angiogenesis in a mouse model of osteosarcoma. UMR-106 cells and EPCs were treated with various concentrations of melittin and cell viability was determined using the MTT assay. EPC adherence, migration and tube forming ability were assessed. Furthermore, SDF-1α, AKT and extracellular signal-regulated kinase (ERK)1/2 expression levels were detected by western blotting. Nude mice were inoculated with UMR-106 cells to establish an osteosarcoma mouse model. The tumors were injected with melittin, and its effects were assessed by immunohistochemistry and immunofluorescence. Melittin decreased the viability of UMR-106 cells and EPCs. In addition, it decreased EPC adhesion, migration and tube formation when compared with control and SDF-1α-treated cells. Melittin decreased the expression of phosphorylated (p)-AKT, p-ERK1/2, SDF-1α and CXCR4 in UMR-106 cells and EPCs when compared with the control. The proportions of cluster of differentiation (CD)34/CD133 double-positive cells were 16.4±10.4% in the control, and 7.0±4.4, 2.9±1.2 and 1.3±0.3% in tumors treated with 160, 320 and 640 µg/kg melittin per day, respectively (P<0.05). At 11 days, melittin reduced the tumor size when compared with that of the control (control, 4.8±1.3 cm3; melittin, 3.2±0.6, 2.6±0.5, and 2.0±0.2 cm3 for 160, 320 and 640 µg/kg, respectively; all P<0.05). Melittin decreased the microvessel density, and SDF-1α and CXCR4 protein expression levels in the tumors. Melittin may decrease the effect of osteosarcoma on EPC-mediated angiogenesis, possibly via inhibition of the SDF-1α/CXCR4 signaling pathway

  19. Store-Operated Ca2+ Entry Is Remodelled and Controls In Vitro Angiogenesis in Endothelial Progenitor Cells Isolated from Tumoral Patients

    PubMed Central

    Dragoni, Silvia; Bottino, Cinzia; Ong, Hwei Ling; Guerra, Germano; Ganini, Carlo; Massa, Margherita; Manzoni, Mariangela; Ambudkar, Indu S.; Genazzani, Armando A.; Rosti, Vittorio; Pedrazzoli, Paolo; Tanzi, Franco; Moccia, Francesco

    2012-01-01

    Background Endothelial progenitor cells (EPCs) may be recruited from bone marrow to sustain tumor vascularisation and promote the metastatic switch. Understanding the molecular mechanisms driving EPC proliferation and tubulogenesis could outline novel targets for alternative anti-angiogenic treatments. Store-operated Ca2+ entry (SOCE), which is activated by a depletion of the intracellular Ca2+ pool, regulates the growth of human EPCs, where is mediated by the interaction between the endoplasmic reticulum Ca2+-sensor, Stim1, and the plasmalemmal Ca2+ channel, Orai1. As oncogenesis may be associated to the capability of tumor cells to grow independently on Ca2+ influx, it is important to assess whether SOCE regulates EPC-dependent angiogenesis also in tumor patients. Methodology/Principal Findings The present study employed Ca2+ imaging, recombinant sub-membranal and mitochondrial aequorin, real-time polymerase chain reaction, gene silencing techniques and western blot analysis to investigate the expression and the role of SOCE in EPCs isolated from peripheral blood of patients affected by renal cellular carcinoma (RCC; RCC-EPCs) as compared to control EPCs (N-EPCs). SOCE, activated by either pharmacological (i.e. cyclopiazonic acid) or physiological (i.e. ATP) stimulation, was significantly higher in RCC-EPCs and was selectively sensitive to BTP-2, and to the trivalent cations, La3+ and Gd3+. Furthermore, 2-APB enhanced thapsigargin-evoked SOCE at low concentrations, whereas higher doses caused SOCE inhibition. Conversely, the anti-angiogenic drug, carboxyamidotriazole (CAI), blocked both SOCE and the intracellular Ca2+ release. SOCE was associated to the over-expression of Orai1, Stim1, and transient receptor potential channel 1 (TRPC1) at both mRNA and protein level The intracellular Ca2+ buffer, BAPTA, BTP-2, and CAI inhibited RCC-EPC proliferation and tubulogenesis. The genetic suppression of Stim1, Orai1, and TRPC1 blocked CPA-evoked SOCE in RCC

  20. Melittin inhibits tumor angiogenesis modulated by endothelial progenitor cells associated with the SDF-1α/CXCR4 signaling pathway in a UMR-106 osteosarcoma xenograft mouse model.

    PubMed

    Qin, Gang; Chen, Yongqiang; Li, Haidong; Xu, Suyang; Li, Yumei; Sun, Jian; Rao, Wu; Chen, Chaowei; Du, Mindong; He, Kaiyi; Ye, Yong

    2016-07-01

    Endothelial progenitor cells (EPCs) are important in tumor angiogenesis. Stromal cell-derived factor-1α (SDF-1α) and its receptor C-X-C chemokine receptor type 4 (CXCR4) are key in stem cell homing. Melittin, a component of bee venom, exerts antitumor activity, however, the underlying mechanisms remain to be elucidated. The present study aimed to assess the effects of melittin on EPCs and angiogenesis in a mouse model of osteosarcoma. UMR‑106 cells and EPCs were treated with various concentrations of melittin and cell viability was determined using the MTT assay. EPC adherence, migration and tube forming ability were assessed. Furthermore, SDF‑1α, AKT and extracellular signal‑regulated kinase (ERK)1/2 expression levels were detected by western blotting. Nude mice were inoculated with UMR‑106 cells to establish an osteosarcoma mouse model. The tumors were injected with melittin, and its effects were assessed by immunohistochemistry and immunofluorescence. Melittin decreased the viability of UMR‑106 cells and EPCs. In addition, it decreased EPC adhesion, migration and tube formation when compared with control and SDF‑1α‑treated cells. Melittin decreased the expression of phosphorylated (p)‑AKT, p‑ERK1/2, SDF‑1α and CXCR4 in UMR‑106 cells and EPCs when compared with the control. The proportions of cluster of differentiation (CD)34/CD133 double‑positive cells were 16.4±10.4% in the control, and 7.0±4.4, 2.9±1.2 and 1.3±0.3% in tumors treated with 160, 320 and 640 µg/kg melittin per day, respectively (P<0.05). At 11 days, melittin reduced the tumor size when compared with that of the control (control, 4.8±1.3 cm3; melittin, 3.2±0.6, 2.6±0.5, and 2.0±0.2 cm3 for 160, 320 and 640 µg/kg, respectively; all P<0.05). Melittin decreased the microvessel density, and SDF‑1α and CXCR4 protein expression levels in the tumors. Melittin may decrease the effect of osteosarcoma on EPC‑mediated angiogenesis, possibly via inhibition

  1. Diazoxide preconditioning of endothelial progenitor cells from streptozotocin-induced type 1 diabetic rats improves their ability to repair diabetic cardiomyopathy.

    PubMed

    Ali, Muhammad; Mehmood, Azra; Anjum, Muhammad Sohail; Tarrar, Moazzam Nazir; Khan, Shaheen N; Riazuddin, Sheikh

    2015-12-01

    Type 1 diabetes mellitus (DM) is a strong risk factor for the development of diabetic cardiomyopathy (DCM) which is the leading cause of morbidity and mortality in the type 1 diabetic patients. Stem cells may act as a therapeutic agent for the repair of DCM. However, deteriorated functional abilities and survival of stem cells derived from type 1 diabetic subjects need to be overcome for obtaining potential outcome of the stem cell therapy. Diazoxide (DZ) a highly selective mitochondrial ATP-sensitive K(+) channel opener has been previously shown to improve the ability of mesenchymal stem cells for the repair of heart failure. In the present study, we evaluated the effects of DZ preconditioning in improving the ability of streptozotocin-induced type 1 diabetes affected bone marrow-derived endothelial progenitor cells (DM-EPCs) for the repair of DCM in the type 1 diabetic rats. DM-EPCs were characterized by immunocytochemistry, flow cytometry, and reverse transcriptase PCR for endothelial cell-specific markers like vWF, VE cadherin, VEGFR2, PECAM, CD34, and eNOS. In vitro studies included preconditioning of DM-EPCs with 200 μM DZ for 30 min followed by exposure to either 200 μM H2O2 for 2 h (for oxidative stress induction) or 30 mM glucose media (for induction of hyperglycemic stress) for 48 h. Non-preconditioned EPCs with and without exposure to H2O2 and 30 mM high glucose served as controls. These cells were then evaluated for survival (by MTT and XTT cell viability assays), senescence, paracrine potential (by ELISA for VEGF), and alteration in gene expression [VEGF, stromal derived factor-1α (SDF-1α), HGF, bFGF, Bcl2, and Caspase-3]. DZ preconditioned DM-EPCs demonstrated significantly increased survival and VEGF release while reduced cell injury and senescence. Furthermore, DZ preconditioned DM-EPCs exhibited up-regulated expression of prosurvival genes (VEGF, SDF-1α, HGF, bFGF, and Bcl2) on exposure to H2O2, and VEGF and Bcl2 on exposure to hyperglycemia

  2. Levels and values of lipoprotein-associated phospholipase A2, galectin-3, RhoA/ROCK, and endothelial progenitor cells in critical limb ischemia: pharmaco-therapeutic role of cilostazol and clopidogrel combination therapy

    PubMed Central

    2014-01-01

    Objective We tested the hypothesis that clopidogrel and cilostazol combination therapy could effectively attenuate systemic inflammatory reaction, facilitate proliferation of circulating endothelial progenitor cell (EPC), and improve the clinical outcomes of critical limb ischemia (CLI) in patients unsuitable for surgical revascularization or percutaneous transluminal angioplasty (PTA). Methods A total 55 patients (mean age, 72 years; 56% female) were consecutively enrolled. Clopidogrel and cilostazol combination therapy was administered throughout the study period. Results As compared with the baseline, circulating endothelial progenitor cell level (as shown by flow cytometry) was significantly increased (p < 0.003), whereas the CLI-related ulcers and painfulness were significantly improved (all p < 0.01) by day 90 after treatment. On the other hand, after clopidogrel and cilostazol combination therapy, galectin-3 level, lipoprotein-associated phospholipase A2 gene expression, and RhoA/ROCK-related protein expression in peripheral blood mononuclear cells were significantly suppressed (all p < 0.01). Eventually, by day 90, 5 patients (9.1%) died of other etiologies, 3 (5.5%) withdrew from the study, 6 (10.9%) required amputation, and the remaining 41 had satisfactory clinical improvement with complete wound healing in 9 (16.4%) patients. Conclusion The results of the present study highlight that clopidogrel and cilostazol combination therapy may be considered to be an alternative method for treating patients with CLI unsuitable for surgical revascularization or PTA. PMID:24742198

  3. An Anti-CD34 Antibody-Functionalized Clinical-Grade POSS-PCU Nanocomposite Polymer for Cardiovascular Stent Coating Applications: A Preliminary Assessment of Endothelial Progenitor Cell Capture and Hemocompatibility

    PubMed Central

    Farhatnia, Yasmin; G, Natasha; Lim, Jing; Teoh, Swee-Hin; Rajadas, Jayakumar; Alavijeh, Mohammad S.; Seifalian, Alexander M.

    2013-01-01

    In situ endothelialization of cardiovascular implants has emerged in recent years as an attractive means of targeting the persistent problems of thrombosis and intimal hyperplasia. This study aimed to investigate the efficacy of immobilizing anti-CD34 antibodies onto a POSS-PCU nanocomposite polymer surface to sequester endothelial progenitor cells (EPCs) from human blood, and to characterize the surface properties and hemocompatibility of this surface. Amine-functionalized fumed silica was used to covalently conjugate anti-CD34 to the polymer surface. Water contact angle, fluorescence microscopy, and scanning electron microscopy were used for surface characterization. Peripheral blood mononuclear cells (PBMCs) were seeded on modified and pristine POSS-PCU polymer films. After 7 days, adhered cells were immunostained for the expression of EPC and endothelial cell markers, and assessed for the formation of EPC colonies. Hemocompatibility was assessed by thromboelastography, and platelet activation and adhesion assays. The number of EPC colonies formed on anti-CD34-coated POSS-PCU surfaces was not significantly higher than that of POSS-PCU (5.0±1.0 vs. 1.7±0.6, p>0.05). However, antibody conjugation significantly improved hemocompatibility, as seen from the prolonged reaction and clotting times, decreased angle and maximum amplitude (p<0.05), as well as decreased platelet adhesion (76.8±7.8 vs. 8.4±0.7, p<0.05) and activation. Here, we demonstrate that POSS-PCU surface immobilized anti-CD34 antibodies selectively captured CD34+ cells from peripheral blood, although only a minority of these were EPCs. Nevertheless, antibody conjugation significantly improves the hemocompatibility of POSS-PCU, and should therefore continue to be explored in combination with other strategies to improve the specificity of EPC capture to promote in situ endothelialization. PMID:24116210

  4. Mechanisms underlying protective effects of trimetazidine on endothelial progenitor cells biological functions against H2O2-induced injury: involvement of antioxidation and Akt/eNOS signaling pathways.

    PubMed

    Wu, Qinqin; Qi, Benling; Liu, Yun; Cheng, Bei; Liu, Lihua; Li, Yuanyuan; Wang, Qian

    2013-05-01

    Trimetazidine (TMZ) is a widely used drug exerting cardioprotective effects against ischemic heart disease through a number of mechanisms in conditions of oxidative stress. However, there are few data regarding the effects of TMZ on endothelial lineage, especially endothelial progenitor cells (EPCs). Thus, we sought to investigate whether TMZ could protect EPCs against oxidative stress injury induced by H2O2 (100 µM) and the preliminary mechanisms involved in vitro. The results showed that pretreatment of EPCs with TMZ (10 µM) protected the proliferation, adhesion, migration, and apoptosis of EPCs against H2O2, accompanied by an increase in superoxide dismutase (SOD) activity, a decrease in malonaldehyde (MDA) content, and increases in eNOS, Akt phosphorylation, and NO production. These TMZ-mediated beneficial effects on EPCs could be attenuated by pre-incubation with the Akt inhibitor triciribine. In conclusion, the present study demonstrates that TMZ ameliorated H2O2-induced impairment of biological functions in EPCs with the involvement of antioxidation and Akt/eNOS signaling pathway. These findings suggest that TMZ mediating preservation of EPCs may contribute to its cardioprotective effects on ischemic heart disease. PMID:23528356

  5. How to utilize Ca2+ signals to rejuvenate the repairative phenotype of senescent endothelial progenitor cells in elderly patients affected by cardiovascular diseases: a useful therapeutic support of surgical approach?

    PubMed Central

    2013-01-01

    Endothelial dysfunction or loss is the early event that leads to a host of severe cardiovascular diseases, such as atherosclerosis, hypertension, brain stroke, myocardial infarction, and peripheral artery disease. Ageing is regarded among the most detrimental risk factor for vascular endothelium and predisposes the subject to atheroscleorosis and inflammatory states even in absence of traditional comorbid conditions. Standard treatment to restore blood perfusion through stenotic arteries are surgical or endovascular revascularization. Unfortunately, ageing patients are not the most amenable candidates for such interventions, due to high operative risk or unfavourable vascular involvement. It has recently been suggested that the transplantation of autologous bone marrow-derived endothelial progenitor cells (EPCs) might constitute an alternative and viable therapeutic option for these individuals. Albeit pre-clinical studies demonstrated the feasibility of EPC-based therapy to recapitulate the diseased vasculature of young and healthy animals, clinical studies provided less impressive results in old ischemic human patients. One hurdle associated to this kind of approach is the senescence of autologous EPCs, which are less abundant in peripheral blood and display a reduced pro-angiogenic activity. Conversely, umbilical cord blood (UCB)-derived EPCs are more suitable for cellular therapeutics due to their higher frequency and sensitivity to growth factors, such as vascular endothelial growth factor (VEGF). An increase in intracellular Ca2+ concentration is central to EPC activation by VEGF. We have recently demonstrated that the Ca2+ signalling machinery driving the oscillatory Ca2+ response to this important growth factor is different in UCB-derived EPCs as compared to their peripheral counterparts. In particular, we focussed on the so-called endothelial colony forming cells (ECFCs), which are the only EPC population belonging to the endothelial lineage and able to

  6. Inactivation of viruses in labile blood derivatives. II. Physical methods

    SciTech Connect

    Horowitz, B.; Wiebe, M.E.; Lippin, A.; Vandersande, J.; Stryker, M.H.

    1985-11-01

    The thermal inactivation of viruses in labile blood derivatives was evaluated by addition of marker viruses (VSV, Sindbis, Sendai, EMC) to anti-hemophilic factor (AHF) concentrates. The rate of virus inactivation at 60 degrees C was decreased by at least 100- to 700-fold by inclusion of 2.75 M glycine and 50 percent sucrose, or 3.0 M potassium citrate, additives which contribute to retention of protein biologic activity. Nonetheless, at least 10(4) infectious units of each virus was inactivated within 10 hours. Increasing the temperature from 60 to 70 or 80 degrees C caused a 90 percent or greater loss in AHF activity. An even greater decline in the rate of virus inactivation was observed on heating AHF in the lyophilized state, although no loss in AHF activity was observed after 72 hours of heating at 60 degrees C. Several of the proteins present in lyophilized AHF concentrates displayed an altered electrophoretic mobility as a result of exposure to 60 degrees C for 24 hours. Exposure of lyophilized AHF to irradiation from a cobalt 60 source resulted in an acceptable yield of AHF at 1.0, but not at 2.0, megarads. At 1 megarad, greater than or equal to 6.0 logs of VSV and 3.3 logs of Sindbis virus were inactivated.

  7. Isolation of Functional Human Endothelial Cells from Small Volumes of Umbilical Cord Blood

    PubMed Central

    Do Kang, Sa; Carlon, Tim A.; Jantzen, Alexandra E.; Lin, Fu-Hsiung; Ley, Melissa M.; Allen, Jason D.; Stabler, Thomas V.; Haley, N. Rebecca; Truskey, George A.; Achneck, Hardean E.

    2013-01-01

    Endothelial cells (ECs) isolated from endothelial progenitor cells in blood have great potential as a therapeutic tool to promote vasculogenesis and angiogenesis and treat cardiovascular diseases. However, current methods to isolate ECs are limited by a low yield with few colonies appearing during isolation. In order to utilize blood-derived ECs for therapeutic applications, a simple method is needed that can produce a high yield of ECs from small volumes of blood without the addition of animal-derived products. For the first time, we show that human endothelial cells can be isolated without the prior separation of blood components through the technique of diluted whole blood incubation (DWBI) utilizing commercially available human serum. We isolated ECs from small volumes of blood (~ 10 ml) via DWBI and characterized them with flow cytometry, immunohistochemistry, and uptake of DiI-labeled acetylated low density lipoprotein (DiI-Ac-LDL). These ECs are functional as demonstrated by their ability to form tubular networks in Matrigel, adhere and align with flow under physiological fluid shear stress, and produce increased nitric oxide under fluid flow. An average of 7.0 ± 2.5 EC colonies that passed all functional tests described above were obtained per 10 ml of blood as compared to only 0.3 ± 0.1 colonies with the traditional method based on density centrifugation. The time until first colony appearance was 8.3 ± 1.2 days for ECs isolated with the DWBI method and 12 ± 1.4 days for ECs isolated with the traditional isolation method. A simplified method, such as DWBI, in combination with advances in isolation yield could enable the use of blood-derived ECs in clinical practice. PMID:23604849

  8. A Chemokine Receptor, CXCR4, Which Is Regulated by Hypoxia-Inducible Factor 2α, Is Crucial for Functional Endothelial Progenitor Cells Migration to Ischemic Tissue and Wound Repair

    PubMed Central

    Tu, Tran Cam; Nagano, Masumi; Yamashita, Toshiharu; Hamada, Hiromi; Ohneda, Kinuko; Kimura, Kenichi

    2016-01-01

    Endothelial progenitor cells (EPCs) have the ability to form new blood vessels and protect ischemic tissues from damage. We previously reported that EPCs with low activity of aldehyde dehydrogenase (Alde-Low EPCs) possess the greater ability to treat ischemic tissues compared with Alde-High EPCs. The expression level of the hypoxia-inducible factors (HIFs), HIF-1α and HIF-2α, was found to be greater in Alde-Low EPCs than in Alde-High EPCs. However, the precise role of the HIF factors in the regulation of EPC activity remains obscure. In this study, we demonstrate a critical role of HIF-2α and its target gene CXCR4 for controlling the migratory activity of EPC to ischemic tissue. We found that coculture of Alde-High EPCs with microvesicles derived from Alde-Low EPCs improved their ability to repair an ischemic skin flap, and the expression of CXCR4 and its ligand SDF1 was significantly increased following the coculture. In Alde-Low EPCs, the expression of CXCR4 was suppressed by short hairpin RNA (shRNA)-mediated HIF-2α, but not HIF-1α downregulation. Chromatin immunoprecipitation assays showed that HIF-2α, but not HIF-1α, binds to the promoter region of CXCR4 gene. The CXCR4 shRNA treatment in Alde-Low EPCs almost completely abrogated their migratory activity to ischemic tissues, whereas the reduction of vascular endothelial growth factor (VEGF) showed much less effect. The CXCR4 overexpression in Alde-High EPCs resulted in a partial, but significant improvement in their repairing ability in an ischemic skin flap. Collectively, these findings indicate that the CXCR4/SDF-1 axis, which is specifically regulated by HIF-2α, plays a crucial role in the regulation of EPC migration to ischemic tissues. PMID:26620723

  9. A Chemokine Receptor, CXCR4, Which Is Regulated by Hypoxia-Inducible Factor 2α, Is Crucial for Functional Endothelial Progenitor Cells Migration to Ischemic Tissue and Wound Repair.

    PubMed

    Tu, Tran Cam; Nagano, Masumi; Yamashita, Toshiharu; Hamada, Hiromi; Ohneda, Kinuko; Kimura, Kenichi; Ohneda, Osamu

    2016-02-01

    Endothelial progenitor cells (EPCs) have the ability to form new blood vessels and protect ischemic tissues from damage. We previously reported that EPCs with low activity of aldehyde dehydrogenase (Alde-Low EPCs) possess the greater ability to treat ischemic tissues compared with Alde-High EPCs. The expression level of the hypoxia-inducible factors (HIFs), HIF-1α and HIF-2α, was found to be greater in Alde-Low EPCs than in Alde-High EPCs. However, the precise role of the HIF factors in the regulation of EPC activity remains obscure. In this study, we demonstrate a critical role of HIF-2α and its target gene CXCR4 for controlling the migratory activity of EPC to ischemic tissue. We found that coculture of Alde-High EPCs with microvesicles derived from Alde-Low EPCs improved their ability to repair an ischemic skin flap, and the expression of CXCR4 and its ligand SDF1 was significantly increased following the coculture. In Alde-Low EPCs, the expression of CXCR4 was suppressed by short hairpin RNA (shRNA)-mediated HIF-2α, but not HIF-1α downregulation. Chromatin immunoprecipitation assays showed that HIF-2α, but not HIF-1α, binds to the promoter region of CXCR4 gene. The CXCR4 shRNA treatment in Alde-Low EPCs almost completely abrogated their migratory activity to ischemic tissues, whereas the reduction of vascular endothelial growth factor (VEGF) showed much less effect. The CXCR4 overexpression in Alde-High EPCs resulted in a partial, but significant improvement in their repairing ability in an ischemic skin flap. Collectively, these findings indicate that the CXCR4/SDF-1 axis, which is specifically regulated by HIF-2α, plays a crucial role in the regulation of EPC migration to ischemic tissues. PMID:26620723

  10. Progenitors for the corneal endothelium and trabecular meshwork: a potential source for personalized stem cell therapy in corneal endothelial diseases and glaucoma.

    PubMed

    Yu, Wing Yan; Sheridan, Carl; Grierson, Ian; Mason, Sharon; Kearns, Victoria; Lo, Amy Cheuk Yin; Wong, David

    2011-01-01

    Several adult stem cell types have been found in different parts of the eye, including the corneal epithelium, conjunctiva, and retina. In addition to these, there have been accumulating evidence that some stem-like cells reside in the transition area between the peripheral corneal endothelium (CE) and the anterior nonfiltering portion of the trabecular meshwork (TM), which is known as the Schwalbe's Ring region. These stem/progenitor cells may supply new cells for the CE and TM. In fact, the CE and TM share certain similarities in terms of their embryonic origin and proliferative capacity in vivo. In this paper, we discuss the putative stem cell source which has the potential for replacement of lost and nonfunctional cells in CE diseases and glaucoma. The future development of personalized stem cell therapies for the CE and TM may reduce the requirement of corneal grafts and surgical treatments in glaucoma. PMID:22187525

  11. Differentiation state determines neural effects on microvascular endothelial cells

    SciTech Connect

    Muffley, Lara A.; Pan, Shin-Chen; Smith, Andria N.; Ga, Maricar; Hocking, Anne M.; Gibran, Nicole S.

    2012-10-01

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

  12. Correlation of ionizing irradiation-induced late pulmonary fibrosis with long-term bone marrow culture fibroblast progenitor cell biology in mice homozygous deletion recombinant negative for endothelial cell adhesion molecules.

    PubMed

    Epperly, Michael W; Guo, Hongliang; Shields, Donna; Zhang, Xichen; Greenberger, Joel S

    2004-01-01

    Ionizing irradiation damage to the lung is associated with an acute inflammatory reaction, followed by a latent period and then late effects including predominantly pulmonary fibrosis. The cells mediating fibrosis have recently been shown to derive from the bone marrow hematopoietic microenvironment. Initiation of late pulmonary irradiation lung damage has been correlated with up-regulation of VCAM-1 and ICAM-1 in pulmonary endothelial cells, followed by infiltration of macrophages and bone marrow-derived fibroblasts forming the fibrotic lesions of organizing alveolitis/fibrosis. To determine whether the absence of expression of VCAM-1, ICAM-1, or other adhesion molecules known to be relevant to inflammatory cell attachment to lung endothelial cells was associated with a decrease in irradiation-induced lung fibrosis, homozygous deletion recombinant knockout mice lacking each of several adhesion molecules were tested compared to littermates for survival and development of organizing alveolitis following 20 Gy irradiation to both lungs. Bone marrow culture longevity has been shown to be a parameter, which correlates with both hematopoietic stem cell reserve and the integrity of fibroblast progenitors of the supportive hematopoietic microenvironment; radiation lung survival data were correlated to longevity of hematopoiesis in long-term bone marrow cultures established from tibia and femur bone marrow of the same mice. Homozygous deletion recombinant negative mice including VCAM-1-/-, ICAM-1-/-, E-Selectin-/-, or L-Selectin-/- were irradiated to 20 Gy to both lungs and followed for survival and percent organizing alveolitis at time of death compared to each normal littermate. A significant increase in survival (median 190 days) was detected with L-Selectin-/- compared to littermate control mice (median 140 days) or other groups. Long-term bone marrow cultures from L-Selectin-/- mice showed no detectable difference in marrow fibroblasts or hematopoietic cell biology

  13. Involvement of marrow-derived endothelial cells in vascularization.

    PubMed

    Larrivée, B; Karsan, A

    2007-01-01

    Until recently, the adult neovasculature was thought to arise only through angiogenesis, the mechanism by which new blood vessels form from preexisting vessels through endothelial cell migration and proliferation. However, recent studies have provided evidence that postnatal neovasculature can also arise though vasculogenesis, a process by which endothelial progenitor cells are recruited and differentiate into mature endothelial cells to form new blood vessels. Evidence for the existence of endothelial progenitors has come from studies demonstrating the ability of bone marrow-derived cells to incorporate into adult vasculature. However, the exact nature of endothelial progenitor cells remains controversial. Because of the lack of definitive markers of endothelial progenitors, the in vivo contribution of progenitor cells to physiological and pathological neovascularization remains unclear. Early studies reported that endothelial progenitor cells actively integrate into the adult vasculature and are critical in the development of many types of vascular-dependent disorders such as neoplastic progression. Moreover, it has been suggested that endothelial progenitor cells can be used as a therapeutic strategy aimed at promoting vascular growth in a variety of ischemic diseases. However, increasing numbers of studies have reported no clear contribution of endothelial progenitors in physiological or pathological angiogenesis. In this chapter, we discuss the origin of the endothelial progenitor cell in the embryo and adult, and we discuss the cell's link to the primitive hematopoietic stem cell. We also review the potential significance of endothelial progenitor cells in the formation of a postnatal vascular network and discuss the factors that may account for the current lack of consensus of the scientific community on this important issue. PMID:17554506

  14. Controlled release of stromal cell-derived factor-1α from silk fibroin-coated coils accelerates intra-aneurysmal organization and occlusion of neck remnant by recruiting endothelial progenitor cells.

    PubMed

    Gao, Yuyuan; Wang, Qiujing; Cui, Xubo; Liu, Yaqi; Zheng, Tao; Chen, Chengwei; Sun, Chengmei; Huang, Shuyun; Wang, Xin; Liu, Yanchao; Jiang, Xiaodan; Zeng, Chi; Quan, Daping

    2014-01-01

    This study is to test the efficacy of stromal cell-derived factor-1α (SDF-1α)-coated coils together with endothelial progenitor cells (EPCs) transplantation in occluding aneurysms. Bone marrow-derived EPC surface markers were analyzed using flow cytometry. The migratory function of EPCs in response to SDF-1α was evaluated using a modified Boyden chamber assay. Capillary-like tube formation was assessed using Matrigel gel. Coil morphologies before and after coating with SDF-1α were observed under a scanning electron microscope. The level of SDF-1α in supernatants was measured by ELISA. Sprague-Dawley rats were randomly allocated into five groups. Histological analysis was performed on days 14 and 28 after coil implantation. The bone marrow-EPCs could express CD133, CD34, and VEGFR-2 and form tubule-like structures in vitro. Migratory ability of EPCs in the presence of SDF-1α-coated coils was similar to that in the presence of 5 ng/ml SDF-1α gradient. Sustained release of SDF-1α was achieved using silk fibroin as a carrier. In SDF-1α-coated coils + EPCs transplantation group, a well-organized fibrous tissue bridging the orifice of aneurysms was shown on days 14 and 28. On day 28, tissue organization was greater in the SDF-1α-coated coils group than in the unmodified coils group. Immunofluorescence showed α-smooth muscle actin-positive cells in organized tissue in sacs. Combined treatment with SDF-1α-coated coils and EPCs transplantation is a safe and effective treatment for rat aneurysms. This may provide a new strategy for endovascular therapy following aneurysmal subarachnoid hemorrhage. PMID:25674201

  15. Effect of High-Dose Atorvastatin Reload on the Release of Endothelial Progenitor Cells in Patients on Long-Term Statin Treatment Who Underwent Percutaneous Coronary Intervention (from the ARMYDA-EPC Study).

    PubMed

    Ricottini, Elisabetta; Madonna, Rosalinda; Grieco, Domenico; Zoccoli, Alice; Stampachiacchiere, Barbara; Patti, Giuseppe; Tonini, Giuseppe; De Caterina, Raffaele; Di Sciascio, Germano

    2016-01-15

    Endothelial progenitor cells (EPCs) may concur to endogenous vascular repair. Previous studies have reported that statin treatment increases EPC levels. We investigated whether this occurs in patients on long-term statin treatment who underwent percutaneous coronary interventions (PCIs). In a phase A study, 53 patients (atorvastatin reload [AR] 80 mg 12 hours before + 40 mg 2 hours before PCI, n = 27; placebo [P], n = 26) were evaluated for EPC mobilization as CD45dim/CD34+/CD133+/KDR+ cell number by flow cytometry. Assays were run at randomization (12 hours before PCI, R), immediately before PCI (T0) at 8 (T8) and 24 hours (T24). In phase B study, 50 patients (AR, n = 25; P, n = 25) were evaluated for early colony formation by Hill colony forming unit (CFU) assay, with sampling at randomization and 24 hours later. In phase A, EPCs levels were similar at randomization between 2 arms (0.23% [0.14 to 0.54] of total events in AR vs 0.22% [0.04 to 0.37] in P group; p = 0.33). At PCI, EPC levels were higher in AR arm (0.42% [0.06 to 0.30] vs 0.19% [0.06 to 030]; p = 0.009). Higher EPC levels in AR group were also found at 8 and 24 hours. In phase B, EPC CFUs/well numbers at randomization were similar in the 2 arms (8 [6 to 12] in AR vs 12 [6 to 20] in P group, p = 0.109). EPC CFU/well at 24 hours became significantly higher in AR arm (17 [10 to 23] vs 5 [2 to 13], p = 0.002). In conclusion, high-dose AR before PCI in patients on long-term statin therapy promptly increases EPCs mobilization, which are capable of early colony formation and may contribute to cardioprotection. PMID:26743348

  16. Bee Venom Accelerates Wound Healing in Diabetic Mice by Suppressing Activating Transcription Factor-3 (ATF-3) and Inducible Nitric Oxide Synthase (iNOS)-Mediated Oxidative Stress and Recruiting Bone Marrow-Derived Endothelial Progenitor Cells.

    PubMed

    Badr, Gamal; Hozzein, Wael N; Badr, Badr M; Al Ghamdi, Ahmad; Saad Eldien, Heba M; Garraud, Olivier

    2016-10-01

    Multiple mechanisms contribute to impaired diabetic wound healing including impaired neovascularization and deficient endothelial progenitor cell (EPC) recruitment. Bee venom (BV) has been used as an anti-inflammatory agent for the treatment of several diseases. Nevertheless, the effect of BV on the healing of diabetic wounds has not been studied. Therefore, in this study, we investigated the impact of BV on diabetic wound closure in a type I diabetic mouse model. Three experimental groups were used: group 1, non-diabetic control mice; group 2, diabetic mice; and group 3, diabetic mice treated with BV. We found that the diabetic mice exhibited delayed wound closure characterized by a significant decrease in collagen production and prolonged elevation of inflammatory cytokines levels in wounded tissue compared to control non-diabetic mice. Additionally, wounded tissue in diabetic mice revealed aberrantly up-regulated expression of ATF-3 and iNOS followed by a marked elevation in free radical levels. Impaired diabetic wound healing was also characterized by a significant elevation in caspase-3, -8, and -9 activity and a marked reduction in the expression of TGF-β and VEGF, which led to decreased neovascularization and angiogenesis of the injured tissue by impairing EPC mobilization. Interestingly, BV treatment significantly enhanced wound closure in diabetic mice by increasing collagen production and restoring the levels of inflammatory cytokines, free radical, TGF-β, and VEGF. Most importantly, BV-treated diabetic mice exhibited mobilized long-lived EPCs by inhibiting caspase activity in the wounded tissue. Our findings reveal the molecular mechanisms underlying improved diabetic wound healing and closure following BV treatment. J. Cell. Physiol. 231: 2159-2171, 2016. © 2016 Wiley Periodicals, Inc. PMID:26825453

  17. Isolation, Culture, and Characterization of Human Umbilical Cord Blood-Derived Mesenchymal Stromal Cells.

    PubMed

    Bieback, Karen; Netsch, Philipp

    2016-01-01

    Umbilical cord blood (CB) is considered one of the youngest available sources of adult stem cells. Besides hematopoietic stem cells, CB has been shown to contain endothelial progenitor cells as well as mesenchymal stromal/stem cells (MSC). To isolate MSC from cord blood, CB is collected into a sterile bag containing the anticoagulant citrate-phosphate-dextrose (CPD). The CB is then processed by density-gradient centrifugation to obtain mononuclear cells (MNC). These are cultured until the outgrowth of fibroblastoid cell colonies appears. After reaching a subconfluent stage, cells are harvested, expanded, and characterized as cord blood mesenchymal stromal cells (CB-MSC) according to standard criteria: plastic adherence, fibroblast morphology, CFU-f assay, proliferation potential, immune phenotype, and differentiation potential.Apparently, the frequency of MSC in CB is extremely low. Thus, not every CB unit will provide adequate MSC isolation yields. Different strategies have been proposed aiming to optimize the isolation success by selecting CB units of optimal quality. It is commonly agreed on that a high CB volume, a high cellular content, and a short time frame between birth and MSC isolation are criteria that will enhance the MSC isolation success.The procedures in this chapter are standardized protocols that were established and optimized in the authors' research laboratory; however, various modifications of the protocols are possible. PMID:27236676

  18. Cord Blood-Derived Hematopoietic Stem/Progenitor Cells: Current Challenges in Engraftment, Infection, and Ex Vivo Expansion

    PubMed Central

    Kita, Katsuhiro; Lee, Jong O.; Finnerty, Celeste C.; Herndon, David N.

    2011-01-01

    Umbilical cord blood has served as an alternative to bone marrow for hematopoietic transplantation since the late 1980s. Numerous clinical studies have proven the efficacy of umbilical cord blood. Moreover, the possible immaturity of cells in umbilical cord blood gives more options to recipients with HLA mismatch and allows for the use of umbilical cord blood from unrelated donors. However, morbidity and mortality rates associated with hematopoietic malignancies still remain relatively high, even after cord blood transplantation. Infections and relapse are the major causes of death after cord blood transplantation in patients with hematopoietic diseases. Recently, new strategies have been introduced to improve these major problems. Establishing better protocols for simple isolation of primitive cells and ex vivo expansion will also be very important. In this short review, we discuss several recent promising findings related to the technical improvement of cord blood transplantation. PMID:21603139

  19. Progenitor Epithelium

    PubMed Central

    Marty-Santos, Leilani

    2015-01-01

    Insulin-producing β cells within the vertebrate fetal pancreas acquire their fate in a step-wise manner. Whereas the intrinsic factors dictating the transcriptional or epigenetic status of pancreatic lineages have been intensely examined, less is known about cell–cell interactions that might constitute a niche for the developing β cell lineage. It is becoming increasingly clear that understanding and recapitulating these steps may instruct in vitro differentiation of embryonic stem cells and/or therapeutic regeneration. Indeed, directed differentiation techniques have improved since transitioning from 2D to 3D cultures, suggesting that the 3D microenvironment in which β cells are born is critical. However, to date, it remains unknown whether the changing architecture of the pancreatic epithelium impacts the fate of cells therein. An emerging challenge in the field is to elucidate how progenitors are allocated during key events, such as the stratification and subsequent resolution of the pre-pancreatic epithelium, as well as the formation of lumens and branches. Here, we assess the progenitor epithelium and examine how it might influence the emergence of pancreatic multipotent progenitors (MPCs), which give rise to β cells and other pancreatic lineages. PMID:26216134

  20. Simultaneous characterization of progenitor cell compartments in adult human liver.

    PubMed

    Porretti, Laura; Cattaneo, Alessandra; Colombo, Federico; Lopa, Raffaella; Rossi, Giorgio; Mazzaferro, Vincenzo; Battiston, Carlo; Svegliati-Baroni, Gianluca; Bertolini, Francesco; Rebulla, Paolo; Prati, Daniele

    2010-01-01

    The human liver is a complex tissue consisting of epithelial, endothelial, hematopoietic, and mesenchymal elements that probably derive from multiple lineage-committed progenitors, but no comprehensive study aimed at identifying and characterizing intrahepatic precursors has yet been published. Cell suspensions for this study were obtained by enzymatic digestion of liver specimens taken from 20 patients with chronic liver disease and 13 multiorgan donors. Stem and progenitor cells were first isolated, amplified, and characterized ex vivo according to previously validated methods, and then optimized flow cytometry was used to assess their relative frequencies and characterize their immunophenotypes in the clinical specimens. Stem and progenitor cells committed to hematopoietic, endothelial, epithelial, and mesenchymal lineages were clearly identifiable in livers from both healthy and diseased subjects. Within the mononuclear liver cell compartment, epithelial progenitors [epithelial cell adhesion molecule (EpCAM)(+)/CD49f(+)/CD29(+)/CD45(-)] accounted for 2.7-3.5% whereas hematopoietic (CD34(+)/CD45(+)), endothelial [vascular endothelial growth factor-2 (KDR)(+)/CD146(+)/CD45(-)], and mesenchymal [CD73(+)/CD105(+)/CD90 (Thy-1)(+)/CD45 (-)] stem cells and progenitors accounted for smaller fractions (0.02-0.6%). The patients' livers had higher percentages of hematopoietic and endothelial precursors than those of the donors. In conclusion, we identified and characterized precursors committed to four different lineages in adult human liver. We also optimized a flow cytometry approach that will be useful in exploring the contribution of these cells to the pathogenesis of liver disease. PMID:19960544

  1. Full-length dysferlin expression driven by engineered human dystrophic blood derived CD133+ stem cells.

    PubMed

    Meregalli, Mirella; Navarro, Claire; Sitzia, Clementina; Farini, Andrea; Montani, Erica; Wein, Nicolas; Razini, Paola; Beley, Cyriaque; Cassinelli, Letizia; Parolini, Daniele; Belicchi, Marzia; Parazzoli, Dario; Garcia, Luis; Torrente, Yvan

    2013-12-01

    The protein dysferlin is abundantly expressed in skeletal and cardiac muscles, where its main function is membrane repair. Mutations in the dysferlin gene are involved in two autosomal recessive muscular dystrophies: Miyoshi myopathy and limb-girdle muscular dystrophy type 2B. Development of effective therapies remains a great challenge. Strategies to repair the dysferlin gene by skipping mutated exons, using antisense oligonucleotides (AONs), may be suitable only for a subset of mutations, while cell and gene therapy can be extended to all mutations. AON-treated blood-derived CD133+ stem cells isolated from patients with Miyoshi myopathy led to partial dysferlin reconstitution in vitro but failed to express dysferlin after intramuscular transplantation into scid/blAJ dysferlin null mice. We thus extended these experiments producing the full-length dysferlin mediated by a lentiviral vector in blood-derived CD133+ stem cells isolated from the same patients. Transplantation of engineered blood-derived CD133+ stem cells into scid/blAJ mice resulted in sufficient dysferlin expression to correct functional deficits in skeletal muscle membrane repair. Our data suggest for the first time that lentivirus-mediated delivery of full-length dysferlin in stem cells isolated from Miyoshi myopathy patients could represent an alternative therapeutic approach for treatment of dysferlinopathies. PMID:24028392

  2. 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.

  3. Postinfarction Functional Recovery Driven by a Three-Dimensional Engineered Fibrin Patch Composed of Human Umbilical Cord Blood-Derived Mesenchymal Stem Cells

    PubMed Central

    Roura, Santiago; Soler-Botija, Carolina; Bagó, Juli R.; Llucià-Valldeperas, Aida; Férnandez, Marco A.; Gálvez-Montón, Carolina; Prat-Vidal, Cristina; Perea-Gil, Isaac; Blanco, Jerónimo

    2015-01-01

    Considerable research has been dedicated to restoring myocardial cell slippage and limiting ventricular remodeling after myocardial infarction (MI). We examined the ability of a three-dimensional (3D) engineered fibrin patch filled with human umbilical cord blood-derived mesenchymal stem cells (UCBMSCs) to induce recovery of cardiac function after MI. The UCBMSCs were modified to coexpress luciferase and fluorescent protein reporters, mixed with fibrin, and applied as an adhesive, viable construct (fibrin-cell patch) over the infarcted myocardium in mice (MI-UCBMSC group). The patch adhered well to the heart. Noninvasive bioluminescence imaging demonstrated early proliferation and differentiation of UCBMSCs within the construct in the postinfarct mice in the MI-UCBMSC group. The implanted cells also participated in the formation of new, functional microvasculature that connected the fibrin-cell patch to both the subjacent myocardial tissue and the host circulatory system. As revealed by echocardiography, the left ventricular ejection fraction and fractional shortening at sacrifice were improved in MI-UCBMSC mice and were markedly reduced in mice treated with fibrin alone and untreated postinfarction controls. In conclusion, a 3D engineered fibrin patch composed of UCBMSCs attenuated infarct-derived cardiac dysfunction when transplanted locally over a myocardial wound. Significance Ischemic heart failure (HF) is the end stage of many cardiovascular diseases, including myocardial infarction. The only definitive treatment for HF is cardiac transplant, which is hampered by limited number of heart donors and graft rejection. In recent times, cellular cardiomyoplasty has been expected to repair infarcted myocardium by implantation of different sources of stem or progenitor cells. However, low cell survival and myocardial implantation rates have motivated the emergence of novel approaches with the objective of generating graftable cell-based implants. Here, the potential

  4. Caspase-1 mediates hyperlipidemia-weakened progenitor cell vessel repair

    PubMed Central

    Li, Ya-Feng; Huang, Xiao; Li, Xinyuan; Gong, Ren; Yin, Ying; Nelson, Jun; Gao, Erhe; Zhang, Hongyu; Hoffman, Nicholas E.; Houser, Steven R.; Madesh, Muniswamy; Tilley, Douglas G.; Choi, Eric T.; Jiang, Xiaohua; Huang, Cong-Xin; Wang, Hong; Yang, Xiao-Feng

    2015-01-01

    Caspase-1 activation senses metabolic danger-associated molecular patterns (DAMPs) and mediates the initiation of inflammation in endothelial cells. Here, we examined whether the caspase-1 pathway is responsible for sensing hyperlipidemia as a DAMP in bone marrow (BM)-derived Stem cell antigen-1 positive (Sca-1+) stem/progenitor cells and weakening their angiogenic ability. Using biochemical methods, gene knockout, cell therapy and myocardial infarction (MI) models, we had the following findings: 1) Hyperlipidemia induces caspase-1 activity in mouse Sca-1+ progenitor cells in vivo; 2) Caspase-1 contributes to hyperlipidemia-induced modulation of vascular cell death-related gene expression in vivo; 3) Injection of Sca-1+ progenitor cells from caspase-1−/− mice improves endothelial capillary density in heart and decreases cardiomyocyte death in a mouse model of MI; and 4) Caspase-1−/− Sca-1+ progenitor cell therapy improves mouse cardiac function after MI. Our results provide insight on how hyperlipidemia activates caspase-1 in Sca-1+ progenitor cells, which subsequently weakens Sca-1+ progenitor cell repair of vasculature injury. These results demonstrate the therapeutic potential of caspase-1 inhibition in improving progenitor cell therapy for MI. PMID:26709768

  5. Circulating Hematopoietic Progenitor Cells are Decreased in COPD

    PubMed Central

    Janssen, William J.; Yunt, Zulma X.; Muldrow, Alaina; Kearns, Mark T.; Kloepfer, Angela; Barthel, Lea; Bratton, Donna L.; Bowler, Russell P.; Henson, Peter M.

    2014-01-01

    Rationale Bone marrow derived progenitor cells participate in the repair of injured vessels. The lungs of individuals with emphysema have reduced alveolar capillary density and increased endothelial apoptosis. We hypothesized that circulating levels of endothelial and hematopoietic progenitor cells would be reduced in this group of patients. Objectives The goal of this study was to measure circulating levels of endothelial progenitor cells (EPCs) and hematopoietic progenitor cells (HPCs) in subjects with COPD and to determine if progenitor levels correlated with disease severity and the presence of emphysema. Methods Peripheral blood mononuclear cells were isolated from 61 patients with COPD and 32 control subjects. Levels of EPCs (CD45dim CD34+ ) and HPCs (CD45+ CD34+ VEGF-R2+) were quantified using multi-parameter flow cytometry. Progenitor cell function was assessed using cell culture assays. All subjects were evaluated with spirometry and CT scanning. Measurements and Main Results HPC levels were reduced in subjects with COPD compared to controls, whereas circulating EPC levels were similar between the two groups. HPC levels correlated with severity of obstruction and were lowest in subjects with severe emphysema. These associations remained after correction for factors known to affect progenitor cell levels including age, smoking status, the use of statin medications and the presence of coronary artery disease. The ability of mononuclear cells to form endothelial cell colony forming units (EC-CFU) was also reduced in subjects with COPD. Conclusions HPC levels are reduced in subjects with COPD and correlate with emphysema phenotype and severity of obstruction. Reduction of HPCs may disrupt maintenance of the capillary endothelium, thereby contributing to the pathogenesis of COPD. PMID:24182349

  6. Development and bioevaluation of nanofibers with blood-derived growth factors for dermal wound healing.

    PubMed

    Bertoncelj, Valentina; Pelipenko, Jan; Kristl, Julijana; Jeras, Matjaž; Cukjati, Marko; Kocbek, Petra

    2014-09-01

    The aim of our work was to produce a modern nanomaterial with incorporated blood-derived growth factors, produced by electrospinning, applicable in treatment of chronic wounds. Platelet-rich plasma was chosen as a natural source of growth factors. Results showed that platelet-rich plasma stimulates keratinocyte and fibroblast cell growth in vitro. Its optimal concentration in growth medium was 2% (v/v) for both types of skin cells, while higher concentrations caused alterations in cell morphology, with reduced cell mobility and proliferation. In the next step hydrophilic nanofibers loaded with platelet-rich plasma were produced from chitosan and poly(ethylene oxide), using electrospinning. The morphology of nanofibers was stable in aqueous conditions for 72 h. It was shown that electrospinning does not adversely affect the biological activity of platelet-rich plasma. The effects of nanofibers with incorporated platelet-rich plasma on cell proliferation, survival, morphology and mobility were examined. Nanofibers limited cell mobility, changed morphology and stimulated cell proliferation. Despite of the small amount of blood-derived growth factors introduced in cell culture via platelet-rich plasma-loaded nanofibers, such nanofibrillar support significantly induced cell proliferation, indicating synergistic effect of nanotopography and incorporated growth factors. The overall results confirm favorable in vitro properties of produced nanofibers, indicating their high potential as a nanomaterial suitable for delivery of platelet-rich plasma in wound healing applications. PMID:24931341

  7. The Possible Roles of Biological Bone Constructed with Peripheral Blood Derived EPCs and BMSCs in Osteogenesis and Angiogenesis.

    PubMed

    Wu, Li; Zhao, Xian; He, Bo; Jiang, Jie; Xie, Xiao-Jie; Liu, Liu

    2016-01-01

    This study aimed to determine the possible potential of partially deproteinized biologic bone (PDPBB) seeded with bone marrow stromal cells (BMSCs) and endothelial progenitor cells (EPCs) in osteogenesis and angiogenesis. BMSCs and EPCs were isolated, identified, and cocultured in vitro, followed by seeding on the PDPBB. Expression of osteogenesis and vascularization markers was quantified by immunofluorescence (IF) staining, immunohistochemistry (IHC), and quantitive real-time polymerase chain reaction (qRT-PCR). Scanning electron microscope (SEM) was also employed to further evaluate the morphologic alterations of cocultured cells in the biologic bone. Results demonstrated that the coculture system combined with BMSCs and EPCs had significant advantages of (i) upregulating the mRNA expression of VEGF, Osteonectin, Osteopontin, and Collagen Type I and (ii) increasing ALP and OC staining compared to the BMSCs or EPCs only group. Moreover, IHC staining for CD105, CD34, and ZO-1 increased significantly in the implanted PDPBB seeded with coculture system, compared to that of BMSCs or EPCs only, respectively. Summarily, the present data provided evidence that PDPBB seeded with cocultured system possessed favorable cytocompatibility, provided suitable circumstances for different cell growth, and had the potential to provide reconstruction for cases with bone defection by promoting osteogenesis and angiogenesis. PMID:27195296

  8. The Possible Roles of Biological Bone Constructed with Peripheral Blood Derived EPCs and BMSCs in Osteogenesis and Angiogenesis

    PubMed Central

    Wu, Li; Zhao, Xian; He, Bo; Jiang, Jie; Xie, Xiao-Jie; Liu, Liu

    2016-01-01

    This study aimed to determine the possible potential of partially deproteinized biologic bone (PDPBB) seeded with bone marrow stromal cells (BMSCs) and endothelial progenitor cells (EPCs) in osteogenesis and angiogenesis. BMSCs and EPCs were isolated, identified, and cocultured in vitro, followed by seeding on the PDPBB. Expression of osteogenesis and vascularization markers was quantified by immunofluorescence (IF) staining, immunohistochemistry (IHC), and quantitive real-time polymerase chain reaction (qRT-PCR). Scanning electron microscope (SEM) was also employed to further evaluate the morphologic alterations of cocultured cells in the biologic bone. Results demonstrated that the coculture system combined with BMSCs and EPCs had significant advantages of (i) upregulating the mRNA expression of VEGF, Osteonectin, Osteopontin, and Collagen Type I and (ii) increasing ALP and OC staining compared to the BMSCs or EPCs only group. Moreover, IHC staining for CD105, CD34, and ZO-1 increased significantly in the implanted PDPBB seeded with coculture system, compared to that of BMSCs or EPCs only, respectively. Summarily, the present data provided evidence that PDPBB seeded with cocultured system possessed favorable cytocompatibility, provided suitable circumstances for different cell growth, and had the potential to provide reconstruction for cases with bone defection by promoting osteogenesis and angiogenesis. PMID:27195296

  9. 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.

  10. Osteogenic differentiation of GFP-labeled human umbilical cord blood derived mesenchymal stem cells after cryopreservation.

    PubMed

    Liu, Guangpeng; Ye, Xinhai; Zhu, Yuchang; Li, Yulin; Sun, Jian; Cui, Lei; Cao, Yilin

    2011-10-01

    The osteogenic capacity of human umbilical cord blood derived mesenchymal stem cells (UCB-MSCs) has been demonstrated both in vitro and in vivo. Therefore, cell labeling and storage are becoming necessary for researching the potential therapeutic use of UCB-MSCs for bone tissue engineering. The aim of this study was to determine the effect of cryopreservation on the osteogenic differentiation of green fluorescent protein (GFP)-marked UCB-MSCs in vitro. MSCs were isolated from full-term human UCB, expanded, transfected with the GFP gene, and then cryopreserved in liquid nitrogen for 4 weeks. After thawing, cell surface antigen markers and osteogenic potential were analyzed, and the luminescence of these cells was observed by fluorescence microscopy. The results demonstrate that cryopreservation has no effect on the cell phenotype, GFP expression or osteogenic differentiation of UCB-MSCs, showing that cryopreserved GFP-labeled UCB-MSCs might be applied for bone tissue engineering. PMID:21684270

  11. Effects of hypoxia on proliferation of human cord blood-derived mesenchymal stem cells.

    PubMed

    Peng, Longying; Shu, Xiaomei; Lang, Changhui; Yu, Xiaohua

    2016-08-01

    The purpose of our study was to examine the influence of hypoxia on proliferation of human umbilical cord blood-derived mesenchymal stem cells (hUCB-MSCs). The mononuclear cells were separated by density gradient centrifugation from human umbilical cord blood and then, respectively, cultured under hypoxia (5 % O2) or normoxia (20 % O2). Their cell morphology, cell surface markers, β-galactosidase staining, cell growth curve, DNA cycle, and the expression of hypoxia-inducible factor-1α (HIF-1α) were evaluated. We found that hypoxia, in part via HIF-1α, improved the proliferation efficiency, and prevented senescence of hUCB-MSCs without altering their morphology and surface markers. These results demonstrated that hypoxia provides a favorable culture condition to promote hUCB-MSCs proliferation in vitro, which is a better way to obtain sufficient numbers of hUCB-MSCs for research and certainly clinical application. PMID:25742732

  12. Bone marrow-derived Kruppel-like Factor 10 Controls Re-endothelialization in Response to Arterial Injury

    PubMed Central

    Wara, Akm Khyrul; Manica, Andre; Marchini, Julio F.; Sun, Xinghui; Icli, Basak; Tesmenitsky, Yevgenia; Croce, Kevin; Feinberg, Mark W.

    2013-01-01

    Objective The objective of this study was to investigate the role of Kruppel-like factor (KLF) 10, a zinc-finger transcription factor, in bone marrow-derived cell responses to arterial endothelial injury. Accumulating evidence indicates that bone marrow-derived progenitors are recruited to sites of vascular injury and contribute to endothelial repair. Approach and Results In response to carotid artery endothelial denudation, KLF10 mRNA expression was markedlyincreased in both bone marrow and circulating lin− progenitor cells. To examine the specific role for KLF10 in arterial re-endothelialization, we used two models of endothelial denudation (wire- and thermal-induced injury) of the carotid artery in WT and KLF10−/− mice. WT mice displayed higher areas of re-endothelialization compared to KLF10−/− mice following endothelial injury using either method. Bone marrow (BM) transplant studies revealed that re-constitution of KLF10−/− mice with WT BM fully rescued the defect in re-endothelialization and increased lin−CD34+KDR+ progenitors in the blood and injured carotid arteries. Conversely, reconstitution of WT mice with KLF10−/−BM re-capitulated the defects in re-endothelialization and peripheral cell progenitors. The media from cultured KLF10−/− BM progenitors was markedly inefficient at promoting endothelial cell growth and migration compared to the media from WT progenitors, indicative of defective paracrine trophic effects from KLF10−/− BM progenitors. Finally, BM-derived KLF10−/− lin− progenitors from reconstituted mice had reduced CXCR4 expression and impaired migratory responses. Conclusions Collectively, these observations demonstrate a protective role for BM-derived KLF10 in paracrine and homing responses important to arterial endothelial injury and highlight KLF10 as a possible therapeutic target to promote endothelial repair in vascular disease states. PMID:23685559

  13. N-cadherin Determines Individual Variations in the Therapeutic Efficacy of Human Umbilical Cord Blood-derived Mesenchymal Stem Cells in a Rat Model of Myocardial Infarction

    PubMed Central

    Lee, Eun Ju; Choi, Eue-Keun; Kang, Soo Kyoung; Kim, Gi-Hwan; Park, Ju Young; Kang, Hyun-Jae; Lee, Sae-Won; Kim, Keum-Hyun; Kwon, Jin Sook; Lee, Ki Hong; Ahn, Youngkeun; Lee, Ho-Jae; Cho, Hyun-Jai; Choi, Soo Jin; Oh, Won Il; Park, Young-Bae; Kim, Hyo-Soo

    2012-01-01

    In this study, we established and characterized human umbilical cord blood-derived mesenchymal stem cells (hUCB-MSCs) from four different donors. However, the hUCB-MSCs showed remarkable variations in their therapeutic efficacy for repairing rat infarcted myocardium (including the process of angiogenesis) 8 weeks after transplantation. In addition, we observed that the level of vascular endothelial growth factor (VEGF) is correlated with the therapeutic efficacy of the four hUCB-MSCs. Next, to investigate the practical application of hUCB-MSCs, we searched for surface signature molecules that could serve as indicators of therapeutic efficacy. The gene for N-cadherin was the only cell surface gene that was highly expressed in the most effective hUCB-MSCs, both at the transcriptional and translational levels. We observed downregulation and upregulation of VEGF in response to N-cadherin blocking and N-cadherin overexpression, respectively. Activation of extracellular signal-regulated kinase (ERK), but not protein kinase B, was increased when N-cadherin expression was increased, whereas disruption of N-cadherin-mediated cell–cell contact induced suppression of ERK activation and led to VEGF downregulation. Moreover, by investigating hUCB-MSCs overexpressing N-cadherin or N-cadherin knockdown hUCB-MSCs, we confirmed the in vivo function of N-cadherin. In addition, we observed that DiI-labeled hUCB-MSCs express N-cadherin in the peri-infarct area and interact with cardiomyocytes. PMID:22068423

  14. Simulated Microgravity Exerts an Age-Dependent Effect on the Differentiation of Cardiovascular Progenitors Isolated from the Human Heart.

    PubMed

    Fuentes, Tania I; Appleby, Nancy; Raya, Michael; Bailey, Leonard; Hasaniya, Nahidh; Stodieck, Louis; Kearns-Jonker, Mary

    2015-01-01

    Microgravity has a profound effect on cardiovascular function, however, little is known about the impact of microgravity on progenitors that reside within the heart. We investigated the effect of simulated microgravity exposure on progenitors isolated from the neonatal and adult human heart by quantifying changes in functional parameters, gene expression and protein levels after 6-7 days of 2D clinorotation. Utilization of neonatal and adult cardiovascular progenitors in ground-based studies has provided novel insight into how microgravity may affect cells differently depending on age. Simulated microgravity exposure did not impact AKT or ERK phosphorylation levels and did not influence cell migration, but elevated transcripts for paracrine factors were identified in neonatal and adult cardiovascular progenitors. Age-dependent responses surfaced when comparing the impact of microgravity on differentiation. Endothelial cell tube formation was unchanged or increased in progenitors from adults whereas neonatal cardiovascular progenitors showed a decline in tube formation (p<0.05). Von Willebrand Factor, an endothelial differentiation marker, and MLC2v and Troponin T, markers for cardiomyogenic differentiation, were elevated in expression in adult progenitors after simulated microgravity. DNA repair genes and telomerase reverse transcriptase which are highly expressed in early stem cells were increased in expression in neonatal but not adult cardiac progenitors after growth under simulated microgravity conditions. Neonatal cardiac progenitors demonstrated higher levels of MESP1, OCT4, and brachyury, markers for early stem cells. MicroRNA profiling was used to further investigate the impact of simulated microgravity on cardiovascular progenitors. Fifteen microRNAs were significantly altered in expression, including microRNAs-99a and 100 (which play a critical role in cell dedifferentiation). These microRNAs were unchanged in adult cardiac progenitors. The effect of

  15. Transcription factor expression in lipopolysaccharide-activated peripheral-blood-derived mononuclear cells

    PubMed Central

    Roach, Jared C.; Smith, Kelly D.; Strobe, Katie L.; Nissen, Stephanie M.; Haudenschild, Christian D.; Zhou, Daixing; Vasicek, Thomas J.; Held, G. A.; Stolovitzky, Gustavo A.; Hood, Leroy E.; Aderem, Alan

    2007-01-01

    Transcription factors play a key role in integrating and modulating biological information. In this study, we comprehensively measured the changing abundances of mRNAs over a time course of activation of human peripheral-blood-derived mononuclear cells (“macrophages”) with lipopolysaccharide. Global and dynamic analysis of transcription factors in response to a physiological stimulus has yet to be achieved in a human system, and our efforts significantly advanced this goal. We used multiple global high-throughput technologies for measuring mRNA levels, including massively parallel signature sequencing and GeneChip microarrays. We identified 92 of 1,288 known human transcription factors as having significantly measurable changes during our 24-h time course. At least 42 of these changes were previously unidentified in this system. Our data demonstrate that some transcription factors operate in a functional range below 10 transcripts per cell, whereas others operate in a range three orders of magnitude greater. The highly reproducible response of many mRNAs indicates feedback control. A broad range of activation kinetics was observed; thus, combinatorial regulation by small subsets of transcription factors would permit almost any timing input to cis-regulatory elements controlling gene transcription. PMID:17913878

  16. Tumorigenicity Evaluation of Umbilical Cord Blood-derived Mesenchymal Stem Cells

    PubMed Central

    Park, Sang-Jin; Kim, Hyun-Jung; Kim, Woojin; Kim, Ok-Sun; Lee, Sunyeong; Han, Su-Yeon; Jeong, Eun Ju; Park, Hyun-shin; Kim, Hea-Won; Moon, Kyoung-Sik

    2016-01-01

    Mesenchymal stem cells (MSCs) have been identified in multiple types of tissue and exhibit characteristic self-renewal and multi-lineage differentiation abilities. However, the possibility of oncogenic transformation after transplantation is concerning. In this study, we investigated the tumorigenic potential of umbilical cord blood-derived MSCs (hUCB-MSCs) relative to MRC-5 and HeLa cells (negative and positive controls, respectively) both in vitro and in vivo. To evaluate tumorigenicity in vitro, anchorage-independent growth was assessed using the soft agar colony formation assay. hUCB-MSCs and MRC-5 cells formed few colonies, while HeLa cells formed a greater number of larger colonies, indicating that hUCB-MSCs and MRC-5 cells do not have anchorage-independent proliferation potential. To detect tumorigenicity in vivo, hUCB-MSCs were implanted as a single subcutaneous injection into BALB/c-nu mice. No tumor formation was observed in mice transplanted with hUCB-MSCs or MRC-5 cells based on macroand microscopic examinations; however, all mice transplanted with HeLa cells developed tumors that stained positive for a human gene according to immunohistochemical analysis. In conclusion, hUCB-MSCs do not exhibit tumorigenic potential based on in vitro and in vivo assays under our experimental conditions, providing further evidence of their safety for clinical applications. PMID:27437093

  17. Monoclonal Antibodies as Probes for the Detection of Porcine Blood-Derived Food Ingredients.

    PubMed

    Ofori, Jack A; Hsieh, Yun-Hwa P

    2016-05-11

    The lack of effective methods to monitor the use of porcine blood-derived food ingredients (PBFIs) is a concern for the billions of individuals who avoid consuming blood. We therefore sought to develop a panel of porcine blood-specific monoclonal antibodies (mAbs) for use as probes in immunoassays for the detection of PBFIs. Ten selected mAbs were identified that react with either a 60 or 90 kDa protein in the plasma fraction or a 12 kDa protein in the red blood cell fraction of porcine blood. Western blot analysis of commercially produced PBFIs revealed that these antigenic proteins are not affected by various manufacturing processes. The utility of these mAbs was demonstrated in a prototype sandwich ELISA developed for this study using mAbs 19C5-E10 and 16F9-C11. The new assay is porcine blood-specific and capable of detecting ≤0.03% (v/v) of PBFIs in cooked (100 °C for 15 min) ground meats or fish. PMID:27135860

  18. Human Umbilical Cord Blood-Derived Mesenchymal Stem Cells Contribute to Chondrogenesis in Coculture with Chondrocytes

    PubMed Central

    Li, Xingfu; Duan, Li; Liang, Yujie; Zhu, Weimin; Xiong, Jianyi; Wang, Daping

    2016-01-01

    Human umbilical cord blood-derived mesenchymal stem cells (hUCB-MSCs) have been shown as the most potential stem cell source for articular cartilage repair. In this study, we aimed to develop a method for long-term coculture of human articular chondrocytes (hACs) and hUCB-MSCs at low density in vitro to determine if the low density of hACs could enhance the hUCB-MSC chondrogenic differentiation as well as to determine the optimal ratio of the two cell types. Also, we compared the difference between direct coculture and indirect coculture at low density. Monolayer cultures of hUCB-MSCs and hACs were investigated at different ratios, at direct cell-cell contact groups for 21 days. Compared to direct coculture, hUCB-MSCs and hACs indirect contact culture significantly increased type II collagen (COL2) and decreased type I collagen (COL1) protein expression levels. SRY-box 9 (SOX9) mRNA levels and protein expression were highest in indirect coculture. Overall, these results indicate that low density direct coculture induces fibrocartilage. However, indirect coculture in conditioned chondrocyte cell culture medium can increase expression of chondrogenic markers and induce hUCB-MSCs differentiation into mature chondrocytes. This work demonstrates that it is possible to promote chondrogenesis of hUCB-MSCs in combination with hACs, further supporting the concept of novel coculture strategies for tissue engineering. PMID:27446948

  19. Endothelial Lessons.

    PubMed

    Vanhoutte, Paul M

    2016-01-01

    This essay focuses on nine important lessons learned during more than thirty years of endothelial research. They include: the danger of hiding behind a word, the confusion generated by abbreviations, the need to define the physiological role of the response studied, the local role of endothelium- dependent responses, the strength of pharmacological analyses, endothelial dysfunction as consequence and cause of disease, the importance of rigorous protocols, the primacy of in vivo studies and the importance of serendipity. PMID:26638800

  20. Mass spectrometry in cancer biomarker research: a case for immunodepletion of abundant blood-derived proteins from clinical tissue specimens

    PubMed Central

    Prieto, DaRue A; Johann, Donald J; Wei, Bih-Rong; Ye, Xiaoying; Chan, King C; Nissley, Dwight V; Simpson, R Mark; Citrin, Deborah E; Mackall, Crystal L; Linehan, W Marston; Blonder, Josip

    2014-01-01

    The discovery of clinically relevant cancer biomarkers using mass spectrometry (MS)-based proteomics has proven difficult, primarily because of the enormous dynamic range of blood-derived protein concentrations and the fact that the 22 most abundant blood-derived proteins constitute approximately 99% of the total plasma protein mass. Immunodepletion of clinical body fluid specimens (e.g., serum/plasma) for the removal of highly abundant proteins is a reasonable and reproducible solution. Often overlooked, clinical tissue specimens also contain a formidable amount of highly abundant blood-derived proteins present in tissue-embedded networks of blood/lymph capillaries and interstitial fluid. Hence, the dynamic range impediment to biomarker discovery remains a formidable obstacle, regardless of clinical sample type (solid tissue and/or body fluid). Thus, we optimized and applied simultaneous immunodepletion of blood-derived proteins from solid tissue and peripheral blood, using clear cell renal cell carcinoma as a model disease. Integrative analysis of data from this approach and genomic data obtained from the same type of tumor revealed concordant key pathways and protein targets germane to clear cell renal cell carcinoma. This includes the activation of the lipogenic pathway characterized by increased expression of adipophilin (PLIN2) along with 'cadherin switching', a phenomenon indicative of transcriptional reprogramming linked to renal epithelial dedifferentiation. We also applied immunodepletion of abundant blood-derived proteins to various tissue types (e.g., adipose tissue and breast tissue) showing unambiguously that the removal of abundant blood-derived proteins represents a powerful tool for the reproducible profiling of tissue proteomes. Herein, we show that the removal of abundant blood-derived proteins from solid tissue specimens is of equal importance to depletion of body fluids and recommend its routine use in the context of biological discovery and

  1. Pulmonary vascular disease in mice xenografted with human BM progenitors from patients with pulmonary arterial hypertension

    PubMed Central

    Farha, Samar; Lichtin, Alan; Graham, Brian; George, Deepa; Aldred, Micheala; Hazen, Stanley L.; Loyd, James; Tuder, Rubin

    2012-01-01

    Hematopoietic myeloid progenitors released into the circulation are able to promote vascular remodeling through endothelium activation and injury. Endothelial injury is central to the development of pulmonary arterial hypertension (PAH), a proliferative vasculopathy of the pulmonary circulation, but the origin of vascular injury is unknown. In the present study, mice transplanted with BM-derived CD133+ progenitor cells from patients with PAH, but not from healthy controls, exhibited morbidity and/or death due to features of PAH: in situ thrombi and endothelial injury, angioproliferative remodeling, and right ventricular hypertrophy and failure. Myeloid progenitors from patients with heritable and/or idiopathic PAH all produced disease in xenografted mice. Analyses of hematopoietic transcription factors and colony formation revealed underlying abnormalities of progenitors that skewed differentiation toward the myeloid-erythroid lineage. The results of the present study suggest a causal role for hematopoietic stem cell abnormalities in vascular injury, right ventricular hypertrophy, and morbidity associated with PAH. PMID:22745307

  2. Morphogenesis of the lymphatic vasculature: A focus on new progenitors and cellular mechanisms important for constructing lymphatic vessels.

    PubMed

    Kazenwadel, Jan; Harvey, Natasha L

    2016-03-01

    Lymphatic vessels serve crucial roles in the regulation of tissue fluid homeostasis, dietary lipid absorption and immune cell trafficking. Defects in lymphatic vessel morphogenesis and function have been associated with lymphedema, obesity, hypertension and tumour metastasis. Morphogenetic events important for construction of the lymphatic vasculature during development include the specification and emergence of lymphatic endothelial progenitor cells, their differentiation and assembly into interconnected vessels and vascular remodeling, ultimately giving rise to a functional vascular network. Despite the embryonic origins of lymphatic endothelial progenitor cells being long debated, work performed over the last decade had overwhelmingly supported at least a great majority of progenitor cells arising from the venous vasculature. Here, we review the most recent advances in the field of lymphatic vessel morphogenesis, with a focus on studies that have identified novel sources of embryonic lymphatic endothelial progenitor cells, together with the cellular mechanisms by which lymphatic vessels are initially assembled. PMID:26228815

  3. Development and validation of risk index for cognitive decline using blood-derived markers

    PubMed Central

    Ayonayon, Hilsa; Harris, Tamara; Phillips, Caroline; Rosano, Caterina; Satterfield, Suzanne; Yaffe, Kristine

    2015-01-01

    Objective: We sought to develop and validate a risk index for prospective cognitive decline in older adults based on blood-derived markers. Methods: The index was based on 8 markers that have been previously associated with cognitive aging: APOE genotype, plasma β-amyloid 42/40 ratio, telomere length, cystatin C, glucose, C-reactive protein, interleukin-6, and albumin. The outcome was person-specific cognitive slopes (Modified Mini-Mental State Examination) from 11 years of follow-up. A total of 1,445 older adults comprised the development sample. An index based on dichotomized markers was divided into low-, medium-, and high-risk categories; the risk categories were validated with the remaining sample (n = 739) using linear regression. Amyloid was measured on a subsample (n = 865) and was included only in a secondary index. Results: The risk categories showed significant differences from each other and were predictive of prospective cognitive decline in the validation sample, even after adjustment for age and baseline cognitive score: the low-risk group (24.8%) declined 0.32 points/y (95% confidence interval [CI]: −0.46, −0.19), the medium-risk group (58.7%) declined 0.55 points/y (95% CI: −0.65, 0.45), and the high-risk group (16.6%) declined 0.69 points/y (95% CI: −0.85, −0.54). Using the secondary index, which included β-amyloid 42/40 (validation n = 279), the low-risk group (26.9%) declined 0.20 points/y (95% CI: −0.42, 0.01), the medium-risk group (61.3%) declined 0.55 points/y (95% CI: −0.72, −0.38), and the high-risk group (11.8%) declined 0.83 points/y (95% CI: −1.14, −0.51). Conclusions: A risk index based on 8 blood-based markers was modestly able to predict cognitive decline over an 11-year follow-up. Further validation in other cohorts is necessary. PMID:25609760

  4. Blood-derived proteins in milk at start of lactation: Indicators of active or passive transfer.

    PubMed

    Wall, Samantha K; Gross, Josef J; Kessler, Evelyne C; Villez, Kris; Bruckmaier, Rupert M

    2015-11-01

    Colostrum has a different composition compared with milk in established lactation. This difference is in part due to the partially open blood-milk barrier, which, when closed, is designed to prevent the interdiffusion of blood and milk components. In the first days of lactation, α-lactalbumin (α-LA), a milk protein, is typically present in blood and several blood-derived proteins are also present in milk, such as IgG1, IgG2, serum albumin (SA), and lactate dehydrogenase (LDH). With the exception of IgG1, which is known to be transferred by active transcellular transport, the other proteins are thought to pass paracellularly through the temporarily open barrier. Along with an exchange of blood and milk components, somatic cell count (SCC) is typically high in colostrum. The decline of these proteins and SCC can be used as indicators to determine transcellular or paracellular transport. Two hypotheses were tested. The first hypothesis was that the decline curve for a protein or SCC would be the same as IgG1, indicating transcellular transport, or the decline curve would be different than IgG1, indicating paracellular transport. The second hypothesis was that the decline curves of SCC and all proteins that are thought to have paracellular transport would be the same. Ten Holstein cows were milked at 4 h after parturition, the next 5 consecutive milkings, and the afternoon milking on d 5, 8, 10, and 14 of lactation for a total of 10 milking time points, and sequential jugular blood samples were also taken. Blood and milk samples were analyzed for the concentrations of LDH, SA, IgG1, IgG2, and α-LA and milk samples were measured for SCC. Protein concentration and SCC curves were generated from all 10 time points and were evaluated using the tau time constant model to determine the rate of decline of the slope of each protein. When examining the first hypothesis, the concentration of IgG1 declined significantly faster in the milk than the proteins IgG2 and LDH, but

  5. Cord blood-derived cytokine-induced killer cellular therapy plus radiation therapy for esophageal cancer: a case report.

    PubMed

    Wang, Liming; Huang, Shigao; Dang, Yazheng; Li, Ming; Bai, Wen; Zhong, Zhanqiang; Zhao, Hongliang; Li, Yang; Liu, Yongjun; Wu, Mingyuan

    2014-12-01

    Esophageal cancer is a serious malignancy with regards to mortality and prognosis. Current treatment options include multimodality therapy mainstays of current treatment including surgery, radiation, and chemotherapy. Cell therapy for esophageal cancer is an advancing area of research. We report a case of esophageal cancer following cord blood-derived cytokine-induced killer cell infusion and adjuvant radiotherapy. Initially, she presented with poor spirit, full liquid diets, and upper abdominal pain. Through cell therapy plus adjuvant radiotherapy, the patient remitted and was self-reliant. Recognition of this curative effect of sequent therapy for esophageal cancer is important to enable appropriate treatment. This case highlights cord blood-derived cytokine-induced killer cell therapy significantly alleviates the adverse reaction of radiation and improves the curative effect. Cell therapy plus adjuvant radiotherapy can be a safe and effective treatment for esophageal cancer. PMID:25526496

  6. Red light, green light: Signals that control endothelial cell proliferation during embryonic vascular development

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The proper regulation of endothelial cell proliferation is critical for vascular development in the embryo. VEGF-A and bFGF, which are important in the induction of mesodermal progenitors to form a capillary plexus, are also key mitogenic signals. Disruption in VEGF-A or bFGF decreases endothelial c...

  7. Circulating Progenitor Cells and Vascular Dysfunction in Chronic Obstructive Pulmonary Disease

    PubMed Central

    Pizarro, Sandra; García-Lucio, Jéssica; Peinado, Víctor I.; Tura-Ceide, Olga; Díez, Marta; Blanco, Isabel; Sitges, Marta; Petriz, Jordi; Torralba, Yolanda; Marín, Pedro; Roca, Josep; Barberà, Joan Albert

    2014-01-01

    Background In chronic obstructive pulmonary disease (COPD), decreased progenitor cells and impairment of systemic vascular function have been suggested to confer higher cardiovascular risk. The origin of these changes and their relationship with alterations in the pulmonary circulation are unknown. Objectives To investigate whether changes in the number of circulating hematopoietic progenitor cells are associated with pulmonary hypertension or changes in endothelial function. Methods 62 COPD patients and 35 controls (18 non-smokers and 17 smokers) without cardiovascular risk factors other than cigarette smoking were studied. The number of circulating progenitors was measured as CD45+CD34+CD133+ labeled cells by flow cytometry. Endothelial function was assessed by flow-mediated dilation. Markers of inflammation and angiogenesis were also measured in all subjects. Results Compared with controls, the number of circulating progenitor cells was reduced in COPD patients. Progenitor cells did not differ between control smokers and non-smokers. COPD patients with pulmonary hypertension showed greater number of progenitor cells than those without pulmonary hypertension. Systemic endothelial function was worse in both control smokers and COPD patients. Interleukin-6, fibrinogen, high sensitivity C-reactive protein, vascular endothelial growth factor and tumor necrosis factor were increased in COPD. In COPD patients, the number of circulating progenitor cells was inversely related to the flow-mediated dilation of systemic arteries. Conclusions Pulmonary and systemic vascular impairment in COPD is associated with cigarette smoking but not with the reduced number of circulating hematopoietic progenitors. The latter appears to be a consequence of the disease itself not related to smoking habit. PMID:25171153

  8. Human umbilical cord blood-derived stem cells and brain-derived neurotrophic factor protect injured optic nerve: viscoelasticity characterization

    PubMed Central

    Lv, Xue-man; Liu, Yan; Wu, Fei; Yuan, Yi; Luo, Min

    2016-01-01

    The optic nerve is a viscoelastic solid-like biomaterial. Its normal stress relaxation and creep properties enable the nerve to resist constant strain and protect it from injury. We hypothesized that stress relaxation and creep properties of the optic nerve change after injury. More-over, human brain-derived neurotrophic factor or umbilical cord blood-derived stem cells may restore these changes to normal. To validate this hypothesis, a rabbit model of optic nerve injury was established using a clamp approach. At 7 days after injury, the vitreous body re-ceived a one-time injection of 50 μg human brain-derived neurotrophic factor or 1 × 106 human umbilical cord blood-derived stem cells. At 30 days after injury, stress relaxation and creep properties of the optic nerve that received treatment had recovered greatly, with patho-logical changes in the injured optic nerve also noticeably improved. These results suggest that human brain-derived neurotrophic factor or umbilical cord blood-derived stem cell intervention promotes viscoelasticity recovery of injured optic nerves, and thereby contributes to nerve recovery. PMID:27212930

  9. Human umbilical cord blood-derived stem cells and brain-derived neurotrophic factor protect injured optic nerve: viscoelasticity characterization.

    PubMed

    Lv, Xue-Man; Liu, Yan; Wu, Fei; Yuan, Yi; Luo, Min

    2016-04-01

    The optic nerve is a viscoelastic solid-like biomaterial. Its normal stress relaxation and creep properties enable the nerve to resist constant strain and protect it from injury. We hypothesized that stress relaxation and creep properties of the optic nerve change after injury. More-over, human brain-derived neurotrophic factor or umbilical cord blood-derived stem cells may restore these changes to normal. To validate this hypothesis, a rabbit model of optic nerve injury was established using a clamp approach. At 7 days after injury, the vitreous body re-ceived a one-time injection of 50 μg human brain-derived neurotrophic factor or 1 × 10(6) human umbilical cord blood-derived stem cells. At 30 days after injury, stress relaxation and creep properties of the optic nerve that received treatment had recovered greatly, with patho-logical changes in the injured optic nerve also noticeably improved. These results suggest that human brain-derived neurotrophic factor or umbilical cord blood-derived stem cell intervention promotes viscoelasticity recovery of injured optic nerves, and thereby contributes to nerve recovery. PMID:27212930

  10. Lung microvascular endothelium is enriched with progenitor cells that exhibit vasculogenic capacity.

    PubMed

    Alvarez, Diego F; Huang, Lan; King, Judy A; ElZarrad, M Khair; Yoder, Mervin C; Stevens, Troy

    2008-03-01

    Endothelial progenitor cells (EPCs) have been isolated postnatally from bone marrow, blood, and both the intima and adventitia of conduit vessels. However, it is unknown whether EPCs can be isolated from the lung microcirculation. Thus we sought to determine whether the microvasculature possesses EPCs capable of de novo vasculogenesis. Rat pulmonary artery (PAEC) and microvascular (PMVEC) endothelial cells were isolated and selected by using a single-cell clonogenic assay. Whereas the majority of PAECs (approximately 60%) were fully differentiated, the majority of PMVECs (approximately 75%) divided, with approximately 50% of the single cells giving rise to large colonies (>2,000 cells/colony). These highly proliferative cells exhibited the capacity to reconstitute the entire proliferative hierarchy of PMVECs, unveiling the existence of resident microvascular endothelial progenitor cells (RMEPCs). RMEPCs expressed endothelial cell markers (CD31, CD144, endothelial nitric oxide synthase, and von Willenbrand factor) and progenitor cell antigens (CD34 and CD309) but did not express the leukocyte marker CD45. Consistent with their origin, RMEPCs interacted with Griffonia simplicifolia and displayed restrictive barrier properties. In vitro and in vivo Matrigel assays revealed that RMEPCs possess vasculogenic capacity, forming ultrastructurally normal de novo vessels. Thus the pulmonary microcirculation is enriched with EPCs that display vasculogenic competence while maintaining functional endothelial microvascular specificity. PMID:18065657

  11. Mechanism study for hypoxia induced differentiation of insulin-producing cells from umbilical cord blood-derived mesenchymal stem cells.

    PubMed

    Sun, Bo; Meng, Xian-Hui; Liu, Rui; Yan, Shancheng; Xiao, Zhong-Dang

    2015-10-23

    Recently, we have successfully obtained functional IPCs efficiently from umbilical cord blood-derived mesenchymal stem cells by using hypoxia treatment. In this study, we further elaborated that the improved function and viability of IPCs are the result of the interaction β cell development pathway and c-Met/HGF axis induced by hypoxia. We found that hypoxia induced c-MET elevation is efficiently initiated the early stage differentiation IPCs from MSCs, and HGF improved the fully differentiation of IPCs by inducing the expression of NGN3. This finding may contribute to understanding β cell development and the development of stem cell therapy for diabetes. PMID:26392316

  12. Production of good manufacturing practice-grade human umbilical cord blood-derived mesenchymal stem cells for therapeutic use.

    PubMed

    Van Pham, Phuc; Phan, Ngoc Kim

    2015-01-01

    Human umbilical cord blood-derived mesenchymal stem cells (UCB-MSCs) are multipotent stem cells that can be differentiated into several specific cell types such as adipocytes, osteoblasts, and chondroblasts. They also were demonstrated to trans-differentiate into other cell lineages such as muscle cells and neurons. Thus, they are considered a promising stem cell source for therapeutic use. Here, we describe a method for production of good manufacturing practice-grade human UCB-MSCs for therapeutic use. The obtained UCB-MSCs are free of allogenous or xenogenous proteins. In addition, these MSCs could maintain the MSC phenotype in long-term culture. PMID:25239529

  13. Hemovascular Progenitors in the Kidney Require Sphingosine-1-Phosphate Receptor 1 for Vascular Development.

    PubMed

    Hu, Yan; Li, Minghong; Göthert, Joachim R; Gomez, R Ariel; Sequeira-Lopez, Maria Luisa S

    2016-07-01

    The close relationship between endothelial and hematopoietic precursors during early development of the vascular system suggested the possibility of a common yet elusive precursor for both cell types. Whether similar or related progenitors for endothelial and hematopoietic cells are present during organogenesis is unclear. Using inducible transgenic mice that specifically label endothelial and hematopoietic precursors, we performed fate-tracing studies combined with colony-forming assays and crosstransplantation studies. We identified a progenitor, marked by the expression of helix-loop-helix transcription factor stem cell leukemia (SCL/Tal1). During organogenesis of the kidney, SCL/Tal1(+) progenitors gave rise to endothelium and blood precursors with multipotential colony-forming capacity. Furthermore, appropriate morphogenesis of the kidney vasculature, including glomerular capillary development, arterial mural cell coating, and lymphatic vessel development, required sphingosine 1-phosphate (S1P) signaling via the G protein-coupled S1P receptor 1 in these progenitors. Overall, these results show that SCL/Tal1(+) progenitors with hemogenic capacity originate and differentiate within the early embryonic kidney by hemovasculogenesis (the concomitant formation of blood and vessels) and underscore the importance of the S1P pathway in vascular development. PMID:26534925

  14. The roles of blood-derived macrophages and resident microglia in the neuroinflammatory response to implanted Intracortical microelectrodes

    PubMed Central

    Ravikumar, Madhumitha; Sunil, Smrithi; Black, James; Barkauskas, Deborah S.; Haung, Alex Y.; Miller, Robert H.; Selkirk, Stephen M.; Capadona, Jeffrey R.

    2014-01-01

    Resident microglia and blood-borne macrophages have both been implicated to play a dominant role in mediating the neuroinflammatory response affecting implanted intracortical microelectrodes. However, the distinction between each cell type has not been demonstrated due to a lack of discriminating cellular markers. Understanding the subtle differences of each cell population in mediating neuroinflammation can aid in determining the appropriate therapeutic approaches to improve microelectrode performance. Therefore, the goal of this study is to characterize the role of infiltrating blood-derived cells, specifically macrophages, in mediating neuroinflammation following intracortical microelectrode implantation. Interestingly, we found no correlation between microglia and neuron populations at the microelectrode-tissue interface. On the other hand, blood-borne macrophages consistently dominated the infiltrating cell population following microelectrode implantation. Most importantly, we found a correlation between increased populations of blood-derived cells (including the total macrophage population) and neuron loss at the microelectrode-tissue interface. Specifically, the total macrophage population was greatest at two and sixteen weeks post implantation, at the same time points when we observed the lowest densities of neuronal survival in closest proximity to the implant. Together, our results suggest a dominant role of infiltrating macrophages, and not resident microglia, in mediating neurodegeneration following microelectrode implantation. PMID:24973296

  15. The roles of blood-derived macrophages and resident microglia in the neuroinflammatory response to implanted intracortical microelectrodes.

    PubMed

    Ravikumar, Madhumitha; Sunil, Smrithi; Black, James; Barkauskas, Deborah S; Haung, Alex Y; Miller, Robert H; Selkirk, Stephen M; Capadona, Jeffrey R

    2014-09-01

    Resident microglia and blood-borne macrophages have both been implicated to play a dominant role in mediating the neuroinflammatory response affecting implanted intracortical microelectrodes. However, the distinction between each cell type has not been demonstrated due to a lack of discriminating cellular markers. Understanding the subtle differences of each cell population in mediating neuroinflammation can aid in determining the appropriate therapeutic approaches to improve microelectrode performance. Therefore, the goal of this study is to characterize the role of infiltrating blood-derived cells, specifically macrophages, in mediating neuroinflammation following intracortical microelectrode implantation. Interestingly, we found no correlation between microglia and neuron populations at the microelectrode-tissue interface. On the other hand, blood-borne macrophages consistently dominated the infiltrating cell population following microelectrode implantation. Most importantly, we found a correlation between increased populations of blood-derived cells (including the total macrophage population) and neuron loss at the microelectrode-tissue interface. Specifically, the total macrophage population was greatest at two and sixteen weeks post implantation, at the same time points when we observed the lowest densities of neuronal survival in closest proximity to the implant. Together, our results suggest a dominant role of infiltrating macrophages, and not resident microglia, in mediating neurodegeneration following microelectrode implantation. PMID:24973296

  16. Umbilical cord blood-derived stem cells improve heat tolerance and hypothalamic damage in heat stressed mice.

    PubMed

    Tseng, Ling-Shu; Chen, Sheng-Hsien; Lin, Mao-Tsun; Lin, Ying-Chu

    2014-01-01

    Heatstroke is characterized by excessive hyperthermia associated with systemic inflammatory responses, which leads to multiple organ failure, in which brain disorders predominate. This definition can be almost fulfilled by a mouse model of heatstroke used in the present study. Unanesthetized mice were exposed to whole body heating (41.2°C for 1 hour) and then returned to room temperature (26°C) for recovery. Immediately after termination of whole body heating, heated mice displayed excessive hyperthermia (body core temperature ~42.5°C). Four hours after termination of heat stress, heated mice displayed (i) systemic inflammation; (ii) ischemic, hypoxic, and oxidative damage to the hypothalamus; (iii) hypothalamo-pituitary-adrenocortical axis impairment (reflected by plasma levels of both adrenocorticotrophic-hormone and corticosterone); (iv) decreased fractional survival; and (v) thermoregulatory deficits (e.g., they became hypothermia when they were exposed to room temperature). These heatstroke reactions can be significantly attenuated by human umbilical cord blood-derived CD34(+) cells therapy. Our data suggest that human umbilical cord blood-derived stem cells therapy may improve outcomes of heatstroke in mice by reducing systemic inflammation as well as hypothalamo-pituitary-adrenocortical axis impairment. PMID:24804231

  17. GAP JUNCTION COMMUNICATION MEDIATES TRANSFORMING GROWTH FACTOR-BETA ACTIVATION AND ENDOTHELIAL-INDUCED MURAL CELL DIFFERENTIATION

    Technology Transfer Automated Retrieval System (TEKTRAN)

    During blood vessel assembly, endothelial cells recruit mesenchymal progenitors and induce their differentiation into mural cells via contact-dependent transforming growth factor-beta (TGF-beta) activation. We investigated whether gap junction channels are formed between endothelial cells and recrui...

  18. Optimal Route for Human Umbilical Cord Blood-Derived Mesenchymal Stem Cell Transplantation to Protect Against Neonatal Hyperoxic Lung Injury: Gene Expression Profiles and Histopathology.

    PubMed

    Sung, Dong Kyung; Chang, Yun Sil; Ahn, So Yoon; Sung, Se In; Yoo, Hye Soo; Choi, Soo Jin; Kim, Soo Yoon; Park, Won Soon

    2015-01-01

    The aim of this study was to determine the optimal route of mesenchymal stem cell (MSC) transplantation. To this end, gene expression profiling was performed to compare the effects of intratracheal (i.t.) versus intravenous (i.v.) MSC administration. Furthermore, the therapeutic efficacy of each route to protect against neonatal hyperoxic lung injury was also determined. Newborn Sprague-Dawley rats were exposed to hyperoxia (90% oxygen) from birth for 14 days. Human umbilical cord blood-derived MSCs labeling with PKH26 were transplanted through either the i.t. (5×10(5)) or i.v. (2×10(6)) route at postnatal day (P) 5. At P14, lungs were harvested for histological, biochemical and microarray analyses. Hyperoxic conditions induced an increase in the mean linear intercept and mean alveolar volume (MAV), indicative of impaired alveolarization. The number of ED-1 positive cells was significantly decreased by both i.t. and i.v. transplantations. However, i.t. administration of MSCs resulted in a greater decrease in MAV and ED-1 positive cells compared to i.v. administration. Moreover, the number of TUNEL-positive cells was significantly decreased in the i.t. group, but not in the i.v. group. Although the i.t. group received only one fourth of the number of MSCs that the i.v. group did, a significantly higher number of donor cell-derived red PKH 26 positivity were recovered in the i.t. group. Hyperoxic conditions induced the up regulation of genes associated with the inflammatory response, such as macrophage inflammatory protein-1 α, tumor necrosis factor-α and inter leukin-6; genes associated with cell death, such as p53 and caspases; and genes associated with fibrosis, such as connective tissue growth factor. In contrast, hyperoxic conditions induced the dwon-regulation of vascular endothelial growth factor and hepatocyte growth factor. These hyperoxia-induced changes in gene expression were decreased in the i.t. group, but not in the i.v. group. Thus, local i.t. MSC

  19. Optimal Route for Human Umbilical Cord Blood-Derived Mesenchymal Stem Cell Transplantation to Protect Against Neonatal Hyperoxic Lung Injury: Gene Expression Profiles and Histopathology

    PubMed Central

    Ahn, So Yoon; Sung, Se In; Yoo, Hye Soo; Choi, Soo Jin; Kim, Soo Yoon; Park, Won Soon

    2015-01-01

    The aim of this study was to determine the optimal route of mesenchymal stem cell (MSC) transplantation. To this end, gene expression profiling was performed to compare the effects of intratracheal (IT) versus intravenous (IV) MSC administration. Furthermore, the therapeutic efficacy of each route to protect against neonatal hyperoxic lung injury was also determined. Newborn Sprague-Dawley rats were exposed to hyperoxia (90% oxygen) from birth for 14 days. Human umbilical cord blood-derived MSCs labeling with PKH26 were transplanted through either the IT (5×105) or IV (2×106) route at postnatal day (P) 5. At P14, lungs were harvested for histological, biochemical and microarray analyses. Hyperoxic conditions induced an increase in the mean linear intercept and mean alveolar volume (MAV), indicative of impaired alveolarization. The number of ED-1 positive cells was significantly decreased by both IT and IV transplantations. However, IT administration of MSCs resulted in a greater decrease in MAV and ED-1 positive cells compared to IV administration. Moreover, the number of TUNEL-positive cells was significantly decreased in the IT group, but not in the IV group. Although the IT group received only one fourth of the number of MSCs that the IV group did, a significantly higher number of donor cell-derived red PKH 26 positivity were recovered in the IT group. Hyperoxic conditions induced the up regulation of genes associated with the inflammatory response, such as macrophage inflammatory protein-1 α, tumor necrosis factor-α and inter leukin-6; genes associated with cell death, such as p53 and caspases; and genes associated with fibrosis, such as connective tissue growth factor. In contrast, hyperoxic conditions induced the dwon-regulation of vascular endothelial growth factor and hepatocyte growth factor. These hyperoxia-induced changes in gene expression were decreased in the IT group, but not in the IV group. Thus, local IT MSC transplantation was more effective

  20. Dengue Virus Infection of Mast Cells Triggers Endothelial Cell Activation ▿

    PubMed Central

    Brown, Michael G.; Hermann, Laura L.; Issekutz, Andrew C.; Marshall, Jean S.; Rowter, Derek; Al-Afif, Ayham; Anderson, Robert

    2011-01-01

    Vascular perturbation is a hallmark of severe forms of dengue disease. We show here that antibody-enhanced dengue virus infection of primary human cord blood-derived mast cells (CBMCs) and the human mast cell-like line HMC-1 results in the release of factor(s) which activate human endothelial cells, as evidenced by increased expression of the adhesion molecules ICAM-1 and VCAM-1. Endothelial cell activation was prevented by pretreatment of mast cell-derived supernatants with a tumor necrosis factor (TNF)-specific blocking antibody, thus identifying TNF as the endothelial cell-activating factor. Our findings suggest that mast cells may represent an important source of TNF, promoting vascular endothelial perturbation following antibody-enhanced dengue virus infection. PMID:21068256

  1. Identification of three molecular and functional subtypes in canine hemangiosarcoma through gene expression profiling and progenitor cell characterization.

    PubMed

    Gorden, Brandi H; Kim, Jong-Hyuk; Sarver, Aaron L; Frantz, Aric M; Breen, Matthew; Lindblad-Toh, Kerstin; O'Brien, Timothy D; Sharkey, Leslie C; Modiano, Jaime F; Dickerson, Erin B

    2014-04-01

    Canine hemangiosarcomas have been ascribed to an endothelial origin based on histologic appearance; however, recent findings suggest that these tumors may arise instead from hematopoietic progenitor cells. To clarify this ontogenetic dilemma, we used genome-wide expression profiling of primary hemangiosarcomas and identified three distinct tumor subtypes associated with angiogenesis (group 1), inflammation (group 2), and adipogenesis (group 3). Based on these findings, we hypothesized that a common progenitor may differentiate into the three tumor subtypes observed in our gene profiling experiment. To investigate this possibility, we cultured hemangiosarcoma cell lines under normal and sphere-forming culture conditions to enrich for tumor cell progenitors. Cells from sphere-forming cultures displayed a robust self-renewal capacity and exhibited genotypic, phenotypic, and functional properties consistent with each of the three molecular subtypes seen in primary tumors, including expression of endothelial progenitor cell (CD133 and CD34) and endothelial cell (CD105, CD146, and αvβ3 integrin) markers, expression of early hematopoietic (CD133, CD117, and CD34) and myeloid (CD115 and CD14) differentiation markers in parallel with increased phagocytic capacity, and acquisition of adipogenic potential. Collectively, these results suggest that canine hemangiosarcomas arise from multipotent progenitors that differentiate into distinct subtypes. Improved understanding of the mechanisms that determine the molecular and phenotypic differentiation of tumor cells in vivo could change paradigms regarding the origin and progression of endothelial sarcomas. PMID:24525151

  2. Identification of Three Molecular and Functional Subtypes in Canine Hemangiosarcoma through Gene Expression Profiling and Progenitor Cell Characterization

    PubMed Central

    Gorden, Brandi H.; Kim, Jong-Hyuk; Sarver, Aaron L.; Frantz, Aric M.; Breen, Matthew; Lindblad-Toh, Kerstin; O'Brien, Timothy D.; Sharkey, Leslie C.; Modiano, Jaime F.; Dickerson, Erin B.

    2015-01-01

    Canine hemangiosarcomas have been ascribed to an endothelial origin based on histologic appearance; however, recent findings suggest that these tumors may arise instead from hematopoietic progenitor cells. To clarify this ontogenetic dilemma, we used genome-wide expression profiling of primary hemangiosarcomas and identified three distinct tumor subtypes associated with angiogenesis (group 1), inflammation (group 2), and adipogenesis (group 3). Based on these findings, we hypothesized that a common progenitor may differentiate into the three tumor subtypes observed in our gene profiling experiment. To investigate this possibility, we cultured hemangiosarcoma cell lines under normal and sphere-forming culture conditions to enrich for tumor cell progenitors. Cells from sphere-forming cultures displayed a robust self-renewal capacity and exhibited genotypic, phenotypic, and functional properties consistent with each of the three molecular subtypes seen in primary tumors, including expression of endothelial progenitor cell (CD133 and CD34) and endothelial cell (CD105, CD146, and αvβ3 integrin) markers, expression of early hematopoietic (CD133, CD117, and CD34) and myeloid (CD115 and CD14) differentiation markers in parallel with increased phagocytic capacity, and acquisition of adipogenic potential. Collectively, these results suggest that canine hemangiosarcomas arise from multipotent progenitors that differentiate into distinct subtypes. Improved understanding of the mechanisms that determine the molecular and phenotypic differentiation of tumor cells in vivo could change paradigms regarding the origin and progression of endothelial sarcomas. PMID:24525151

  3. Photodynamic inactivation of Escherichia coli and Streptococcus mitis by cationic zinc(II) phthalocyanines in media with blood derivatives.

    PubMed

    Spesia, Mariana B; Rovera, Marisa; Durantini, Edgardo N

    2010-06-01

    The photodynamic inactivation (PDI) of Escherichia coli and Streptococcus mitis sensitized by cationic phthalocyanines was studied in different media containing blood derivatives. First, the activity of zinc(II) tetramethyltetrapyridino[3,4-b:3',4'-g:3'',4''-l:3''',4'''-q]porphyrazinium (ZnAPc4+), zinc(II) 2,9,16,23-tetrakis[4-(N-methylpyridyloxy)]phthalocyanine (ZnPPc4+) and zinc(II) 2,9,16,23-tetrakis[2-(N,N,N-trimethylamino)ethoxy]phthalocyanine (ZnEPc4+) were compared to photoinactivate these bacteria in saline solutions. After visible light irradiation, a higher photoinactivation of E. coli cells was found for ZnPPc4+, while ZnEPc4+ was the more effective sensitizer to eradicate S. mitis cells. In the presence of human red blood (HRB) cells, two aspects were analyzed: the photohemolysis induced by these cationic phthalocyanines and the PDI of bacteria in medium containing erythrocytes. The highest photohemolytic damage was produced by ZnPPc4+, which can be avoided using azida ion as photoprotective quencher. In both bacteria, the photoinactivation is possible in presence of HRB cells. Mainly, ZnEPc4+ is effective to photoinactivate S. mitis with a low hemolysis of erythrocytes. However, inactivation of E. coli by ZnPPc4+ decreases in medium with HRB cells, further when azide ion is added to avoid hemolysis. The presence of plasma considerable reduces the photocytotoxic effect, which mainly affects the eradication of E. coli. However, the PDI of S. mitis by ZnEPc4+ is even possible in presence of blood derivatives. PMID:20153568

  4. Stromal vascular progenitors in adult human adipose tissue

    PubMed Central

    Zimmerlin, Ludovic; Donnenberg, Vera S.; Pfeifer, Melanie E.; Meyer, E. Michael; Péault, Bruno; Rubin, J. Peter; Donnenberg, Albert D.

    2014-01-01

    Background The in vivo progenitor of culture-expanded mesenchymal-like adipose-derived stem cells (ADSC) remains elusive, owing in part to the complex organization of stromal cells surrounding the small vessels, and the rapidity with which adipose stromal vascular cells adopt a mesenchymal phenotype in vitro. Methods Immunohistostaining of intact adipose tissue was used to identify 3 markers (CD31, CD34, CD146) which together unambiguously discriminate histologically distinct inner and outer rings of vessel-associated stromal cells, as well as capillary and small vessel endothelial cells. These markers were used in multiparameter flow cytometry in conjunction with stem/progenitor markers (CD90, CD117) to further characterize stromal vascular fraction (SVF) subpopulations. Two mesenchymal and two endothelial populations were isolated by high speed flow cytometric sorting, expanded in short term culture and tested for adipogenesis. Results The inner layer of stromal cells in contact with small vessel endothelium (pericytes) was CD146+/α-SMA+/CD90±/CD34−/CD31−; the outer adventitial stromal ring (designated supra adventitial-adipose stromal cells, SA-ASC) was CD146−/α-SMA−/CD90+/CD34+/CD31−. Capillary endothelial cells were CD31+/CD34+/CD90+ (endothelial progenitor), while small vessel endothelium was CD31+/CD34−/CD90− (endothelial mature). Flow cytometry confirmed these expression patterns and revealed a CD146+/CD90+/CD34+/CD31− pericyte subset that may be transitional between pericytes and SA-ASC. Pericytes had the most potent adipogenic potential, followed by the more numerous SA-ASC. Endothelial populations had significantly reduced adipogenic potential compared to unsorted expanded SVF cells. Conclusions In adipose tissue perivascular stromal cells are organized in two discrete layers, the innermost consisting of CD146+/CD34− pericytes, and the outermost of CD146−/CD34+ SA-ASC, both of which have adipogenic potential in culture. A CD146+/CD

  5. Stem cells and progenitor cells in renal disease.

    PubMed

    Haller, Hermann; de Groot, Kirsten; Bahlmann, Ferdinand; Elger, Marlies; Fliser, Danilo

    2005-11-01

    Stem cells and progenitor cells are necessary for repair and regeneration of injured renal tissue. Infiltrating or resident stem cells can contribute to the replacement of lost or damaged tissue. However, the regulation of circulating progenitor cells is not well understood. We have analyzed the effects of erythropoietin on circulating progenitor cells and found that low levels of erythropoietin induce mobilization and differentiation of endothelial progenitor cells. In an animal model of 5/6 nephrectomy we could demonstrate that erythropoietin ameliorates tissue injury. Full regeneration of renal tissue demands the existence of stem cells and an adequate local "milieu," a so-called stem cell niche. We have previously described a stem cell niche in the kidneys of the dogfish, Squalus acanthus. Further analysis revealed that in the regenerating zone of the shark kidney, stem cells exist that can be induced by loss of renal tissue to form new glomeruli. Such animal models improve our understanding of stem cell behavior in the kidney and may eventually contribute to novel therapies. PMID:16221168

  6. Secondary Sphere Formation Enhances the Functionality of Cardiac Progenitor Cells

    PubMed Central

    Cho, Hyun-Jai; Lee, Ho-Jae; Youn, Seock-Won; Koh, Seok-Jin; Won, Joo-Yun; Chung, Yeon-Ju; Cho, Hyun-Ju; Yoon, Chang-Hwan; Lee, Sae-Won; Lee, Eun Ju; Kwon, Yoo-Wook; Lee, Hae-Young; Lee, Sang Hun; Ho, Won-Kyung; Park, Young-Bae; Kim, Hyo-Soo

    2012-01-01

    Loss of cardiomyocytes impairs cardiac function after myocardial infarction (MI). Recent studies suggest that cardiac stem/progenitor cells could repair the damaged heart. However, cardiac progenitor cells are difficult to maintain in terms of purity and multipotency when propagated in two-dimensional culture systems. Here, we investigated a new strategy that enhances potency and enriches progenitor cells. We applied the repeated sphere formation strategy (cardiac explant → primary cardiosphere (CS) formation → sphere-derived cells (SDCs) in adherent culture condition → secondary CS formation by three-dimensional culture). Cells in secondary CS showed higher differentiation potentials than SDCs. When transplanted into the infarcted myocardium, secondary CSs engrafted robustly, improved left ventricular (LV) dysfunction, and reduced infarct sizes more than SDCs did. In addition to the cardiovascular differentiation of transplanted secondary CSs, robust vascular endothelial growth factor (VEGF) synthesis and secretion enhanced neovascularization in the infarcted myocardium. Microarray pathway analysis and blocking experiments using E-selectin knock-out hearts, specific chemicals, and small interfering RNAs (siRNAs) for each pathway revealed that E-selectin was indispensable to sphere initiation and ERK/Sp1/VEGF autoparacrine loop was responsible for sphere maturation. These results provide a simple strategy for enhancing cellular potency for cardiac repair. Furthermore, this strategy may be implemented to other types of stem/progenitor cell-based therapy. PMID:22713697

  7. Hmga2 regulates self-renewal of retinal progenitors

    PubMed Central

    Parameswaran, Sowmya; Xia, Xiaohuan; Hegde, Ganapati; Ahmad, Iqbal

    2014-01-01

    In vertebrate retina, histogenesis occurs over an extended period. To sustain the temporal generation of diverse cell types, retinal progenitor cells (RPCs) must self-renew. However, self-renewal and regulation of RPCs remain poorly understood. Here, we demonstrate that cell-extrinsic factors coordinate with the epigenetic regulator high-mobility group AT-hook 2 (Hmga2) to regulate self-renewal of late retinal progenitor cells (RPCs). We observed that a small subset of RPCs was capable of clonal propagation and retained multipotentiality of parents in the presence of endothelial cells (ECs), known self-renewal regulators in various stem cell niches. The self-renewing effects, also observed in vivo, involve multiple intercellular signaling pathways, engaging Hmga2. As progenitors exhaust during retinal development, expression of Hmga2 progressively decreases. Analyses of Hmga2-expression perturbation, in vitro and in vivo, revealed that Hmga2 functionally helps to mediate cell-extrinsic influences on late-retinal progenitor self-renewal. Our results provide a framework for integrating the diverse intercellular influences elicited by epigenetic regulators for self-renewal in a dynamic stem cell niche: the developing vertebrate retina. PMID:25336737

  8. JAK-STAT and AKT pathway-coupled genes in erythroid progenitor cells through ontogeny

    PubMed Central

    2012-01-01

    Background It has been reported that the phosphatidylinositol 3-kinase (PI3K)-AKT signaling pathway regulates erythropoietin (EPO)-induced survival, proliferation, and maturation of early erythroid progenitors. Erythroid cell proliferation and survival have also been related to activation of the JAK-STAT pathway. The goal of this study was to observe the function of EPO activation of JAK-STAT and PI3K/AKT pathways in the development of erythroid progenitors from hematopoietic CD34+ progenitor cells, as well as to distinguish early EPO target genes in human erythroid progenitors during ontogeny. Methods Hematopoietic CD34+ progenitor cells, isolated from fetal and adult hematopoietic tissues, were differentiated into erythroid progenitor cells. We have used microarray analysis to examine JAK-STAT and PI3K/AKT related genes, as well as broad gene expression modulation in these human erythroid progenitor cells. Results In microarray studies, a total of 1755 genes were expressed in fetal liver, 3844 in cord blood, 1770 in adult bone marrow, and 1325 genes in peripheral blood-derived erythroid progenitor cells. The erythroid progenitor cells shared 1011 common genes. Using the Ingenuity Pathways Analysis software, we evaluated the network pathways of genes linked to hematological system development, cellular growth and proliferation. The KITLG, EPO, GATA1, PIM1 and STAT3 genes represent the major connection points in the hematological system development linked genes. Some JAK-STAT signaling pathway-linked genes were steadily upregulated throughout ontogeny (PIM1, SOCS2, MYC, PTPN11), while others were downregulated (PTPN6, PIAS, SPRED2). In addition, some JAK-STAT pathway related genes are differentially expressed only in some stages of ontogeny (STATs, GRB2, CREBB). Beside the continuously upregulated (AKT1, PPP2CA, CHUK, NFKB1) and downregulated (FOXO1, PDPK1, PIK3CG) genes in the PI3K-AKT signaling pathway, we also observed intermittently regulated gene expression

  9. Transcript Analysis Reveals a Specific HOX Signature Associated with Positional Identity of Human Endothelial Cells

    PubMed Central

    Toshner, Mark; Dunmore, Benjamin J.; McKinney, Eoin F.; Southwood, Mark; Caruso, Paola; Upton, Paul D.; Waters, John P.; Ormiston, Mark L.; Skepper, Jeremy N.; Nash, Gerard; Rana, Amer A.; Morrell, Nicholas W.

    2014-01-01

    The endothelial cell has a remarkable ability for sub-specialisation, adapted to the needs of a variety of vascular beds. The role of developmental programming versus the tissue contextual environment for this specialization is not well understood. Here we describe a hierarchy of expression of HOX genes associated with endothelial cell origin and location. In initial microarray studies, differential gene expression was examined in two endothelial cell lines: blood derived outgrowth endothelial cells (BOECs) and pulmonary artery endothelial cells. This suggested shared and differential patterns of HOX gene expression between the two endothelial lines. For example, this included a cluster on chromosome 2 of HOXD1, HOXD3, HOXD4, HOXD8 and HOXD9 that was expressed at a higher level in BOECs. Quantative PCR confirmed the higher expression of these HOXs in BOECs, a pattern that was shared by a variety of microvascular endothelial cell lines. Subsequently, we analysed publically available microarrays from a variety of adult cell and tissue types using the whole “HOX transcriptome” of all 39 HOX genes. Using hierarchical clustering analysis the HOX transcriptome was able to discriminate endothelial cells from 61 diverse human cell lines of various origins. In a separate publically available microarray dataset of 53 human endothelial cell lines, the HOX transcriptome additionally organized endothelial cells related to their organ or tissue of origin. Human tissue staining for HOXD8 and HOXD9 confirmed endothelial expression and also supported increased microvascular expression of these HOXs. Together these observations suggest a significant involvement of HOX genes in endothelial cell positional identity. PMID:24651450

  10. Genetic Tagging During Human Mesoderm Differentiation Reveals Tripotent Lateral Plate Mesodermal Progenitors.

    PubMed

    Chin, Chee Jia; Cooper, Aaron R; Lill, Georgia R; Evseenko, Denis; Zhu, Yuhua; He, Chong Bin; Casero, David; Pellegrini, Matteo; Kohn, Donald B; Crooks, Gay M

    2016-05-01

    Although clonal studies of lineage potential have been extensively applied to organ specific stem and progenitor cells, much less is known about the clonal origins of lineages formed from the germ layers in early embryogenesis. We applied lentiviral tagging followed by vector integration site analysis (VISA) with high-throughput sequencing to investigate the ontogeny of the hematopoietic, endothelial and mesenchymal lineages as they emerge from human embryonic mesoderm. In contrast to studies that have used VISA to track differentiation of self-renewing stem cell clones that amplify significantly over time, we focused on a population of progenitor clones with limited self-renewal capability. Our analyses uncovered the critical influence of sampling on the interpretation of lentiviral tag sharing, particularly among complex populations with minimal clonal duplication. By applying a quantitative framework to estimate the degree of undersampling we revealed the existence of tripotent mesodermal progenitors derived from pluripotent stem cells, and the subsequent bifurcation of their differentiation into bipotent endothelial/hematopoietic or endothelial/mesenchymal progenitors. Stem Cells 2016;34:1239-1250. PMID:26934332

  11. Endothelial cells regulate neural crest and second heart field morphogenesis

    PubMed Central

    Milgrom-Hoffman, Michal; Michailovici, Inbal; Ferrara, Napoleone; Zelzer, Elazar; Tzahor, Eldad

    2014-01-01

    ABSTRACT Cardiac and craniofacial developmental programs are intricately linked during early embryogenesis, which is also reflected by a high frequency of birth defects affecting both regions. The molecular nature of the crosstalk between mesoderm and neural crest progenitors and the involvement of endothelial cells within the cardio–craniofacial field are largely unclear. Here we show in the mouse that genetic ablation of vascular endothelial growth factor receptor 2 (Flk1) in the mesoderm results in early embryonic lethality, severe deformation of the cardio–craniofacial field, lack of endothelial cells and a poorly formed vascular system. We provide evidence that endothelial cells are required for migration and survival of cranial neural crest cells and consequently for the deployment of second heart field progenitors into the cardiac outflow tract. Insights into the molecular mechanisms reveal marked reduction in Transforming growth factor beta 1 (Tgfb1) along with changes in the extracellular matrix (ECM) composition. Our collective findings in both mouse and avian models suggest that endothelial cells coordinate cardio–craniofacial morphogenesis, in part via a conserved signaling circuit regulating ECM remodeling by Tgfb1. PMID:24996922

  12. Circulating endothelial cells in cardiovascular disease.

    PubMed

    Boos, Christopher J; Lip, Gregory Y H; Blann, Andrew D

    2006-10-17

    Quantification of circulating endothelial cells (CECs) in peripheral blood is developing as a novel and reproducible method of assessing endothelial damage/dysfunction. The CECs are thought to be mature cells that have detached from the intimal monolayer in response to endothelial injury and are a different cell population to endothelial progenitor cells (EPCs). The EPCs are nonleukocytes derived from the bone marrow that are believed to have proliferative potential and may be important in vascular regeneration. Currently accepted methods of CEC quantification include the use of immunomagnetic bead separation (with cell counting under fluorescence microscopy) and flow cytometry. Several recent studies have shown increased numbers of CECs in cardiovascular disease and its risk factors, such as unstable angina, acute myocardial infarction, stroke, diabetes mellitus, and critical limb ischemia, but no change in stable intermittent claudication, essential hypertension, or atrial fibrillation. Furthermore, CEC quantification at 48 h after acute myocardial infarction has been shown to be an accurate predictor of major adverse coronary events and death at both 1 month and 1 year. This article presents an overview of the pathophysiology of CECs in the setting of cardiovascular disease and a brief comparison with EPCs. PMID:17045885

  13. Functional Kidney Bioengineering with Pluripotent Stem-Cell-Derived Renal Progenitor Cells and Decellularized Kidney Scaffolds.

    PubMed

    Du, Chan; Narayanan, Karthikeyan; Leong, Meng Fatt; Ibrahim, Mohammed Shahrudin; Chua, Ying Ping; Khoo, Vanessa Mei Hui; Wan, Andrew C A

    2016-08-01

    Recent advances in developmental biology and stem cell technology have led to the engineering of functional organs in a dish. However, the limited size of these organoids and absence of a large circulatory system poses limits to its clinical translation. To overcome these issues, decellularized whole kidney scaffolds with native microstructure and extracellular matrix (ECM) are employed for kidney bioengineering, using human-induced pluripotent-stem-cell-derived renal progenitor cells and endothelial cells. To demonstrate ECM-guided cellular assembly, the present work is focused on generating the functional unit of the kidney, the glomerulus. In the repopulated organ, the presence of endothelial cells broadly upregulates the expression level of genes related to renal development. When the cellularized native scaffolds are implanted in SCID mice, glomeruli assembly can be achieved by co-culture of the renal progenitors and endothelial cells. These individual glomerular units are shown to be functional in the context of the whole organ using a simulated bio-reactor set-up with urea and creatinine excretion and albumin reabsorption. Our results indicate that the repopulation of decellularized native kidney using clinically relevant, expandable patient-specific renal progenitors and endothelial cells may be a viable approach for the generation of a functional whole kidney. PMID:27294565

  14. Eotaxin-Rich Proangiogenic Hematopoietic Progenitor Cells and CCR3+ Endothelium in the Atopic Asthmatic Response.

    PubMed

    Asosingh, Kewal; Vasanji, Amit; Tipton, Aaron; Queisser, Kimberly; Wanner, Nicholas; Janocha, Allison; Grandon, Deepa; Anand-Apte, Bela; Rothenberg, Marc E; Dweik, Raed; Erzurum, Serpil C

    2016-03-01

    Angiogenesis is closely linked to and precedes eosinophilic infiltration in asthma. Eosinophils are recruited into the airway by chemoattractant eotaxins, which are expressed by endothelial cells, smooth muscles cells, epithelial cells, and hematopoietic cells. We hypothesized that bone marrow-derived proangiogenic progenitor cells that contain eotaxins contribute to the initiation of angiogenesis and inflammation in asthma. Whole-lung allergen challenge of atopic asthma patients revealed vascular activation occurs within hours of challenge and before airway inflammation. The eotaxin receptor CCR3 was expressed at high levels on submucosal endothelial cells in patients and a murine model of asthma. Ex vivo exposure of murine endothelial cells to eotaxins induced migration and angiogenesis. In mechanistic studies, wild-type mice transplanted with eotaxin-1/2-deficient bone marrow had markedly less angiogenesis and inflammation in an atopic asthma model, whereas adoptive transfer of proangiogenic progenitor cells from wild-type mice in an atopic asthma model into the eotaxin-1/2-deficient mice led to angiogenesis and airway inflammation. The findings indicate that Th2-promoting hematopoietic progenitor cells are rapidly recruited to the lung upon allergen exposure and release eotaxins that coordinately activate endothelial cells, angiogenesis, and airway inflammation. PMID:26810221

  15. Endothelin-1 supports clonal derivation and expansion of cardiovascular progenitors derived from human embryonic stem cells.

    PubMed

    Soh, Boon-Seng; Ng, Shi-Yan; Wu, Hao; Buac, Kristina; Park, Joo-Hye C; Lian, Xiaojun; Xu, Jiejia; Foo, Kylie S; Felldin, Ulrika; He, Xiaobing; Nichane, Massimo; Yang, Henry; Bu, Lei; Li, Ronald A; Lim, Bing; Chien, Kenneth R

    2016-01-01

    Coronary arteriogenesis is a central step in cardiogenesis, requiring coordinated generation and integration of endothelial cell and vascular smooth muscle cells. At present, it is unclear whether the cell fate programme of cardiac progenitors to generate complex muscular or vascular structures is entirely cell autonomous. Here we demonstrate the intrinsic ability of vascular progenitors to develop and self-organize into cardiac tissues by clonally isolating and expanding second heart field cardiovascular progenitors using WNT3A and endothelin-1 (EDN1) human recombinant proteins. Progenitor clones undergo long-term expansion and differentiate primarily into endothelial and smooth muscle cell lineages in vitro, and contribute extensively to coronary-like vessels in vivo, forming a functional human-mouse chimeric circulatory system. Our study identifies EDN1 as a key factor towards the generation and clonal derivation of ISL1(+) vascular intermediates, and demonstrates the intrinsic cell-autonomous nature of these progenitors to differentiate and self-organize into functional vasculatures in vivo. PMID:26952167

  16. Endothelin-1 supports clonal derivation and expansion of cardiovascular progenitors derived from human embryonic stem cells

    PubMed Central

    Soh, Boon-Seng; Ng, Shi-Yan; Wu, Hao; Buac, Kristina; Park, Joo-Hye C.; Lian, Xiaojun; Xu, Jiejia; Foo, Kylie S.; Felldin, Ulrika; He, Xiaobing; Nichane, Massimo; Yang, Henry; Bu, Lei; Li, Ronald A.; Lim, Bing; Chien, Kenneth R.

    2016-01-01

    Coronary arteriogenesis is a central step in cardiogenesis, requiring coordinated generation and integration of endothelial cell and vascular smooth muscle cells. At present, it is unclear whether the cell fate programme of cardiac progenitors to generate complex muscular or vascular structures is entirely cell autonomous. Here we demonstrate the intrinsic ability of vascular progenitors to develop and self-organize into cardiac tissues by clonally isolating and expanding second heart field cardiovascular progenitors using WNT3A and endothelin-1 (EDN1) human recombinant proteins. Progenitor clones undergo long-term expansion and differentiate primarily into endothelial and smooth muscle cell lineages in vitro, and contribute extensively to coronary-like vessels in vivo, forming a functional human–mouse chimeric circulatory system. Our study identifies EDN1 as a key factor towards the generation and clonal derivation of ISL1+ vascular intermediates, and demonstrates the intrinsic cell-autonomous nature of these progenitors to differentiate and self-organize into functional vasculatures in vivo. PMID:26952167

  17. TNFα and Endothelial Cells Modulate Notch Signaling in the Bone Marrow Microenvironment during Inflammation

    PubMed Central

    Fernandez, Luis; Rodriguez, Sonia; Huang, Hui; Chora, Angelo; Mumaw, Christin; Cruz, Eugenia; Pollok, Karen; Cristina, Filipa; Price, Joanne E.; Ferkowicz, Michael J.; Scadden, David T.; Clauss, Matthias; Cardoso, Angelo A.; Carlesso, Nadia

    2009-01-01

    Objective Homeostasis of the hematopoietic compartment is challenged and maintained during conditions of stress by mechanisms that are poorly defined. To understand how the bone marrow (BM) microenvironment influences hematopoiesis, we explored the role of Notch signaling and bone marrow endothelial cells in providing microenvironmental cues to hematopoietic cells in the presence of inflammatory stimuli. Methods The human BM endothelial cell line BMEC and primary human BM endothelial cells were analyzed for expression of Notch ligands and the ability to expand hematopoietic progenitors in an in vitro co-culture system. In vivo experiments were carried out to identify modulation of Notch signaling in BM endothelial and hematopoietic cells in mice challenged with TNFα or LPS, or in Tie2-tmTNFα transgenic mice characterized by constitutive TNFα activation. Results BM endothelial cells were found to express Jagged ligands and to greatly support progenitor’s colony-forming ability. This effect was markedly decreased by Notch antagonists and augmented by increasing levels of Jagged2. Physiologic upregulation of Jagged2 expression on BMEC was observed upon TNFα activation. Injection of TNFα or LPS upregulated 3 to 4 fold Jagged2 expression on murine BM endothelial cells in vivo and resulted in increased Notch activation on murine hematopoietic stem/progenitor cells. Similarly, constitutive activation of endothelial cells in Tie2-tmTNFα mice was characterized by increased expression of Jagged2 and by augmented Notch activation on hematopoietic stem/progenitor cells. Conclusions Our results provide the first evidence that BM endothelial cells promote expansion of hematopoietic progenitor cells by a Notch-dependent mechanism and that TNFα and LPS can modulate the levels of Notch ligand expression and Notch activation in the bone marrow microenvironment in vivo. PMID:18439488

  18. 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

  19. Human Umbilical Cord Blood-Derived Serum for Culturing the Supportive Feeder Cells of Human Pluripotent Stem Cell Lines

    PubMed Central

    Rungsiwiwut, Ruttachuk; Ingrungruanglert, Praewphan; Numchaisrika, Pranee; Virutamasen, Pramuan; Phermthai, Tatsanee; Pruksananonda, Kamthorn

    2016-01-01

    Although human pluripotent stem cells (hPSCs) can proliferate robustly on the feeder-free culture system, genetic instability of hPSCs has been reported in such environment. Alternatively, feeder cells enable hPSCs to maintain their pluripotency. The feeder cells are usually grown in a culture medium containing fetal bovine serum (FBS) prior to coculture with hPSCs. The use of FBS might limit the clinical application of hPSCs. Recently, human cord blood-derived serum (hUCS) showed a positive effect on culture of mesenchymal stem cells. It is interesting to test whether hUCS can be used for culture of feeder cells of hPSCs. This study was aimed to replace FBS with hUCS for culturing the human foreskin fibroblasts (HFFs) prior to feeder cell preparation. The results showed that HFFs cultured in hUCS-containing medium (HFF-hUCS) displayed fibroblastic features, high proliferation rates, short population doubling times, and normal karyotypes after prolonged culture. Inactivated HFF-hUCS expressed important genes, including Activin A, FGF2, and TGFβ1, which have been implicated in the maintenance of hPSC pluripotency. Moreover, hPSC lines maintained pluripotency, differentiation capacities, and karyotypic stability after being cocultured for extended period with inactivated HFF-hUCS. Therefore, the results demonstrated the benefit of hUCS for hPSCs culture system. PMID:26839561

  20. Conditioned Media from Human Umbilical Cord Blood-Derived Mesenchymal Stem Cells Inhibits Melanogenesis by Promoting Proteasomal Degradation of MITF

    PubMed Central

    Lim, Hoon; Shin, Ji Hyun; Park, So Jung; Jo, Yoon Kyung; Oh, Wonil; Yang, Yoon Sun; Cho, Dong-Hyung; Kim, Ju-Yeon

    2015-01-01

    Human umbilical cord blood-derived mesenchymal stem cells (hUCB-MSCs) secrete various beneficial molecules, which have anti-apoptotic activity and cell proliferation. However, the effect of hUCB-MSCs in melanogenesis is largely unclear. In this study, we show that conditioned media (CM) derived from hUCB-MSCs inhibit melanogenesis by regulating microphthalmia-associated transcription factor (MITF) expression via the ERK signalling pathway. Treatment of hUCB-MSC-CM strongly inhibited the alpha-melanocyte stimulating hormone-induced hyperpigmentation in melanoma cells as well as melanocytes. Treatment of hUCB-MSC-CM induced ERK1/2 activation in melanocytes. In addition, inhibition of ERK1/2 suppressed the anti-pigmentation activity of the hUCB-MSC-CM in melanocytes and in vitro artificial skin models. We also found that the expression of MITF was appreciably diminished while expression of phosphorylated MITF, which leads to its proteasomal degradation, was increased in cells treated with hUCB-MSC-CM. These results suggested that hUCB-MSC-CM significantly suppresses melanin synthesis via MITF degradation by the ERK pathway activation. PMID:26024475

  1. Poly(ethylene glycol)-containing hydrogels promote the release of primary granules from human blood-derived polymorphonuclear leukocytes

    PubMed Central

    Cohen, Hannah Caitlin; Lieberthal, Tyler Jacob; Kao, W. John

    2014-01-01

    Polymorphonuclear leukocytes (PMNs) are recruited to sites of injury and biomaterial implants. Once activated, PMNs can exocytose their granule subsets to recruit monocytes (MCs) and mediate MC/macrophage activation. We investigated the release of myeloperoxidase (MPO), a primary granule marker, and matrix metalloproteinase-9 (MMP-9), a tertiary granule marker, from human blood-derived PMNs cultured on poly(ethylene glycol) (PEG) hydrogels, polydimethylsiloxane (PDMS), tissue culture polystyrene (TCPS) and gelatin-PEG (GP) hydrogels, with and without the presence of the bacterial peptide formyl-Met-Leu-Phe. Supernatants from PMN cultures on PEG-containing hydrogels (i.e., PEG and GP hydrogels) had higher concentrations of MPO than those from PMN cultures on PDMS or TCPS at 2 hours. PMNs on all biomaterials released comparable levels of MMP-9 at 2 hours, indicating that PMNs cultured on PEG-containing hydrogels have different mechanisms of release for primary and tertiary granules. Src family kinases were involved in the release of MPO from PMNs cultured on PEG hydrogels, TCPS and GP hydrogels and in the release of MMP-9 from PMNs cultured on all four materials. The increased release of primary granules from PMNs on PEG-containing hydrogels did not significantly increase MC chemotaxis, indicating that additional co-effectors in the dynamic inflammatory milieu in vivo modulate PMN-mediated MC recruitment. PMID:24497370

  2. Preclinical Study of Cell Therapy for Osteonecrosis of the Femoral Head with Allogenic Peripheral Blood-Derived Mesenchymal Stem Cells

    PubMed Central

    Fu, Qiang; Tang, Ning-Ning; Zhang, Qian; Liu, Yi; Peng, Jia-Chen; Fang, Ning; Yu, Li-Mei; Liu, Jin-Wei

    2016-01-01

    Purpose To explore the value of transplanting peripheral blood-derived mesenchymal stem cells from allogenic rabbits (rPBMSCs) to treat osteonecrosis of the femoral head (ONFH). Materials and Methods rPBMSCs were separated/cultured from peripheral blood after granulocyte colony-stimulating factor mobilization. Afterwards, mobilized rPBMSCs from a second passage labeled with PKH26 were transplanted into rabbit ONFH models, which were established by liquid nitrogen freezing, to observe the effect of rPBMSCs on ONFH repair. Then, the mRNA expressions of BMP-2 and PPAR-γ in the femoral head were assessed by RT-PCR. Results After mobilization, the cultured rPBMSCs expressed mesenchymal markers of CD90, CD44, CD29, and CD105, but failed to express CD45, CD14, and CD34. The colony forming efficiency of mobilized rPBMSCs ranged from 2.8 to 10.8 per million peripheral mononuclear cells. After local transplantation, survival of the engrafted cells reached at least 8 weeks. Therein, BMP-2 was up-regulated, while PPAR-γ mRNA was down-regulated. Additionally, bone density and bone trabeculae tended to increase gradually. Conclusion We confirmed that local transplantation of rPBMSCs benefits ONFH treatment and that the beneficial effects are related to the up-regulation of BMP-2 expression and the down-regulation of PPAR-γ expression. PMID:27189298

  3. Early Umbilical Cord Blood-Derived Stem Cell Transplantation Does Not Prevent Neurological Deterioration in Mucopolysaccharidosis Type III.

    PubMed

    Welling, Lindsey; Marchal, Jan Pieter; van Hasselt, Peter; van der Ploeg, Ans T; Wijburg, Frits A; Boelens, Jaap Jan

    2015-01-01

    Mucopolysaccharidosis type III (MPS III), or Sanfilippo disease, is a neurodegenerative lysosomal storage disease (LSD) caused by defective lysosomal degradation of heparan sulfate (HS). No effective disease-modifying therapy is yet available. In contrast to some other neuronopathic LSDs, bone marrow-derived hematopoietic stem cell transplantation (HSCT) fails to prevent neurological deterioration in MPS III patients. We report on the 5-year outcome of early transplantation, i.e., before onset of clinical neurological disease, in combination with the use of umbilical cord blood-derived hematopoietic stem cells (UCBT), in two MPS III patients. Both patients had a normal developmental quotient at the time of UCBT. One patient had a combination of mutations predicting a classical severe phenotype (MPS IIIA), and one patient (MPS IIIB) had mutations predicting a very attenuated phenotype. Transplantation was uncomplicated with full engraftment of donor cells in both.Both patients showed progressive neurological deterioration with regression of cognitive skills and behavioral disturbances during 5 years after successful UCBT, comparable to the natural history of patients with the same combination of mutations. The concentration of HS in CSF in the patient with the attenuated phenotype of MPS IIIB 2 years after UCBT was very high and in the range of untreated MPS III patients.We conclude that the course of cognitive development, behavioral problems, and absence of biochemical correction in CSF demonstrate the absence of relevant effect of UCBT in MPS III patients, even when performed before clinical onset of CNS disease. PMID:25256447

  4. Effects of donors’ age and passage number on the biological characteristics of menstrual blood-derived stem cells

    PubMed Central

    Chen, Jinyang; Du, Xiaochun; Chen, Qian; Xiang, Charlie

    2015-01-01

    We investigated the effects of donor age and passage number on the biological characteristics of menstrual blood-derived stem cells (MenSCs) by comparing MenSCs derived from donors with three different age ranges and after different passage times. Continuous passage, flat cloning, cell proliferation assays, flow cytometric phenotyping and whole human genome microarray were performed to systematically analyze the relationship between the self-renewal ability of MenSCs as well as their potential to maintain their stem cell characteristics and to resist aging. The results demonstrated that the immunophenotypes and in vitro cultural characteristics of MenSCs did not change significantly with the progression of aging. However, some important signal pathways including MAPK, the insulin signaling pathway, pathways involved in carcinogenesis such as PPAR and P53, and cytokines and their receptors, as well as other pathways associated with immune response and aging, changed to various extents under the conditions of aging after a long time in vitro. The enriched differentially-expressed genes were mainly involved in transcriptional regulation, stress response, cell proliferation, development and apoptosis. The key differentiallyexpressed genes associated with age and passage number were identified for use as biomarkers of cell aging. PMID:26823782

  5. Functional Blood Progenitor Markers in Developing Human Liver Progenitors.

    PubMed

    Goldman, Orit; Cohen, Idan; Gouon-Evans, Valerie

    2016-08-01

    In the early fetal liver, hematopoietic progenitors expand and mature together with hepatoblasts, the liver progenitors of hepatocytes and cholangiocytes. Previous analyses of human fetal livers indicated that both progenitors support each other's lineage maturation and curiously share some cell surface markers including CD34 and CD133. Using the human embryonic stem cell (hESC) system, we demonstrate that virtually all hESC-derived hepatoblast-like cells (Hep cells) transition through a progenitor stage expressing CD34 and CD133 as well as GATA2, an additional hematopoietic marker that has not previously been associated with human hepatoblast development. Dynamic expression patterns for CD34, CD133, and GATA2 in hepatoblasts were validated in human fetal livers collected from the first and second trimesters of gestation. Knockdown experiments demonstrate that each gene also functions to regulate hepatic fate mostly in a cell-autonomous fashion, revealing unprecedented roles of fetal hematopoietic progenitor markers in human liver progenitors. PMID:27509132

  6. Progenitors of type Ia supernovae

    NASA Astrophysics Data System (ADS)

    Maeda, Keiichi; Terada, Yukikatsu

    2016-07-01

    Natures of progenitors of type Ia Supernovae (SNe Ia) have not yet been clarified. There has been long and intensive discussion on whether the so-called single degenerate (SD) scenario or the double degenerate (DD) scenario, or anything else, could explain a major population of SNe Ia, but the conclusion has not yet been reached. With rapidly increasing observational data and new theoretical ideas, the field of studying the SN Ia progenitors has been quickly developing, and various new insights have been obtained in recent years. This paper aims at providing a summary of the current situation regarding the SN Ia progenitors, both in theory and observations. It seems difficult to explain the emerging diversity seen in observations of SNe Ia by a single population, and we emphasize that it is important to clarify links between different progenitor scenarios and different sub-classes of SNe Ia.

  7. Mesenchymal Stem Cell-Derived Microvesicles Support Ex Vivo Expansion of Cord Blood-Derived CD34(+) Cells.

    PubMed

    Xie, Hui; Sun, Li; Zhang, Liming; Liu, Teng; Chen, Li; Zhao, Aiqi; Lei, Qian; Gao, Fei; Zou, Ping; Li, Qiubai; Guo, An-Yuan; Chen, Zhichao; Wang, Hongxiang

    2016-01-01

    Mesenchymal stem cells (MSCs) are known to support the characteristic properties of hematopoietic stem and progenitor cells (HSPCs) in the bone marrow hematopoietic microenvironment. MSCs are used in coculture systems as a feeder layer for the ex vivo expansion of umbilical cord blood (CB) to increase the relatively low number of HSPCs in CB. Findings increasingly suggest that MSC-derived microvesicles (MSC-MVs) play an important role in the biological functions of their parent cells. We speculate that MSC-MVs may recapitulate the hematopoiesis-supporting effects of their parent cells. In the current study, we found MSC-MVs containing microRNAs that are involved in the regulation of hematopoiesis. We also demonstrated that MSC-MVs could improve the expansion of CB-derived mononuclear cells and CD34(+) cells and generate a greater number of primitive progenitor cells in vitro. Additionally, when MSC-MVs were added to the CB-MSC coculture system, they could improve the hematopoiesis-supporting effects of MSCs. These findings highlight the role of MSC-MVs in the ex vivo expansion of CB, which may offer a promising therapeutic approach in CB transplantation. PMID:27042183

  8. Mesenchymal Stem Cell-Derived Microvesicles Support Ex Vivo Expansion of Cord Blood-Derived CD34+ Cells

    PubMed Central

    Xie, Hui; Sun, Li; Zhang, Liming; Liu, Teng; Chen, Li; Zhao, Aiqi; Lei, Qian; Gao, Fei; Zou, Ping; Li, Qiubai; Guo, An-yuan; Chen, Zhichao; Wang, Hongxiang

    2016-01-01

    Mesenchymal stem cells (MSCs) are known to support the characteristic properties of hematopoietic stem and progenitor cells (HSPCs) in the bone marrow hematopoietic microenvironment. MSCs are used in coculture systems as a feeder layer for the ex vivo expansion of umbilical cord blood (CB) to increase the relatively low number of HSPCs in CB. Findings increasingly suggest that MSC-derived microvesicles (MSC-MVs) play an important role in the biological functions of their parent cells. We speculate that MSC-MVs may recapitulate the hematopoiesis-supporting effects of their parent cells. In the current study, we found MSC-MVs containing microRNAs that are involved in the regulation of hematopoiesis. We also demonstrated that MSC-MVs could improve the expansion of CB-derived mononuclear cells and CD34+ cells and generate a greater number of primitive progenitor cells in vitro. Additionally, when MSC-MVs were added to the CB-MSC coculture system, they could improve the hematopoiesis-supporting effects of MSCs. These findings highlight the role of MSC-MVs in the ex vivo expansion of CB, which may offer a promising therapeutic approach in CB transplantation. PMID:27042183

  9. Mesp1 Marked Cardiac Progenitor Cells Repair Infarcted Mouse Hearts

    PubMed Central

    Liu, Yu; Chen, Li; Diaz, Andrea Diaz; Benham, Ashley; Xu, Xueping; Wijaya, Cori S.; Fa’ak, Faisal; Luo, Weijia; Soibam, Benjamin; Azares, Alon; Yu, Wei; Lyu, Qiongying; Stewart, M. David; Gunaratne, Preethi; Cooney, Austin; McConnell, Bradley K.; Schwartz, Robert J.

    2016-01-01

    Mesp1 directs multipotential cardiovascular cell fates, even though it’s transiently induced prior to the appearance of the cardiac progenitor program. Tracing Mesp1-expressing cells and their progeny allows isolation and characterization of the earliest cardiovascular progenitor cells. Studying the biology of Mesp1-CPCs in cell culture and ischemic disease models is an important initial step toward using them for heart disease treatment. Because of Mesp1’s transitory nature, Mesp1-CPC lineages were traced by following EYFP expression in murine Mesp1Cre/+; Rosa26EYFP/+ ES cells. We captured EYFP+ cells that strongly expressed cardiac mesoderm markers and cardiac transcription factors, but not pluripotent or nascent mesoderm markers. BMP2/4 treatment led to the expansion of EYFP+ cells, while Wnt3a and Activin were marginally effective. BMP2/4 exposure readily led EYFP+ cells to endothelial and smooth muscle cells, but inhibition of the canonical Wnt signaling was required to enter the cardiomyocyte fate. Injected mouse pre-contractile Mesp1-EYFP+ CPCs improved the survivability of injured mice and restored the functional performance of infarcted hearts for at least 3 months. Mesp1-EYFP+ cells are bona fide CPCs and they integrated well in infarcted hearts and emerged de novo into terminally differentiated cardiac myocytes, smooth muscle and vascular endothelial cells. PMID:27538477

  10. Mesp1 Marked Cardiac Progenitor Cells Repair Infarcted Mouse Hearts.

    PubMed

    Liu, Yu; Chen, Li; Diaz, Andrea Diaz; Benham, Ashley; Xu, Xueping; Wijaya, Cori S; Fa'ak, Faisal; Luo, Weijia; Soibam, Benjamin; Azares, Alon; Yu, Wei; Lyu, Qiongying; Stewart, M David; Gunaratne, Preethi; Cooney, Austin; McConnell, Bradley K; Schwartz, Robert J

    2016-01-01

    Mesp1 directs multipotential cardiovascular cell fates, even though it's transiently induced prior to the appearance of the cardiac progenitor program. Tracing Mesp1-expressing cells and their progeny allows isolation and characterization of the earliest cardiovascular progenitor cells. Studying the biology of Mesp1-CPCs in cell culture and ischemic disease models is an important initial step toward using them for heart disease treatment. Because of Mesp1's transitory nature, Mesp1-CPC lineages were traced by following EYFP expression in murine Mesp1(Cre/+); Rosa26(EYFP/+) ES cells. We captured EYFP+ cells that strongly expressed cardiac mesoderm markers and cardiac transcription factors, but not pluripotent or nascent mesoderm markers. BMP2/4 treatment led to the expansion of EYFP+ cells, while Wnt3a and Activin were marginally effective. BMP2/4 exposure readily led EYFP+ cells to endothelial and smooth muscle cells, but inhibition of the canonical Wnt signaling was required to enter the cardiomyocyte fate. Injected mouse pre-contractile Mesp1-EYFP+ CPCs improved the survivability of injured mice and restored the functional performance of infarcted hearts for at least 3 months. Mesp1-EYFP+ cells are bona fide CPCs and they integrated well in infarcted hearts and emerged de novo into terminally differentiated cardiac myocytes, smooth muscle and vascular endothelial cells. PMID:27538477

  11. Low immunogenicity of allogeneic human umbilical cord blood-derived mesenchymal stem cells in vitro and in vivo

    SciTech Connect

    Lee, Miyoung; Jeong, Sang Young; Ha, Jueun; Kim, Miyeon; Jin, Hye Jin; Kwon, Soon-Jae; Chang, Jong Wook; Choi, Soo Jin; Oh, Wonil; Yang, Yoon Sun; Kim, Jae-Sung; Jeon, Hong Bae

    2014-04-18

    Highlights: • hUCB-MSCs maintained low immunogenicity even after immune challenge in vitro. • Humanized NSG mice were established using human UCB CD34+ cells. • Repeated intravenous hUCB-MSC injection into mice did not lead to immune responses and adverse events. • Allogeneic hUCB-MSCs maintained low immunogenicity in vitro and in vivo. - Abstract: Evaluation of the immunogenicity of human mesenchymal stem cells (MSCs) in an allogeneic setting during therapy has been hampered by lack of suitable models due to technical and ethical limitations. Here, we show that allogeneic human umbilical cord blood derived-MSCs (hUCB-MSCs) maintained low immunogenicity even after immune challenge in vitro. To confirm these properties in vivo, a humanized mouse model was established by injecting isolated hUCB-derived CD34+ cells intravenously into immunocompromised NOD/SCID IL2γnull (NSG) mice. After repeated intravenous injection of human peripheral blood mononuclear cells (hPBMCs) or MRC5 cells into these mice, immunological alterations including T cell proliferation and increased IFN-γ, TNF-α, and human IgG levels, were observed. In contrast, hUCB-MSC injection did not elicit these responses. While lymphocyte infiltration in the lung and small intestine and reduced survival rates were observed after hPBMC or MRC5 transplantation, no adverse events were observed following hUCB-MSC introduction. In conclusion, our data suggest that allogeneic hUCB-MSCs have low immunogenicity in vitro and in vivo, and are therefore “immunologically safe” for use in allogeneic clinical applications.

  12. Improving the neuronal differentiation efficiency of umbilical cord blood-derived mesenchymal stem cells cultivated under appropriate conditions

    PubMed Central

    Rafieemehr, Hassan; Kheirandish, Maryam; Soleimani, Masoud

    2015-01-01

    Objective(s): Umbilical cord blood-derived mesenchymal stromal cells (UCB-MSCs) are ideally suited for use in various cell-based therapies. We investigated a novel induction protocol (NIP) to improve the neuronal differentiation of human UCB-MSCs under appropriate conditions. Materials and Methods: This experimental study was performed in Iranian Blood Transfusion Organization (IBTO), Tehran, Iran. UCB-MSCs were cultured in DMEM medium supplemented with 10% FBS in a humidified incubator in equilibration with 5% CO2 at 37°C. For neuronal differentiation of UCB-MSCs, DMEM was removed and replaced with pre-induction medium containing RA, bFGF, EGF, and basal medium for two days. Then, NGF, IBMX, AsA, and Neurobasal medium were used for six days for this purpose. Real-time PCR was performed to analyze the neuronal differentiation of UCB-MSCs for the first time in Iran. Results: We found that the maximum and minimum levels of gene expression were related to GFAP and nestin, respectively. In addition, our study showed that compared to other neuronal inducers, RA might play the main role in neuronal differentiation and fate of MSCs compared to other neuronal inducers. Conclusion: Our data showed that the combination of chemical (RA, IBMX, AsA) and growth factors (NGF, EGF, bFGF) in NIP may improve the efficiency of neuronal differentiation of UCB-MSCs and may provide a new method for easy and quick application of UCB-MSCs in regenerative medicine in the future. However, the functionality of neuron-like cells must be carefully assessed in animal experiments prior to use in clinical applications. PMID:26949497

  13. Therapeutic Effects of Umbilical Cord Blood Derived Mesenchymal Stem Cell-Conditioned Medium on Pulmonary Arterial Hypertension in Rats

    PubMed Central

    Lee, Jae Chul; Cha, Choong Ik; Kim, Dong-Sik; Choe, Soo Young

    2015-01-01

    Background: Human umbilical cord blood-derived mesenchymal stem cells (hUCB-MSCs) may have multiple therapeutic applications for cell based therapy including the treatment of pulmonary artery hypertension (PAH). As low survival rates and potential tumorigenicity of implanted cells could undermine the mesenchymal stem cell (MSC) cell-based therapy, we chose to investigate the use of conditioned medium (CM) from a culture of MSC cells as a feasible alternative. Methods: CM was prepared by culturing hUCB-MSCs in three-dimensional spheroids. In a rat model of PAH induced by monocrotaline, we infused CM or the control unconditioned culture media via the tail-vein of 6-week-old Sprague-Dawley rats. Results: Compared with the control unconditioned media, CM infusion reduced the ventricular pressure, the right ventricle/(left ventricle+interventricular septum) ratio, and maintained respiratory function in the treated animals. Also, the number of interleukin 1α (IL-1α), chemokine (C-C motif) ligand 5 (CCL5), and tissue inhibitor of metalloproteinase 1 (TIMP-1)–positive cells increased in lung samples and the number of terminal deoxynucleotidyl transferase–mediated deoxyuridine triphosphate nick-end labeling technique (TUNEL)–positive cells decreased significantly in the CM treated animals. Conclusions: From our in vivo data in the rat model, the observed decreases in the TUNEL staining suggest a potential therapeutic benefit of the CM in ameliorating PAH-mediated lung tissue damage. Increased IL-1α, CCL5, and TIMP-1 levels may play important roles in this regard. PMID:26471341

  14. Transmissible cytotoxicity of multiple myeloma cells by cord blood-derived NK cells is mediated by vesicle trafficking

    PubMed Central

    Martin-Antonio, B; Najjar, A; Robinson, S N; Chew, C; Li, S; Yvon, E; Thomas, M W; Mc Niece, I; Orlowski, R; Muñoz-Pinedo, C; Bueno, C; Menendez, P; Fernández de Larrea, C; Urbano-Ispizua, A; Shpall, E J; Shah, N

    2015-01-01

    Natural killer cells (NK) are important effectors of anti-tumor immunity, activated either by the downregulation of HLA-I molecules on tumor cells and/or the interaction of NK-activating receptors with ligands that are overexpressed on target cells upon tumor transformation (including NKG2D and NKP30). NK kill target cells by the vesicular delivery of cytolytic molecules such as Granzyme-B and Granulysin activating different cell death pathways, which can be Caspase-3 dependent or Caspase-3 independent. Multiple myeloma (MM) remains an incurable neoplastic plasma-cell disorder. However, we previously reported the encouraging observation that cord blood-derived NK (CB-NK), a new source of NK, showed anti-tumor activity in an in vivo murine model of MM and confirmed a correlation between high levels of NKG2D expression by MM cells and increased efficacy of CB-NK in reducing tumor burden. We aimed to characterize the mechanism of CB-NK-mediated cytotoxicity against MM cells. We show a Caspase-3- and Granzyme-B-independent cell death, and we reveal a mechanism of transmissible cell death between cells, which involves lipid–protein vesicle transfer from CB-NK to MM cells. These vesicles are secondarily transferred from recipient MM cells to neighboring MM cells amplifying the initial CB-NK cytotoxicity achieved. This indirect cytotoxicity involves the transfer of NKG2D and NKP30 and leads to lysosomal cell death and decreased levels of reactive oxygen species in MM cells. These findings suggest a novel and unique mechanism of CB-NK cytotoxicity against MM cells and highlight the importance of lipids and lipid transfer in this process. Further, these data provide a rationale for the development of CB-NK-based cellular therapies in the treatment of MM. PMID:25168239

  15. The Potential Utility of Blood-Derived Biochemical Markers as Indicators of Early Clinical Trends Following Severe Traumatic Brain Injury

    PubMed Central

    DeFazio, Michael; Rammo, Richard; Robles, Jaime R.; Bramlett, Helen M.; Dietrich, W. Dalton; Bullock, M. Ross

    2016-01-01

    OBJECTIVE Severe traumatic brain injury (TBI) is a dynamic neuropathologic process in which a substantial proportion of patients die within the first 48-hours. The assessment of injury severity and prognosis are of primary concern in the initial management of severe TBI. Supplemental testing that aids in the stratification of patients at high risk for deterioration may significantly improve posttraumatic management in the acute setting. METHODS This retrospective study assessed the utility of both single-marker and multimarker models as predictive indicators of acute clinical status after severe TBI. Forty-four patients who sustained severe TBI (admission Glasgow Coma Scale [GCS] score ≤8) were divided into two cohorts according to a dichotomized clinical outcome at 72 hours after admission: Poor status (death or GCS score ≤8) and improved status (GCS score improved to >8). Threshold values for clinical status prediction were calculated for serum S-100B, matrix metalloproteinase-9, and plasma D-dimer, upon admission and at 24 hours after TBI by the use of receiver operating characteristic analysis. Performance characteristics of these single-marker predictors were compared with those derived from a multimarker logistic regression analysis. RESULTS Biomarkers with the greatest predictive value for poor status at 72 hours included serum S-100B on admission, as well as plasma D-dimer and serum S-100B at 24 hours, for which, associations were strongly significant. Multimarker analysis indicated no substantial improvement in prediction accuracy over the best single predictors during this time frame. CONCLUSION In conjunction with other clinical, physical, and radiologic evidence, blood-derived biochemical markers may serve to enhance prediction of early clinical trends after severe TBI. PMID:23313262

  16. The Potential of Menstrual Blood-Derived Stem Cells in Differentiation to Epidermal Lineage: A Preliminary Report

    PubMed Central

    Faramarzi, Hossein; Mehrabani, Davood; Fard, Maryam; Akhavan, Maryam; Zare, Sona; Bakhshalizadeh, Shabnam; Manafi, Amir; Kazemnejad, Somaieh; Shirazi, Reza

    2016-01-01

    BACKGROUND Menstrual blood-derived stem cells (MenSCs) are a novel source of stem cells that can be easily isolated non-invasively from female volunteered donor without ethical consideration. These mesenchymal-like stem cells have high rate of proliferation and possess multi lineage differentiation potency. This study was undertaken to isolate the MenSCs and assess their potential in differentiation into epidermal lineage. METHODS About 5-10 ml of menstrual blood (MB) was collected using sterile Diva cups inserted into vagina during menstruation from volunteered healthy fertile women aged between 22-30 years. MB was transferred into Falcon tubes containing phosphate buffered saline (PBS) without Ca2+ or Mg2+ supplemented with 2.5 µg/ml fungizone, 100 µg/mL streptomycin, 100 U/mL penicillin and 0.5 mM EDTA. Mononuclear cells were separated using Ficoll-Hypaque density gradient centrifugation and washed out in PBS. The cell pellet was suspended in DMEM-F12 medium supplemented with 10% FBS and cultured in tissue culture plates. The isolated cells were co-cultured with keratinocytes derived from the foreskin of healthy newborn male aged 2-10 months who was a candidate for circumcision for differentiation into epidermal lineage. RESULTS The isolated MenSCs were adhered to the plate and exhibited spindle-shaped morphology. Flow cytometric analysis revealed the expression of mesenchymal markers of CD10, CD29, CD73, and CD105 and lack of hematopoietic stem cells markers. An early success in derivation of epidermal lineage from MenSCs was visible. CONCLUSION The MenSCs are a real source to design differentiation to epidermal cells that can be used non-invasively in various dermatological lesions and diseases. PMID:27308237

  17. Lineage Reprogramming of Fibroblasts into Proliferative Induced Cardiac Progenitor Cells by Defined Factors.

    PubMed

    Lalit, Pratik A; Salick, Max R; Nelson, Daryl O; Squirrell, Jayne M; Shafer, Christina M; Patel, Neel G; Saeed, Imaan; Schmuck, Eric G; Markandeya, Yogananda S; Wong, Rachel; Lea, Martin R; Eliceiri, Kevin W; Hacker, Timothy A; Crone, Wendy C; Kyba, Michael; Garry, Daniel J; Stewart, Ron; Thomson, James A; Downs, Karen M; Lyons, Gary E; Kamp, Timothy J

    2016-03-01

    Several studies have reported reprogramming of fibroblasts into induced cardiomyocytes; however, reprogramming into proliferative induced cardiac progenitor cells (iCPCs) remains to be accomplished. Here we report that a combination of 11 or 5 cardiac factors along with canonical Wnt and JAK/STAT signaling reprogrammed adult mouse cardiac, lung, and tail tip fibroblasts into iCPCs. The iCPCs were cardiac mesoderm-restricted progenitors that could be expanded extensively while maintaining multipotency to differentiate into cardiomyocytes, smooth muscle cells, and endothelial cells in vitro. Moreover, iCPCs injected into the cardiac crescent of mouse embryos differentiated into cardiomyocytes. iCPCs transplanted into the post-myocardial infarction mouse heart improved survival and differentiated into cardiomyocytes, smooth muscle cells, and endothelial cells. Lineage reprogramming of adult somatic cells into iCPCs provides a scalable cell source for drug discovery, disease modeling, and cardiac regenerative therapy. PMID:26877223

  18. Endothelial Outgrowth Cells: Function and Performance in Vascular Grafts

    PubMed Central

    Glynn, Jeremy J.

    2014-01-01

    The clinical need for vascular grafts continues to grow. Tissue engineering strategies have been employed to develop vascular grafts for patients lacking sufficient autologous vessels for grafting. Restoring a functional endothelium on the graft lumen has been shown to greatly improve the long-term patency of small-diameter grafts. However, obtaining an autologous source of endothelial cells for in vitro endothelialization is invasive and often not a viable option. Endothelial outgrowth cells (EOCs), derived from circulating progenitor cells in peripheral blood, provide an alternative cell source for engineering an autologous endothelium. This review aims at highlighting the role of EOCs in the regulation of processes that are central to vascular graft performance. To characterize EOC performance in vascular grafts, this review identifies the characteristics of EOCs, defines functional performance criteria for EOCs in vascular grafts, and summarizes the existing work in developing vascular grafts with EOCs. PMID:24004404

  19. Endothelial Dysfunction in Idiopathic Sudden Sensorineural Hearing Loss: A Review.

    PubMed

    Quaranta, Nicola; De Ceglie, Vincenzo; D'Elia, Alessandra

    2016-04-20

    An endothelial dysfunction has been described in idiopathic sudden sensorineural hearing loss (ISSHL) patients. The purpose of our review was to: i) identify, evaluate and review recent research about cardiovascular risk factors involvement and signs of endothelial dysfunction in ISSHL; ii) implication of these discovering in clinical practice and future research. A Medline literature search was conducted to identify any study on the involvement of endothelial dysfunction in ISSHL, published in the English language in the last decade. The following MEDLINE search terms were used: sudden sensorineural hearing loss (SSHL) and endothelial dysfunction (text words). Additional studies were identified by hand searching the references of original articles and review articles. Studies were not excluded on the basis of the qualitative or quantitative definitions of SSHL, treatment regimens, or outcome measures. Data were extracted from included papers by a reviewer. Information on the patients, investigations, methods, interventions, and outcomes were systematically analyzed. Characteristics and results of all included studies were reviewed systematically. High levels of adhesion molecules, hyperhomocysteinemia and lower folate levels, unbalanced oxidative status, a lower value of flow-mediated dilatation of brachial artery and a reduced percentage of circulating endothelial progenitor cells in patients affected by ISSHL support the hypothesis that this syndrome should be considered as a microcirculation disorder based on endothelial dysfunction and drive clinicians to implement all the traditional strategies used for preventing cardiovascular events, to also reduce the likelihood of ISSHL occurrence. PMID:27588164

  20. Endothelial Dysfunction in Idiopathic Sudden Sensorineural Hearing Loss: A Review

    PubMed Central

    Quaranta, Nicola; De Ceglie, Vincenzo; D’Elia, Alessandra

    2016-01-01

    An endothelial dysfunction has been described in idiopathic sudden sensorineural hearing loss (ISSHL) patients. The purpose of our review was to: i) identify, evaluate and review recent research about cardiovascular risk factors involvement and signs of endothelial dysfunction in ISSHL; ii) implication of these discovering in clinical practice and future research. A Medline literature search was conducted to identify any study on the involvement of endothelial dysfunction in ISSHL, published in the English language in the last decade. The following MEDLINE search terms were used: sudden sensorineural hearing loss (SSHL) and endothelial dysfunction (text words). Additional studies were identified by hand searching the references of original articles and review articles. Studies were not excluded on the basis of the qualitative or quantitative definitions of SSHL, treatment regimens, or outcome measures. Data were extracted from included papers by a reviewer. Information on the patients, investigations, methods, interventions, and outcomes were systematically analyzed. Characteristics and results of all included studies were reviewed systematically. High levels of adhesion molecules, hyperhomocysteinemia and lower folate levels, unbalanced oxidative status, a lower value of flow-mediated dilatation of brachial artery and a reduced percentage of circulating endothelial progenitor cells in patients affected by ISSHL support the hypothesis that this syndrome should be considered as a microcirculation disorder based on endothelial dysfunction and drive clinicians to implement all the traditional strategies used for preventing cardiovascular events, to also reduce the likelihood of ISSHL occurrence. PMID:27588164

  1. Embryonic Heart Progenitors and Cardiogenesis

    PubMed Central

    Brade, Thomas; Pane, Luna S.; Moretti, Alessandra; Chien, Kenneth R.; Laugwitz, Karl-Ludwig

    2013-01-01

    The mammalian heart is a highly specialized organ, comprised of many different cell types arising from distinct embryonic progenitor populations during cardiogenesis. Three precursor populations have been identified to contribute to different myocytic and nonmyocytic cell lineages of the heart: cardiogenic mesoderm cells (CMC), the proepicardium (PE), and cardiac neural crest cells (CNCCs). This review will focus on molecular cues necessary for proper induction, expansion, and lineage-specific differentiation of these progenitor populations during cardiac development in vivo. Moreover, we will briefly discuss how the knowledge gained on embryonic heart progenitor biology can be used to develop novel therapeutic strategies for the management of congenital heart disease as well as for improvement of cardiac function in ischemic heart disease. PMID:24086063

  2. The Progenitors of Thermonuclear Supernovae

    SciTech Connect

    Piersanti, L.; Straniero, O.; Tornambe, A.; Dominguez, I.

    2009-05-03

    In the framework of the rotating Double Degenerate Scenario for type Ia Supernovae progenitors, we show that the dichotomy between explosive events in early and late type galaxies can be easily explained. Assuming that more massive progenitors produce slow-decline (high-luminosity) light curve, it comes out that, at the current age of the Universe, in late type galaxies the continuous star formation provides very massive exploding objects (prompt component) corresponding to slow-decline (bright) SNe; on the other hand, in early type galaxies, where star formation ended many billions years ago, only low mass ''normal luminosity'' objects (delayed component) are present.

  3. Erythrocyte Enrichment in Hematopoietic Progenitor Cell Cultures Based on Magnetic Susceptibility of the Hemoglobin

    PubMed Central

    Jin, Xiaoxia; Abbot, Stewart; Zhang, Xiaokui; Kang, Lin; Voskinarian-Berse, Vanessa; Zhao, Rui; Kameneva, Marina V.; Moore, Lee R.; Chalmers, Jeffrey J.; Zborowski, Maciej

    2012-01-01

    Using novel media formulations, it has been demonstrated that human placenta and umbilical cord blood-derived CD34+ cells can be expanded and differentiated into erythroid cells with high efficiency. However, obtaining mature and functional erythrocytes from the immature cell cultures with high purity and in an efficient manner remains a significant challenge. A distinguishing feature of a reticulocyte and maturing erythrocyte is the increasing concentration of hemoglobin and decreasing cell volume that results in increased cell magnetophoretic mobility (MM) when exposed to high magnetic fields and gradients, under anoxic conditions. Taking advantage of these initial observations, we studied a noninvasive (label-free) magnetic separation and analysis process to enrich and identify cultured functional erythrocytes. In addition to the magnetic cell separation and cell motion analysis in the magnetic field, the cell cultures were characterized for cell sedimentation rate, cell volume distributions using differential interference microscopy, immunophenotyping (glycophorin A), hemoglobin concentration and shear-induced deformability (elongation index, EI, by ektacytometry) to test for mature erythrocyte attributes. A commercial, packed column high-gradient magnetic separator (HGMS) was used for magnetic separation. The magnetically enriched fraction comprised 80% of the maturing cells (predominantly reticulocytes) that showed near 70% overlap of EI with the reference cord blood-derived RBC and over 50% overlap with the adult donor RBCs. The results demonstrate feasibility of label-free magnetic enrichment of erythrocyte fraction of CD34+ progenitor-derived cultures based on the presence of paramagnetic hemoglobin in the maturing erythrocytes. PMID:22952572

  4. Strategies and Techniques to Enhance the In Situ Endothelialization of Small-Diameter Biodegradable Polymeric Vascular Grafts

    PubMed Central

    Hibino, Narutoshi; Fisher, John P.

    2013-01-01

    Due to the lack of success in small-diameter (<6 mm) prosthetic vascular grafts, a variety of strategies have evolved utilizing a tissue-engineering approach. Much of this work has focused on enhancing the endothelialization of these grafts. A healthy, confluent endothelial layer provides dynamic control over homeo-stasis, influencing and preventing thrombosis and smooth muscle cell proliferation that can lead to intimal hyperplasia. Strategies to improve endothelialization of biodegradable polymeric grafts have encompassed both chemical and physical modifications to graft surfaces, many focusing on the recruitment of endothelial and endothelial progenitor cells. This review aims to provide a compilation of current and developing strategies that utilize in situ endothelialization to improve vascular graft outcomes, providing a context for the future directions of vascular tissue-engineering strategies that do not require preprocedural cell seeding. PMID:23252992

  5. Progenitor Cells and Podocyte Regeneration

    PubMed Central

    Shankland, Stuart J.; Pippin, Jeffrey W.; Duffield, Jeremy S.

    2014-01-01

    The very limited ability of adult podocytes to proliferate in vivo is clinically significant because: podocytes form a vascular barrier which is functionally critical to the nephron; podocyte hypoplasia is a characteristic of disease; and inadequate regeneration of podocytes is a major cause of persistent podocyte hypoplasia. Excessive podocyte loss or inadequate replacement leads to glomerulosclerosis in many progressive kidney diseases. Thus, restoration of podocyte cell density is almost certainly reliant on regeneration by podocyte progenitors. However such putative progenitors have remained elusive until recently. In this review we describe the developmental processes leading to podocyte and parietal epithelial cell (PEC) formation during glomerulogenesis. We compare evidence that in normal human kidneys PECs expressing ‘progenitor’ markers CD133 and CD24 can differentiate into podocytes in vitro and in vivo with evidence from animal models suggesting a more limited role of PEC-capacity to serve as podocyte progenitors in adults. We will highlight tantalizing new evidence that specialized vascular wall cells of afferent arterioles including those which produce renin in healthy kidney, provide a novel local progenitor source of new PECs and podocytes in response to podocyte hypoplasia in the adult, and draw comparisons with glomerulogenesis. PMID:25217270

  6. Concise Review: Evidence for CD34 as a Common Marker for Diverse Progenitors

    PubMed Central

    Sidney, Laura E; Branch, Matthew J; Dunphy, Siobhán E; Dua, Harminder S; Hopkinson, Andrew

    2014-01-01

    CD34 is a transmembrane phosphoglycoprotein, first identified on hematopoietic stem and progenitor cells. Clinically, it is associated with the selection and enrichment of hematopoietic stem cells for bone marrow transplants. Due to these historical and clinical associations, CD34 expression is almost ubiquitously related to hematopoietic cells, and it is a common misconception that CD34-positive (CD34+) cells in nonhematopoietic samples represent hematopoietic contamination. The prevailing school of thought states that multipotent mesenchymal stromal cells (MSC) do not express CD34. However, strong evidence demonstrates CD34 is expressed not only by MSC but by a multitude of other nonhematopoietic cell types including muscle satellite cells, corneal keratocytes, interstitial cells, epithelial progenitors, and vascular endothelial progenitors. In many cases, the CD34+ cells represent a small proportion of the total cell population and also indicate a distinct subset of cells with enhanced progenitor activity. Herein, we explore common traits between cells that express CD34, including associated markers, morphology and differentiation potential. We endeavor to highlight key similarities between CD34+ cells, with a focus on progenitor activity. A common function of CD34 has yet to be elucidated, but by analyzing and understanding links between CD34+ cells, we hope to be able to offer an insight into the overlapping properties of cells that express CD34. Stem Cells 2014;32:1380–1389 PMID:24497003

  7. Using cultured endothelial cells to study endothelial barrier dysfunction: Challenges and opportunities.

    PubMed

    Aman, Jurjan; Weijers, Ester M; van Nieuw Amerongen, Geerten P; Malik, Asrar B; van Hinsbergh, Victor W M

    2016-08-01

    Despite considerable progress in the understanding of endothelial barrier regulation and the identification of approaches that have the potential to improve endothelial barrier function, no drug- or stem cell-based therapy is presently available to reverse the widespread vascular leak that is observed in acute respiratory distress syndrome (ARDS) and sepsis. The translational gap suggests a need to develop experimental approaches and tools that better mimic the complex environment of the microcirculation in which the vascular leak develops. Recent studies have identified several elements of this microenvironment. Among these are composition and stiffness of the extracellular matrix, fluid shear stress, interaction of endothelial cells (ECs) with pericytes, oxygen tension, and the combination of toxic and mechanic injurious stimuli. Development of novel cell culture techniques that integrate these elements would allow in-depth analysis of EC biology that closely approaches the (patho)physiological conditions in situ. In parallel, techniques to isolate organ-specific ECs, to define EC heterogeneity in its full complexity, and to culture patient-derived ECs from inducible pluripotent stem cells or endothelial progenitor cells are likely to advance the